Microorganisms

14 pages, 2035 KB

Open AccessEditor’s ChoiceArticle

Evaluation of a Commercial Multiplex Real-Time PCR with Melting Curve Analysis for the Detection of Mycobacterium tuberculosis Complex and Five Nontuberculous Mycobacterial Species

by Keun Ju Kim, Yunhee Chang, Seung Gyu Yun, Myung-Hyun Nam and Yunjung Cho

Microorganisms 2025, 13(1), 26; https://doi.org/10.3390/microorganisms13010026 - 26 Dec 2024

Cited by 3 | Viewed by 3239

Abstract

Background: Accurate and timely diagnosis of mycobacterial infections, including Mycobacterium tuberculosis complex (MTBC) and nontuberculous mycobacteria (NTM), is crucial for effective disease management. Methods: This study evaluated the performance of the NeoPlex TB/NTM-5 Detection Kit (NeoPlex assay, Seongnam, Republic of Korea), a multiplex [...] Read more.

Background: Accurate and timely diagnosis of mycobacterial infections, including Mycobacterium tuberculosis complex (MTBC) and nontuberculous mycobacteria (NTM), is crucial for effective disease management. Methods: This study evaluated the performance of the NeoPlex TB/NTM-5 Detection Kit (NeoPlex assay, Seongnam, Republic of Korea), a multiplex real-time PCR assay that incorporates melting curve analysis, compared with the line-probe assay (LPA). The NeoPlex assay could simultaneously detect and differentiate MTBC from five other NTM species: Mycobacterium intracellulare, Mycobacterium avium, Mycobacterium kansasii, Mycobacterium abscessus, and Mycobacterium massiliense. A total of 91 acid-fast bacillus culture-positive samples, comprising 36 MTBC and 55 NTM isolates, were collected from the Korea University Anam Hospital. Results: The NeoPlex assay successfully detected nucleic acids in 87 of the 91 isolates (95.6%). Notably, it identified additional mycobacterial nucleic acids not detected by the LPA in eight isolates. These findings were confirmed via DNA sequencing. The assay had 100% sensitivity and specificity for M. intracellulare, M. abscessus, M. massilense, NTM, and MTBC, whereas it had 100% specificity and sensitivity of 90.9% and 75.0% for M. avium and M. kansasii, respectively. Conclusions: These results highlight the potential of the NeoPlex assay to enhance rapid and accurate diagnosis of mycobacterial infections, particularly in settings in which prompt treatment initiation is essential. Full article

(This article belongs to the Special Issue Detection and Analysis of Clinical Microbial Infections, Second Edition)

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14 pages, 2051 KB

Open AccessEditor’s ChoiceArticle

Facultatively Anaerobic Staphylococci Enable Anaerobic Cutibacterium Species to Grow and Form Biofilms Under Aerobic Conditions

by Jeffrey B. Kaplan, Michael Assa, Noor Mruwat, Miloslav Sailer, Suresh Regmi and Khalaf Kridin

Microorganisms 2024, 12(12), 2601; https://doi.org/10.3390/microorganisms12122601 - 16 Dec 2024

Cited by 3 | Viewed by 4052

Abstract

Facultatively anaerobic Staphylococcus spp. and anaerobic Cutibacterium spp. are among the most prominent bacteria on human skin. Although skin microbes generally grow as multispecies biofilms, few studies have investigated the interaction between staphylococci and Cutibacterium spp. in dual-species biofilms. Here, we measured the [...] Read more.

Facultatively anaerobic Staphylococcus spp. and anaerobic Cutibacterium spp. are among the most prominent bacteria on human skin. Although skin microbes generally grow as multispecies biofilms, few studies have investigated the interaction between staphylococci and Cutibacterium spp. in dual-species biofilms. Here, we measured the mono- and dual-species biofilm formation of four staphylococcal species (S. epidermidis, S. hominis, S. capitis, and S. aureus) and two Cutibacterium spp. (C. acnes and C. avidum) cultured in vitro under both aerobic and anaerobic conditions. The biofilms were quantitated by rinsing them to remove planktonic cells, detaching the biofilm bacteria via sonication, and enumerating the cells by dilution plating. When cultured alone, staphylococci formed biofilms under both aerobic and anaerobic conditions, whereas Cutibacterium spp. formed biofilms only under anaerobic conditions. In co-culture, staphylococcal biofilm formation was unaffected by the presence of Cutibacterium spp., regardless of oxygen availability. However, Cutibacterium spp. biofilm formation was significantly enhanced in the presence of staphylococci, enabling robust growth under both anaerobic and aerobic conditions. Fluorescence confocal microscopy of the aerobic dual-species biofilms suggested that staphylococci create anaerobic niches at the base of the biofilm where C. acnes can grow. These findings demonstrate that staphylococci facilitate the colonization of Cutibacterium spp. in oxygen-rich environments, potentially explaining their presence in high numbers on the oxygen-exposed stratum corneum. Full article

(This article belongs to the Special Issue Advanced Research on Microbial Biofilms—a Themed Issue in Honor of Professor Gianfranco Donelli)

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18 pages, 2351 KB

Open AccessEditor’s ChoiceArticle

Bacterial Diversity in the Different Ecological Niches Related to the Yonghwasil Pond (Republic of Korea)

by Myung Kyum Kim, Bong-Soon Lim, Chang Seok Lee and Sathiyaraj Srinivasan

Microorganisms 2024, 12(12), 2547; https://doi.org/10.3390/microorganisms12122547 - 11 Dec 2024

Cited by 2 | Viewed by 2200

Abstract

The bacteriome profile was studied in freshwater ecosystems within the Yonghwasil pond, situated at the National Institute of Ecology, Seocheon-gun, Chungcheongnam-do, central western Korea. Six samples from water, mud, and soil niches were assessed, specifically from lake water, bottom mud (sediment), and root-soil [...] Read more.

The bacteriome profile was studied in freshwater ecosystems within the Yonghwasil pond, situated at the National Institute of Ecology, Seocheon-gun, Chungcheongnam-do, central western Korea. Six samples from water, mud, and soil niches were assessed, specifically from lake water, bottom mud (sediment), and root-soil samples of Bulrush, wild rice, Reed, and Korean Willow. Notably, the phylum Actinobacteria exhibited an upward trend moving from water to mud to soil samples, whereas Chloroflexi showed a contrasting decrease. Across the board, Proteobacteria emerged as the reigning phylum, and subsequent dominance was attributed to Firmicutes and Actinobacteria. The water samples were characterized by an enriched presence of Cyanobacteria and Bacteroidetes, whereas the mud samples distinctly housed a higher concentration of Chloroflexi. Assessing biodiversity through OTU and ACE indices revealed a subdued species richness in the water samples. On the contrary, mud samples stood out with the highest OTU and ACE metrics, signifying a microbially diverse habitat. Bulrush, wild rice, Reed, and Willow samples showed intermediate microbial diversity. The Shannon index further corroborated the pronounced microbial diversity in mud and Bulrush habitats with others. This research elucidates the microbial intricacies across different habitats within Yonghwasil Pond, emphasizing the pivotal role of environmental matrices in shaping bacterial communities. Full article

(This article belongs to the Special Issue Plants and Microorganisms: Role of Microorganisms in Monitoring and Improving Environmental Health)

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22 pages, 4826 KB

Open AccessEditor’s ChoiceArticle

Geography, Antimicrobial Resistance, and Genomics of Salmonella enterica (Serotypes Newport and Anatum) from Meat in Mexico (2021–2023)

by Eduardo Canek Reynoso, Enrique Jesús Delgado-Suárez, Cindy Fabiola Hernández-Pérez, Yaselda Chavarin-Pineda, Elizabeth Ernestina Godoy-Lozano, Geny Fierros-Zárate, Omar Alejandro Aguilar-Vera, Santiago Castillo-Ramírez, Luz del Carmen Sierra Gómez-Pedroso and Luisa María Sánchez-Zamorano

Microorganisms 2024, 12(12), 2485; https://doi.org/10.3390/microorganisms12122485 - 3 Dec 2024

Cited by 3 | Viewed by 3402

Abstract

Salmonella enterica non-typhoidal is a major contributor to diarrheal diseases, with over 2600 serovars identified across diverse environments. In Mexico, serovars Newport and Anatum have shown a marked increase, especially in foodborne disease, posing a public health problem. We conducted a cross-sectional study from [...] Read more.

Salmonella enterica non-typhoidal is a major contributor to diarrheal diseases, with over 2600 serovars identified across diverse environments. In Mexico, serovars Newport and Anatum have shown a marked increase, especially in foodborne disease, posing a public health problem. We conducted a cross-sectional study from 2021 to 2023 using active epidemiological surveillance to assess contamination in ground beef and pork at butcher shops nationwide. It involved isolation, phenotypic antimicrobial resistance, comparative genomics, spatial distribution, antimicrobial-resistance genes, and pangenome analysis. A total of 402 non-typhoidal S. enterica strains were isolated, including 59 Newport and 50 Anatum. After curating for redundancy, 45 Newport and 32 Anatum strains remained. We found that 75% of Newport strains exhibited multidrug resistance (MDR), compared to 25% of Anatum strains. Salmonella Newport also showed a broader distribution and stronger antibiotic-resistance capacity, particularly due to genes such as mphA and ramA. Our pangenome analysis showed a predominance of cell maintenance and survival-process genes in the accessory genome of both serotypes. Considering unique genes, Salmonella Anatum and Newport showed a notorious abundance of genes with functions related to replication, recombination, and repair. The substantial rise of Anatum and Newport strains in meat samples for human consumption presents an epidemiological alert, highlighting the critical need for stringent surveillance programs to mitigate human and ecosystem health risks. Full article

(This article belongs to the Special Issue Molecular Epidemiology of Antimicrobial Resistance, 2nd Edition)

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17 pages, 4373 KB

Open AccessEditor’s ChoiceArticle

Effective Inhibitor Removal from Wastewater Samples Increases Sensitivity of RT-dPCR and Sequencing Analyses and Enhances the Stability of Wastewater-Based Surveillance

by Nico Linzner, Alexander Bartel, Vera Schumacher, José Horacio Grau, Emanuel Wyler, Henrike Preuß, Sonja Garske, Julia Bitzegeio, Elisabeth Barbara Kirst, Karsten Liere, Sebastian Hoppe, Tatiana A. Borodina, Janine Altmüller, Markus Landthaler, Martin Meixner, Daniel Sagebiel and Uta Böckelmann

Microorganisms 2024, 12(12), 2475; https://doi.org/10.3390/microorganisms12122475 - 2 Dec 2024

Cited by 6 | Viewed by 6551

Abstract

Wastewater-based surveillance (WBS) is a proven tool for monitoring population-level infection events. Wastewater contains high concentrations of inhibitors, which contaminate the total nucleic acids (TNA) extracted from these samples. We found that TNA extracts from raw influent of Berlin wastewater treatment plants contained [...] Read more.

Wastewater-based surveillance (WBS) is a proven tool for monitoring population-level infection events. Wastewater contains high concentrations of inhibitors, which contaminate the total nucleic acids (TNA) extracted from these samples. We found that TNA extracts from raw influent of Berlin wastewater treatment plants contained highly variable amounts of inhibitors that impaired molecular analyses like dPCR and next-generation sequencing (NGS). By using dilutions, we were able to detect inhibitory effects. To enhance WBS sensitivity and stability, we applied a combination of PCR inhibitor removal and TNA dilution (PIR+D). This approach led to a 26-fold increase in measured SARS-CoV-2 concentrations, practically reducing the detection limit. Additionally, we observed a substantial increase in the stability of the time series. We define suitable stability as a mean absolute error (MAE) below 0.1 log₁₀ copies/L and a geometric mean relative absolute error (GMRAE) below 26%. Using PIR+D, the MAE could be reduced from 0.219 to 0.097 and the GMRAE from 65.5% to 26.0%, and even further in real-world WBS. Furthermore, PIR+D improved SARS-CoV-2 genome alignment and coverage in amplicon-based NGS for low to medium concentrations. In conclusion, we strongly recommend both the monitoring and removal of inhibitors from samples for WBS. Full article

(This article belongs to the Special Issue Surveillance of SARS-CoV-2 Employing Wastewater)

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14 pages, 5382 KB

Open AccessEditor’s ChoiceArticle

Rhizosphere Shifts: Reduced Fungal Diversity and Microbial Community Functionality Enhance Plant Adaptation in Continuous Cropping Systems

by Jichao Li, Yingmei Zuo and Jinyu Zhang

Microorganisms 2024, 12(12), 2420; https://doi.org/10.3390/microorganisms12122420 - 25 Nov 2024

Cited by 5 | Viewed by 1900

Abstract

Continuous cropping problems constitute threats to perennial plant health and survival. Soil conditioners have the potential to enhance plant disease resistance in continuous cropping systems. However, how microbes and metabolites of the rhizosphere respond to soil conditioner addition remains largely unknown, but this [...] Read more.

Continuous cropping problems constitute threats to perennial plant health and survival. Soil conditioners have the potential to enhance plant disease resistance in continuous cropping systems. However, how microbes and metabolites of the rhizosphere respond to soil conditioner addition remains largely unknown, but this knowledge is paramount to providing innovative strategies to enhance plant adaptation in continuous cropping systems. Here, we found that a biochar conditioner significantly improved plant survival rates in a continuous cropping system. The biochar-induced rhizosphere significantly alters the fungal community, causing a decline in fungal diversity and the downregulation of soil microbial community functionality. Specifically, the biochar-induced rhizosphere causes a reduction in the relative abundance of pathogenic Fusarium sp. and phenolic acid concentration, whose variations are the primary causes of continuous cropping problems. Conversely, we observed an unexpected bacterial diversity increase in rhizospheric and non-rhizospheric soils. Our research further identified key microbial taxa in the biochar-induced rhizosphere, namely, Monographella, Acremonium, Geosmithia, and Funneliformis, which enhance soil nutrient availability, suppress Fusarium sp., mitigate soil acidification, and reduce phenolic acid concentrations. Collectively, we highlight the critical role of regular microbial communities and metabolites in determining plant health during continuous cropping and propose a synthetic microbial community framework for further optimizing the ecological functions of the rhizosphere. Full article

(This article belongs to the Special Issue State-of-the-Art Environmental Microbiology in China (2023–2024))

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12 pages, 1398 KB

Open AccessEditor’s ChoiceArticle

Wound Gel Formulations Containing Poloxamer 407 and Polyhexanide Have In Vitro Antimicrobial and Antibiofilm Activity Against Wound-Associated Microbial Pathogens

by Jeyachchandran Visvalingam, Nandadeva Yakandawala, Suresh Regmi, Adetola Adeniji, Parveen Sharma and Miloslav Sailer

Microorganisms 2024, 12(11), 2362; https://doi.org/10.3390/microorganisms12112362 - 19 Nov 2024

Cited by 4 | Viewed by 2864

Abstract

Chronic wounds are often caused or exacerbated by microbial biofilms that are highly resistant to antimicrobial treatments and that prevent healing. This study compared the antimicrobial and antibiofilm activity of nine topical wound treatments, comprising gels with different concentrations of poloxamer 407 (20–26%) [...] Read more.

Chronic wounds are often caused or exacerbated by microbial biofilms that are highly resistant to antimicrobial treatments and that prevent healing. This study compared the antimicrobial and antibiofilm activity of nine topical wound treatments, comprising gels with different concentrations of poloxamer 407 (20–26%) and different pH levels (4–6) and containing polyhexanide (PHMB) as an antimicrobial agent; the effects of pH on wound gels containing this agent have not been previously reported. The wound gel formulations were tested against six common wound-associated microbial pathogens: Staphylococcus aureus, S. epidermidis, Pseudomonas aeruginosa, Escherichia coli, Acinetobacter baumannii, and Candida albicans. Time-kill assays were used to assess antimicrobial activity against planktonic forms of each species, and a colony biofilm model was used to assess antibiofilm activity against existing biofilms as well as inhibition of new biofilm formation. Biofilm inhibition activity was also assessed in the presence of common wound dressing materials. Wound gels with higher pH levels exhibited stronger antimicrobial activity, while poloxamer 407 concentrations >20% negatively impacted antimicrobial activity. Wound gel formulations were identified that had antimicrobial, antibiofilm, and biofilm inhibition activity against all tested species in vitro. Biofilm inhibition activity was not affected by contact with common wound dressings. Further development of these wound gels may provide a valuable new option for the treatment and prevention of chronic wounds. Full article

(This article belongs to the Special Issue Pathogenic Biofilms: Physiology, Molecular Mechanisms and Counter Strategies)

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10 pages, 1665 KB

Open AccessEditor’s ChoiceArticle

Ten-Year Antimicrobial Resistance Trend in Uropathogenic Escherichia coli (UPEC) Isolated from Dogs and Cats Admitted to a Veterinary Teaching Hospital in Italy

by Alessandro Bellato, Patrizia Robino, Maria Cristina Stella, Daniela Scalas, Paolo Savarino, Renato Zanatta, Giovanni Re and Patrizia Nebbia

Microorganisms 2024, 12(11), 2175; https://doi.org/10.3390/microorganisms12112175 - 29 Oct 2024

Cited by 2 | Viewed by 2955

Abstract

Urinary tract infections (UTIs) are a common occurrence in cats and dogs. Surveillance of antibiotic resistance trends helps in the prudent selection of suitable antimicrobial agents. However, there are limited available data on this matter in Italy. This retrospective study aimed to investigate [...] Read more.

Urinary tract infections (UTIs) are a common occurrence in cats and dogs. Surveillance of antibiotic resistance trends helps in the prudent selection of suitable antimicrobial agents. However, there are limited available data on this matter in Italy. This retrospective study aimed to investigate the trends of antimicrobial resistance in uropathogenic Escherichia coli (UPEC) isolated from cats and dogs over ten years (January 2014 to October 2023). Three hundred thirty-nine UPEC strains were isolated from urine samples submitted to the Veterinary Teaching Hospital of Torino (Italy). Antimicrobial susceptibility testing was conducted for up to 11 classes of antibacterials, categorized into four categories (A, B, C, and D) following the European Medicine Agency guidelines for prudent antimicrobial use in animals. The results reveal a higher resistance towards compounds in categories C and D, while fewer isolates were resistant to antibacterials in categories B and A. Resistance has steadily increased from 2014 to 2019. Starting from 2020, a decline in resistance is evident in all four categories. The reduction is more pronounced for the categories subject to the greatest restrictions under European and national legislation. The change in resistance trend is in line with findings from other European countries and food-production animals in Italy. Full article

(This article belongs to the Special Issue Recent Research on Antimicrobial Stewardship)

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16 pages, 3063 KB

Open AccessEditor’s ChoiceArticle

Aging-Induced Changes in Cutibacterium acnes and Their Effects on Skin Elasticity and Wrinkle Formation

by YeonGyun Jung, Ikwhan Kim, Da-Ryung Jung, Ji Hoon Ha, Eun Kyung Lee, Jin Mo Kim, Jin Young Kim, Jun-Hwan Jang, Jun-Tae Bae, Jae-Ho Shin and Yoon Soo Cho

Microorganisms 2024, 12(11), 2179; https://doi.org/10.3390/microorganisms12112179 - 29 Oct 2024

Cited by 4 | Viewed by 4138

Abstract

Skin aging involves biomechanical changes like decreased elasticity, increased wrinkle formation, and altered barrier function. The skin microbiome significantly impacts this process. Here, we investigated the effects of decreased Cutibacterium acnes abundance and increase in other skin microorganisms on skin biomechanical properties in [...] Read more.

Skin aging involves biomechanical changes like decreased elasticity, increased wrinkle formation, and altered barrier function. The skin microbiome significantly impacts this process. Here, we investigated the effects of decreased Cutibacterium acnes abundance and increase in other skin microorganisms on skin biomechanical properties in 60 healthy Koreans from Seoul, divided into younger (20–29 years) and older (60–75 years) groups. Metagenomic sequencing and skin assessments showed that the older group exhibited decreased C. acnes dominance and increased microbial diversity, correlating with reduced skin elasticity and increased wrinkles. In the younger age group, the enriched pathways included zeatin biosynthesis, distinct biotin metabolism pathways, and cofactor and vitamin metabolism in the younger age group, whereas pathways related to lipid metabolism, energy metabolism, and responses to environmental stressors, including UV damage and pollution, were enriched in the older group, according to functional analysis results. Network analysis indicated higher microbial connectivity in the younger group, suggesting a more stable community, whereas the older group’s community displayed higher modularity, indicating more independent and specialized clusters. This study enhances our understanding of the impact of skin microbiome changes on skin aging, particularly the anti-aging effects of C. acnes. Future research should focus on the physiological mechanisms of skin microbiota on skin aging and explore therapeutic potentials to enhance skin health. Full article

(This article belongs to the Special Issue Advances in Human Microbiomes)

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14 pages, 2125 KB

Open AccessEditor’s ChoiceArticle

Genomic Analysis of Antimicrobial Resistance in Pseudomonas aeruginosa from a “One Health” Perspective

by Celia García-Rivera, Carmen Molina-Pardines, José M. Haro-Moreno, Mónica Parra Grande, Juan Carlos Rodríguez and Mario López-Pérez

Microorganisms 2024, 12(9), 1770; https://doi.org/10.3390/microorganisms12091770 - 27 Aug 2024

Cited by 4 | Viewed by 2466

Abstract

The “One Health” approach provides a comprehensive framework for understanding antimicrobial resistance. This perspective is of particular importance in the study of Pseudomonas aeruginosa, as it is not only a pathogen that affects humans but also persists in environmental reservoirs. To assess [...] Read more.

The “One Health” approach provides a comprehensive framework for understanding antimicrobial resistance. This perspective is of particular importance in the study of Pseudomonas aeruginosa, as it is not only a pathogen that affects humans but also persists in environmental reservoirs. To assess evolutionary selection for niche-specific traits, a genomic comparison of 749 P. aeruginosa strains from three environments (clinical, aquatic, and soil) was performed. The results showed that the environment does indeed exert selective pressure on specific traits. The high percentage of persistent genome, the lack of correlation between phylogeny and origin of the isolate, and the high intrinsic resistance indicate that the species has a high potential for pathogenicity and resistance, regardless of the reservoir. The flexible genome showed an enrichment of metal resistance genes, which could act as a co-selection of antibiotic resistance genes. In the plasmids, resistance genes were found in multigenic clusters, with the presence of a mobile integron being prominent. This integron was identified in several pathogenic strains belonging to distantly related taxa with a worldwide distribution, showing the risk of rapid evolution of resistance. These results provide a more complete understanding of the evolution of P. aeruginosa, which could help develop new prevention strategies. Full article

(This article belongs to the Section Antimicrobial Agents and Resistance)

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24 pages, 2645 KB

Open AccessEditor’s ChoiceArticle

The Diatom Diversity and Ecological Status of a Tufa-Depositing River through eDNA Metabarcoding vs. a Morphological Approach—A Case Study of the Una River (Bosnia and Herzegovina)

by Jasmina Kamberović, Marija Gligora Udovič, Antonija Kulaš, Kálmán Tapolczai, Sandi Orlić, Amela Jusufović, Almina Gajić, Petar Žutinić, Adisa Ahmić and Belma Kalamujić Stroil

Microorganisms 2024, 12(8), 1722; https://doi.org/10.3390/microorganisms12081722 - 21 Aug 2024

Cited by 6 | Viewed by 5953

Abstract

Tufa deposits in karst rivers are unique habitats created by mutual interactions between specific environmental and biotope features and inhabited by diatoms as a highly abundant and diverse algal group. This pilot study aimed to investigate the diversity of diatom communities on tufa [...] Read more.

Tufa deposits in karst rivers are unique habitats created by mutual interactions between specific environmental and biotope features and inhabited by diatoms as a highly abundant and diverse algal group. This pilot study aimed to investigate the diversity of diatom communities on tufa depositing habitats and assess the Una River’s ecological status using a comparative molecular and morphological approach for diatom identification. The 312 base pairs of the rbcL gene were barcoded and analyzed using MiSeq reads and amplicon sequence variants (ASVs) obtained by the DADA2 pipeline. The reference database Diat.barcode v7 was used for taxonomic assignment. The morphological identification of the diatoms was carried out in parallel. In total, the combined dataset revealed 46 taxa identified at genus rank, 125 on the subgenus, and 145 on combined taxonomy rank. The metabarcoding approach mostly leads to a lower number of identified taxa at species rank (58 in molecular vs. 119 in optical inventory), resulting in higher values of beta diversity and heterogeneity in diatom assemblages in samples obtained by morphological approach. Despite the high percentage of taxonomically not assigned diatom ASVs to the species rank, high Shannon diversity index values and a similar number of taxa per locations compared to the morphological approach were obtained. Taxa Achnanthidium minutissimum (Kützing) Czarnecki, Achnanthidium pyrenaicum (Hustedt) H.Kobayasi, Amphora pediculus (Kützing) Grunow, Diatoma vulgaris Bory, Navicula cryptotenella Lange-Bertalot, and Navicula tripunctata (O.F.Müller) Bory were identified at all locations in both inventories. Although limited consistency in the diatom abundances between the two inventory datasets was found, a similar grouping of samples was observed connected to the river’s longitudinal gradient. The data obtained using molecular approach in most sites indicated a mostly lower ecological status (good or moderate) compared to the data obtained from the morphological approach (high, good, and moderate). The potential of environmental DNA (eDNA) diatom metabarcoding for water monitoring and diversity studies is undeniable, but to fully realize the benefits of these methods in the future, it is essential to standardize protocols and expand the reference database for species found in specific habitats, such as tufa deposits. Full article

(This article belongs to the Special Issue Molecular Ecology of Microalgae and Cyanobacteria)

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16 pages, 3751 KB

Open AccessEditor’s ChoiceArticle

Exploring the Role of the Environment as a Reservoir of Antimicrobial-Resistant Campylobacter: Insights from Wild Birds and Surface Waters

by Louise Hock, Cécile Walczak, Juliette Mosser, Catherine Ragimbeau and Henry-Michel Cauchie

Microorganisms 2024, 12(8), 1621; https://doi.org/10.3390/microorganisms12081621 - 8 Aug 2024

Cited by 3 | Viewed by 2239

Abstract

Antimicrobial resistance (AMR) is a growing global health challenge, compromising bacterial infection treatments and necessitating robust surveillance and mitigation strategies. The overuse of antimicrobials in humans and farm animals has made them hotspots for AMR. However, the spread of AMR genes in wildlife [...] Read more.

Antimicrobial resistance (AMR) is a growing global health challenge, compromising bacterial infection treatments and necessitating robust surveillance and mitigation strategies. The overuse of antimicrobials in humans and farm animals has made them hotspots for AMR. However, the spread of AMR genes in wildlife and the environment represents an additional challenge, turning these areas into new AMR hotspots. Among the AMR bacteria considered to be of high concern for public health, Campylobacter has been the leading cause of foodborne infections in the European Union since 2005. This study examines the prevalence of AMR genes and virulence factors in Campylobacter isolates from wild birds and surface waters in Luxembourg. The findings reveal a significant prevalence of resistant Campylobacter strains, with 12% of C. jejuni from wild birds and 37% of C. coli from surface waters carrying resistance genes, mainly against key antibiotics like quinolones and tetracycline. This study underscores the crucial role of the environment in the spread of AMR bacteria and genes, highlighting the urgent need for enhanced surveillance and control measures to curb AMR in wildlife and environmental reservoirs and reduce transmission risks to humans. This research supports One Health approaches to tackling antimicrobial resistance and protecting human, animal, and environmental health. Full article

(This article belongs to the Special Issue Pathogen Infection in Wildlife 2.0)

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11 pages, 2961 KB

Open AccessEditor’s ChoiceArticle

A Bead Biofilm Reactor for High-Throughput Growth and Translational Applications

by Annika Gilmore, Marissa Badham, Winston Rudisin, Nicholas Ashton and Dustin Williams

Microorganisms 2024, 12(8), 1588; https://doi.org/10.3390/microorganisms12081588 - 5 Aug 2024

Cited by 3 | Viewed by 2444

Abstract

Bacteria in natural ecosystems such as soil, dirt, or debris preferentially reside in the biofilm phenotype. When a traumatic injury, such as an open fracture, occurs, these naturally dwelling biofilms and accompanying foreign material can contaminate the injury site. Given their high tolerance [...] Read more.

Bacteria in natural ecosystems such as soil, dirt, or debris preferentially reside in the biofilm phenotype. When a traumatic injury, such as an open fracture, occurs, these naturally dwelling biofilms and accompanying foreign material can contaminate the injury site. Given their high tolerance of systemic levels of antibiotics that may be administered prophylactically, biofilms may contribute to difficult-to-treat infections. In most animal models, planktonic bacteria are used as initial inocula to cause infection, and this might not accurately mimic clinically relevant contamination and infection scenarios. Further, few approaches and systems utilize the same biofilm and accompanying substrate throughout the experimental continuum. In this study, we designed a unique reactor to grow bacterial biofilms on up to 50 silica beads that modeled environmental wound contaminants. The data obtained indicated that the reactor system repeatably produced mature Staphylococcus aureus and Pseudomonas aeruginosa biofilms on the silica beads, with an average of 5.53 and 6.21 log₁₀ colony-forming units per mm², respectively. The bead substrates are easily manipulable for in vitro or in vivo applications, thus improving translatability. Taken together, the bead biofilm reactor presented herein may be a useful system for repeatably growing established biofilms on silica beads that could be used for susceptibility testing and as initial inocula in future animal models of trauma-related injuries. Full article

(This article belongs to the Special Issue Advance Research on Bacterial Biofilm)

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17 pages, 3909 KB

Open AccessEditor’s ChoiceArticle

Harnessing Machine Learning to Uncover Hidden Patterns in Azole-Resistant CYP51/ERG11 Proteins

by Otávio Guilherme Gonçalves de Almeida and Marcia Regina von Zeska Kress

Microorganisms 2024, 12(8), 1525; https://doi.org/10.3390/microorganisms12081525 - 25 Jul 2024

Cited by 2 | Viewed by 1861

Abstract

Fungal resistance is a public health concern due to the limited availability of antifungal resources and the complexities associated with treating persistent fungal infections. Azoles are thus far the primary line of defense against fungi. Specifically, azoles inhibit the conversion of lanosterol to [...] Read more.

Fungal resistance is a public health concern due to the limited availability of antifungal resources and the complexities associated with treating persistent fungal infections. Azoles are thus far the primary line of defense against fungi. Specifically, azoles inhibit the conversion of lanosterol to ergosterol, producing defective sterols and impairing fluidity in fungal plasmatic membranes. Studies on azole resistance have emphasized specific point mutations in CYP51/ERG11 proteins linked to resistance. Although very insightful, the traditional approach to studying azole resistance is time-consuming and prone to errors during meticulous alignment evaluation. It relies on a reference-based method using a specific protein sequence obtained from a wild-type (WT) phenotype. Therefore, this study introduces a machine learning (ML)-based approach utilizing molecular descriptors representing the physiochemical attributes of CYP51/ERG11 protein isoforms. This approach aims to unravel hidden patterns associated with azole resistance. The results highlight that descriptors related to amino acid composition and their combination of hydrophobicity and hydrophilicity effectively explain the slight differences between the resistant non-wild-type (NWT) and WT (nonresistant) protein sequences. This study underscores the potential of ML to unravel nuanced patterns in CYP51/ERG11 sequences, providing valuable molecular signatures that could inform future endeavors in drug development and computational screening of resistant and nonresistant fungal lineages. Full article

(This article belongs to the Special Issue Healthcare-Associated Infections and Antimicrobial Therapy)

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13 pages, 3691 KB

Open AccessEditor’s ChoiceArticle

Sensitive and Accurate Quantification of Enterovirus-D68 (EV-D68) Viral Loads Using Droplet Digital PCR (ddPCR)

by Cassandra S. Grizer, Zhaozhang Li and Joseph J. Mattapallil

Microorganisms 2024, 12(8), 1502; https://doi.org/10.3390/microorganisms12081502 - 23 Jul 2024

Cited by 3 | Viewed by 2773

Abstract

Enterovirus-D68 (EV-D68) is a reemerging virus that has been associated with numerous outbreaks in children in the past 10 years. Most assays examining viral infection kinetics have relied on the use of quantitative RT-PCR (qRT-PCR) assays as an assay of choice. Though valuable, [...] Read more.

Enterovirus-D68 (EV-D68) is a reemerging virus that has been associated with numerous outbreaks in children in the past 10 years. Most assays examining viral infection kinetics have relied on the use of quantitative RT-PCR (qRT-PCR) assays as an assay of choice. Though valuable, there are inherent limitations that introduce variability, thereby reducing its value when comparing results across the field. Unlike the qRT-PCR assay that uses a standard curve to determine the copy number of viral RNA, the droplet digital PCR assay (ddPCR) directly quantifies the absolute number of copies within a given sample, which in turn makes the assay highly sensitive and accurate. Here, we have developed an EV-D68-specific ddPCR assay that effectively quantifies EV-D68 RNA copies in both cells and supernatants within a dynamic range of 6.7 × 10⁻³ copies/μL to 1.2 × 10⁴ copies/μL of the sample. The assay was highly specific for a broad range of EV-D68 isolates (Fermon, US/MO/14-18947, US/MO/14-18949, US/KY/14-18953, USA/2018-23088, USA/2020-23336 and EV-D68-infected human nasal turbinate samples from the 2022 outbreak) without cross-reactivity to other viruses such as Enterovirus-A71 (EV-A71), Human Parechovirus (HPeV)-1 and -2, Coxsackievirus (CV)-B1, Human Coronavirus (HCoV)-NL63, SARS-CoV-2, Influenza-A and B, Rhinovirus, and Respiratory Syncytial Virus (RSV)-A2, which are known to cause infection in children. The assay was able to readily quantify EV-D68 in infected cells and supernatants along with nasal turbinate samples collected from children during the 2022 outbreak. Our results suggest that the assay can be readily translated to accurately quantify viral loads in tissues and body fluids such as plasma and lung or nasal aspirates. Full article

(This article belongs to the Section Virology)

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18 pages, 2280 KB

Open AccessEditor’s ChoiceArticle

Climate Change Stressors, Phosphate Limitation, and High Irradiation Interact to Increase Alexandrium minutum Toxicity and Modulate Encystment Rates

by Marta Sixto, Pilar Riobó, Francisco Rodríguez, Patricio A. Díaz and Rosa I. Figueroa

Microorganisms 2024, 12(7), 1480; https://doi.org/10.3390/microorganisms12071480 - 19 Jul 2024

Cited by 3 | Viewed by 2519

Abstract

The changes in the cell physiology (growth rate, cell size, and cell DNA content), photosynthetic efficiency, toxicity, and sexuality under variable light and nutrient (phosphates) conditions were evaluated in cultures of the dinoflagellate Alexandrium minutum obtained from a red tide in the Ría [...] Read more.

The changes in the cell physiology (growth rate, cell size, and cell DNA content), photosynthetic efficiency, toxicity, and sexuality under variable light and nutrient (phosphates) conditions were evaluated in cultures of the dinoflagellate Alexandrium minutum obtained from a red tide in the Ría de Vigo (NW Spain). The cells were grown at low (40 and 150 µE m⁻² s⁻¹), moderate (400 µE m⁻² s⁻¹), and high (800 µE m⁻² s⁻¹) light intensities in a medium with phosphate (P+) and without (P−). Cultures were acclimated to the irradiance conditions for one week, and the experiment was run for ~1 month. The cell size and DNA content were monitored via flow cytometry. Two different clonal strains were employed as a monoculture (in a P− or P+ medium) or, to foster sexuality and resting cyst formation, as a mixed culture (only in a P− medium). A. minutum growth was favored by increasing light intensities until 400 µE m⁻² s⁻¹. The DNA content analyses indicated the accumulation of S-phase cells at the highest light intensities (400 and 800 µE m⁻² s⁻¹) and therefore the negative effects on cell cycle progression. Only when the cells were grown in a P− medium did higher light intensities trigger dose-dependent, significantly higher toxicities in all the A. minutum cultures. This result suggests that the toxicity level is responsive to the combined effects of (high) light and (low) P stress. The cell size was not significantly affected by the light intensity or P conditions. The optimal light intensity for resting cyst formation was 150 µE m⁻² s⁻¹, with higher irradiances reducing the total encystment yield. Encystment was not observed at the lowest light intensity tested, indicative of the key role of low-level irradiance in gamete and/or zygote formation, in contrast to the stressor effect of excessive irradiance on planozygote formation and/or encystment. Full article

(This article belongs to the Special Issue Phytoplankton Ecology and Harmful Algal Bloom Surveillance in Marine Ecosystems)

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27 pages, 4308 KB

Open AccessEditor’s ChoiceArticle

The Microbiome of Fertilization-Stage Maize Silks (Style) Encodes Genes and Expresses Traits That Potentially Promote Survival in Pollen/Style Niches and Host Reproduction

by Michelle E. H. Thompson and Manish N. Raizada

Microorganisms 2024, 12(7), 1473; https://doi.org/10.3390/microorganisms12071473 - 19 Jul 2024

Cited by 5 | Viewed by 2970

Abstract

Within flowers, the style channel receives pollen and transmits male gametes inside elongating pollen tubes to ovules. The styles of maize/corn are called silks. Fertilization-stage silks possess complex microbiomes, which may partially derive from pollen. These microbiomes lack functional analysis. We hypothesize that [...] Read more.

Within flowers, the style channel receives pollen and transmits male gametes inside elongating pollen tubes to ovules. The styles of maize/corn are called silks. Fertilization-stage silks possess complex microbiomes, which may partially derive from pollen. These microbiomes lack functional analysis. We hypothesize that fertilization-stage silk microbiomes promote host fertilization to ensure their own vertical transmission. We further hypothesize that these microbes encode traits to survive stresses within the silk (water/nitrogen limitation) and pollen (dehydration/aluminum) habitats. Here, bacteria cultured from fertilization-stage silks of 14 North American maize genotypes underwent genome mining and functional testing, which revealed osmoprotection, nitrogen-fixation, and aluminum-tolerance traits. Bacteria contained auxin biosynthesis genes, and testing confirmed indole compound secretion, which is relevant, since pollen delivers auxin to silks to stimulate egg cell maturation. Some isolates encoded biosynthetic/transport compounds known to regulate pollen tube guidance/growth. The isolates encoded ACC deaminase, which degrades the precursor for ethylene that otherwise accelerates silk senescence. The findings suggest that members of the microbiome of fertilization-stage silks encode adaptations to survive the stress conditions of silk/pollen and have the potential to express signaling compounds known to impact reproduction. Overall, whereas these microbial traits have traditionally been assumed to primarily promote vegetative plant growth, this study proposes they may also play selfish roles during host reproduction. Full article

(This article belongs to the Section Plant Microbe Interactions)

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19 pages, 7972 KB

Open AccessEditor’s ChoiceArticle

Rapid Classification and Differentiation of Sepsis-Related Pathogens Using FT-IR Spectroscopy

by Shwan Ahmed, Jawaher Albahri, Sahand Shams, Silvana Sosa-Portugal, Cassio Lima, Yun Xu, Rachel McGalliard, Trevor Jones, Christopher M. Parry, Dorina Timofte, Enitan D. Carrol, Howbeer Muhamadali and Royston Goodacre

Microorganisms 2024, 12(7), 1415; https://doi.org/10.3390/microorganisms12071415 - 12 Jul 2024

Cited by 6 | Viewed by 2728

Abstract

Sepsis is a life-threatening condition arising from a dysregulated host immune response to infection, leading to a substantial global health burden. The accurate identification of bacterial pathogens in sepsis is essential for guiding effective antimicrobial therapy and optimising patient outcomes. Traditional culture-based bacterial [...] Read more.

Sepsis is a life-threatening condition arising from a dysregulated host immune response to infection, leading to a substantial global health burden. The accurate identification of bacterial pathogens in sepsis is essential for guiding effective antimicrobial therapy and optimising patient outcomes. Traditional culture-based bacterial typing methods present inherent limitations, necessitating the exploration of alternative diagnostic approaches. This study reports the successful application of Fourier-transform infrared (FT-IR) spectroscopy in combination with chemometrics as a potent tool for the classification and discrimination of microbial species and strains, primarily sourced from individuals with invasive infections. These samples were obtained from various children with suspected sepsis infections with bacteria and fungi originating at different sites. We conducted a comprehensive analysis utilising 212 isolates from 14 distinct genera, comprising 202 bacterial and 10 fungal isolates. With the spectral analysis taking several weeks, we present the incorporation of quality control samples to mitigate potential variations that may arise between different sample plates, especially when dealing with a large sample size. The results demonstrated a remarkable consistency in clustering patterns among 14 genera when subjected to principal component analysis (PCA). Particularly, Candida, a fungal genus, was distinctly recovered away from bacterial samples. Principal component discriminant function analysis (PC-DFA) allowed for distinct discrimination between different bacterial groups, particularly Gram-negative and Gram-positive bacteria. Clear differentiation was also observed between coagulase-negative staphylococci (CNS) and Staphylococcus aureus isolates, while methicillin-resistant S. aureus (MRSA) was also separated from methicillin-susceptible S. aureus (MSSA) isolates. Furthermore, highly accurate discrimination was achieved between Enterococcus and vancomycin-resistant enterococci isolates with 98.4% accuracy using partial least squares-discriminant analysis. The study also demonstrates the specificity of FT-IR, as it effectively discriminates between individual isolates of Streptococcus and Candida at their respective species levels. The findings of this study establish a strong groundwork for the broader implementation of FT-IR and chemometrics in clinical and microbiological applications. The potential of these techniques for enhanced microbial classification holds significant promise in the diagnosis and management of invasive bacterial infections, thereby contributing to improved patient outcomes. Full article

(This article belongs to the Special Issue Sepsis: Underlying Mechanisms, Pathogen-Host Interactions, Precision Diagnostic, Therapeutic Approaches, and Supportive Management)

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11 pages, 3900 KB

Open AccessEditor’s ChoiceArticle

Oral Microbiota Alterations in Subjects with SARS-CoV-2 Displaying Prevalence of the Opportunistic Fungal Pathogen Candida albicans

by Areej A. Alfaifi, Johanna B. Holm, Tristan W. Wang, Jonathan Lim, Timothy F. Meiller, Peter Rock, Ahmed S. Sultan and Mary Ann Jabra-Rizk

Microorganisms 2024, 12(7), 1356; https://doi.org/10.3390/microorganisms12071356 - 2 Jul 2024

Cited by 4 | Viewed by 2681

Abstract

The oral cavity remains an underappreciated site for SARS-CoV-2 infection despite the myriad of oral conditions in COVID-19 patients. Recently, SARS-CoV-2 was shown to replicate in the salivary gland cells causing tissue inflammation. Given the established association between inflammation and microbiome disruption, we [...] Read more.

The oral cavity remains an underappreciated site for SARS-CoV-2 infection despite the myriad of oral conditions in COVID-19 patients. Recently, SARS-CoV-2 was shown to replicate in the salivary gland cells causing tissue inflammation. Given the established association between inflammation and microbiome disruption, we comparatively profiled oral microbial differences at a metagenomic level in a cohort of hospitalized COVID-19 patients and matched healthy controls. Specifically, we aimed to evaluate colonization by the opportunistic fungal pathogen Candida albicans, the etiologic agent of oral candidiasis. Comprehensive shotgun metagenomic analysis indicated that, overall, COVID-19 patients exhibited significantly reduced bacterial and viral diversity/richness; we identified 12 differentially abundant bacterial species to be negatively associated with COVID-19, and the functional pathways of certain bacteria to be highly associated with COVID-19 status. Strikingly, C. albicans was recovered from approximately half of the COVID-19 subjects but not from any of the healthy controls. The prevalence of Candida is likely linked to immune hypo-dysregulation caused by COVID-19 favoring Candida proliferation, warranting investigations into the interplay between Candida and SARS-CoV2 and potential therapeutic approaches directed toward oral candidiasis. Collectively, our findings prompt a reassessment of oral opportunistic infection risks during COVID-19 disease and their potential long-term impacts on oral health. Full article

(This article belongs to the Special Issue Microbial Interactions including Candida concerning Health and Disease)

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19 pages, 3554 KB

Open AccessEditor’s ChoiceArticle

Metagenomic Investigation of the Short-Term Temporal and Spatial Dynamics of the Bacterial Microbiome and the Resistome Downstream of a Wastewater Treatment Plant in the Iskar River in Bulgaria

by Deyan Donchev, Ivan N. Ivanov, Ivan Stoikov and Monika Ivanova

Microorganisms 2024, 12(6), 1250; https://doi.org/10.3390/microorganisms12061250 - 20 Jun 2024

Cited by 7 | Viewed by 2955

Abstract

Waste Water Treatment Plants (WWTP) aim to reduce contamination in effluent water; however, studies indicate antimicrobial resistance genes (ARGs) persist post-treatment, potentially leading to their spread from human populated areas into the environment. This study evaluated the impact of a large WWTP serving [...] Read more.

Waste Water Treatment Plants (WWTP) aim to reduce contamination in effluent water; however, studies indicate antimicrobial resistance genes (ARGs) persist post-treatment, potentially leading to their spread from human populated areas into the environment. This study evaluated the impact of a large WWTP serving 125,000 people on the Iskar River in Bulgaria, by characterizing the spatial and short-term temporal dynamics in bacterial community dynamics and resistance profiles of the surface water. Pairs of samples were collected biweekly on four dates from two different locations, one about 800 m after the WWTP effluents and the other 10 km downstream. Taxonomic classification revealed the dominance of Pseudomonodota and Bacteriodota, notably the genera Flavobacterium, Aquirufa, Acidovorax, Polynucleobacter, and Limnohabitans. The taxonomic structure corresponded with both lentic and lotic freshwater habitats, with Flavobacterium exhibiting a significant decrease over the study period. Principal Coordinate Analysis revealed statistically significant differences in bacterial community composition between samples collected on different dates. Differential abundance analysis identified notable enrichment of Polynucleobacter and Limnohabitans. There were shifts within the enriched or depleted bacterial taxa between early and late sampling dates. High relative abundance of the genes erm(B), erm(F), mph(E), msr(E) (macrolides); tet(C), tet(O), tet(W), tet(Q) and tet(X) (tetracyclines); sul1 and sul2 (sulphonamides); and cfxA3, cfxA6 (beta-lactams) were detected, with trends of increased presence in the latest sampling dates and in the location closer to the WWTP. Of note, genes conferring resistance to carbapenems blaOXA-58 and blaIMP-33-like were identified. Co-occurrence analysis of ARGs and mobile genetic elements on putative plasmids showed few instances, and the estimated human health risk score (0.19) according to MetaCompare2.0 was low. In total, 29 metagenome-assembled genomes were recovered, with only a few harbouring ARGs. This study enhances our understanding of freshwater microbial community dynamics and antibiotic resistance profiles, highlighting the need for continued ARGs monitoring. Full article

(This article belongs to the Special Issue Water Microorganisms Associated with Human Health)

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14 pages, 1555 KB

Open AccessEditor’s ChoiceArticle

Nisin Inhibition of Gram-Negative Bacteria

by Adam M. Charest, Ethan Reed, Samantha Bozorgzadeh, Lorenzo Hernandez, Natalie V. Getsey, Liam Smith, Anastasia Galperina, Hadley E. Beauregard, Hailey A. Charest, Mathew Mitchell and Margaret A. Riley

Microorganisms 2024, 12(6), 1230; https://doi.org/10.3390/microorganisms12061230 - 19 Jun 2024

Cited by 20 | Viewed by 6767

Abstract

Aims: This study investigates the activity of the broad-spectrum bacteriocin nisin against a large panel of Gram-negative bacterial isolates, including relevant plant, animal, and human pathogens. The aim is to generate supportive evidence towards the use/inclusion of bacteriocin-based therapeutics and open avenues for [...] Read more.

Aims: This study investigates the activity of the broad-spectrum bacteriocin nisin against a large panel of Gram-negative bacterial isolates, including relevant plant, animal, and human pathogens. The aim is to generate supportive evidence towards the use/inclusion of bacteriocin-based therapeutics and open avenues for their continued development. Methods and Results: Nisin inhibitory activity was screened against a panel of 575 strains of Gram-negative bacteria, encompassing 17 genera. Nisin inhibition was observed in 309 out of 575 strains, challenging the prevailing belief that nisin lacks effectiveness against Gram-negative bacteria. The genera Acinetobacter, Helicobacter, Erwinia, and Xanthomonas exhibited particularly high nisin sensitivity. Conclusions: The findings of this study highlight the promising potential of nisin as a therapeutic agent for several key Gram-negative plant, animal, and human pathogens. These results challenge the prevailing notion that nisin is less effective or ineffective against Gram-negative pathogens when compared to Gram-positive pathogens and support future pursuits of nisin as a complementary therapy to existing antibiotics. Significance and Impact of Study: This research supports further exploration of nisin as a promising therapeutic agent for numerous human, animal, and plant health applications, offering a complementary tool for infection control in the face of multidrug-resistant bacteria. Full article

(This article belongs to the Special Issue The Potential of Antimicrobial Activity and Antibiofilm Activity of Bacteriocins)

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17 pages, 3726 KB

Open AccessEditor’s ChoiceArticle

Development of Multiplex RT qPCR Assays for Simultaneous Detection and Quantification of Faecal Indicator Bacteria in Bathing Recreational Waters

by Marina Carrasco-Acosta and Pilar Garcia-Jimenez

Microorganisms 2024, 12(6), 1223; https://doi.org/10.3390/microorganisms12061223 - 18 Jun 2024

Cited by 4 | Viewed by 2992

Abstract

In this study, we designed and validated in silico and experimentally a rapid, sensitive, and specific multiplex RT qPCR for the detection and quantification of faecal indicator bacteria (FIB) used as microbiological references in marine bathing water regulations (Escherichia coli and intestinal [...] Read more.

In this study, we designed and validated in silico and experimentally a rapid, sensitive, and specific multiplex RT qPCR for the detection and quantification of faecal indicator bacteria (FIB) used as microbiological references in marine bathing water regulations (Escherichia coli and intestinal enterococci). The 16S rRNA gene was used to quantify group-specific enterococci and Escherichia/Shigella and species-specific such as Enterococcus faecalis and E. faecium. Additionally, a ybbW gene encoding allantoin transporter protein was used to detect E. coli. An assessment of marine coastal systems (i.e., marine water and sediment) revealed that intestinal enterococci were the predominant group compared to Escherichia/Shigella. The low contribution of E. faecalis to the intestinal enterococci group was reported. As E. faecalis and E. faecium were reported at low concentrations, it is assumed that other enterococci of faecal origin are contributing to the high gene copy number of this group-specific enterococci. Moreover, low 16S rRNA gene copy numbers with respect to E. faecalis and E. faecium were reported in seawater compared to marine sediment. We conclude that marine sediments can affect the quantification of FIBs included in bathing water regulations. Valuing the quality of the marine coastal system through sediment monitoring is recommended. Full article

(This article belongs to the Collection Advances in Public Health Microbiology)

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11 pages, 1467 KB

Open AccessEditor’s ChoiceArticle

Point-of-Care Method T2Bacteria^®Panel Enables a More Sensitive and Rapid Diagnosis of Bacterial Blood Stream Infections and a Shorter Time until Targeted Therapy than Blood Culture

by Tamara Clodi-Seitz, Sebastian Baumgartner, Michael Turner, Theresa Mader, Julian Hind, Christoph Wenisch, Alexander Zoufaly and Elisabeth Presterl

Microorganisms 2024, 12(5), 967; https://doi.org/10.3390/microorganisms12050967 - 11 May 2024

Cited by 7 | Viewed by 3034

Abstract

Background: Rapid diagnosis and identification of pathogens are pivotal for appropriate therapy of blood stream infections. The T2Bacteria^®Panel, a culture-independent assay for the detection of Escherichia coli, Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, and [...] Read more.

Background: Rapid diagnosis and identification of pathogens are pivotal for appropriate therapy of blood stream infections. The T2Bacteria^®Panel, a culture-independent assay for the detection of Escherichia coli, Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, and Pseudomonas aeruginosa in blood, was evaluated under real-world conditions as a point-of-care method including patients admitted to the internal medicine ward due to suspected blood stream infection. Methods: Patients were assigned to two groups (standard of care—SOC vs. T2). In the SOC group 2 × 2 blood culture samples were collected, in the T2 group the T2Bacteria^®Panel was performed additionally for pathogen identification. Results: A total of 94 patients were included. Pathogens were detected in 19 of 50 patients (38%) in the T2 group compared to 16 of 44 patients (36.4%) in the SOC group. The median time until pathogen detection was significantly shorter in the T2 group (4.5 h vs. 60 h, p < 0.001), as well as the time until targeted therapy (antibiotic with the narrowest spectrum and maximal effectiveness) (6.4 h vs. 42.2 h, p = 0.043). Conclusions: The implementation of the T2Bacteria^®Panel for patients with sepsis leads to an earlier targeted antimicrobial therapy resulting in earlier sufficient treatment and decreased excessive usage of broad-spectrum antimicrobials. Full article

(This article belongs to the Special Issue Bacteremia and Sepsis)

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12 pages, 3053 KB

Open AccessEditor’s ChoiceArticle

Growth and Cell Size of Microalga Auxenochlorella protothecoides AS-1 under Different Trophic Modes

by Haoyu Chen, Ana Sosa and Feng Chen

Microorganisms 2024, 12(4), 835; https://doi.org/10.3390/microorganisms12040835 - 20 Apr 2024

Cited by 7 | Viewed by 3828

Abstract

Certain microalgal species can grow with different trophic strategies depending on the availability of nutrient resources. They can use the energy from light or an organic substrate, or both, and can therefore be called autotrophs, heterotrophs, or mixotrophs. We recently isolated a microalgal [...] Read more.

Certain microalgal species can grow with different trophic strategies depending on the availability of nutrient resources. They can use the energy from light or an organic substrate, or both, and can therefore be called autotrophs, heterotrophs, or mixotrophs. We recently isolated a microalgal strain from the microplastic biofilm, which was identified as Auxenochlorella protothecoides, AS-1. Strain AS-1 grew rapidly in bacterial culture media and exhibited different growth rates and cell sizes under different trophic conditions. We compared the growth performance of AS-1 under the three different trophic modes. AS-1 reached a high biomass (>4 g/L) in 6 days under mixotrophic growth conditions with a few organic carbons as a substrate. In contrast, poor autotrophic growth was observed for AS-1. Different cell sizes, including daughter and mother cells, were observed under the different growth modes. We applied a Coulter Counter to measure the size distribution patterns of AS-1 under different trophic modes. We showed that the cell size distribution of AS-1 was affected by different growth modes. Compared to the auto-, hetero- and mixotrophic modes, AS-1 achieved higher biomass productivity by increasing cell number and cell size in the presence of organic substrate. The mechanisms and advantages of having more mother cells with organic substrates are still unclear and warrant further investigations. The work here provides the growth information of a newly isolated A. protothecoides AS-1 which will be beneficial to future downstream applications. Full article

(This article belongs to the Section Molecular Microbiology and Immunology)

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20 pages, 2786 KB

Open AccessEditor’s ChoiceArticle

Arbuscular Mycorrhizal Fungi and Rhizobium Improve Nutrient Uptake and Microbial Diversity Relative to Dryland Site-Specific Soil Conditions

by Rosalie B. Calderon and Sadikshya R. Dangi

Microorganisms 2024, 12(4), 667; https://doi.org/10.3390/microorganisms12040667 - 27 Mar 2024

Cited by 17 | Viewed by 4326

Abstract

Arbuscular mycorrhizal fungi (AMF) and rhizobium play a significant role in plant symbiosis. However, their influence on the rhizosphere soil microbiome associated with nutrient acquisition and soil health is not well defined in the drylands of Montana (MT), USA. This study investigated the [...] Read more.

Arbuscular mycorrhizal fungi (AMF) and rhizobium play a significant role in plant symbiosis. However, their influence on the rhizosphere soil microbiome associated with nutrient acquisition and soil health is not well defined in the drylands of Montana (MT), USA. This study investigated the effect of microbial inoculants as seed treatment on pea yield, nutrient uptake, potential microbial functions, and rhizosphere soil microbial communities using high-throughput sequencing of 16S and ITS rRNA genes. The experiment was conducted under two contrasting dryland conditions with four treatments: control, single inoculation with AMF or Rhizobium, and dual inoculations of AMF and Rhizobium (AMF+Rhizobium). Our findings revealed that microbial inoculation efficacy was site-specific. AMF+Rhizobium synergistically increased grain yield at Sidney dryland field site (DFS) 2, while at Froid site, DFS 1, AMF improved plant resilience to acidic soil but contributed a marginal yield under non-nutrient limiting conditions. Across dryland sites, the plants’ microbial dependency on AMF+Rhizobium (12%) was higher than single inoculations of AMF (8%) or Rhizobium (4%) alone. Variations in microbial community structure and composition indicate a site-specific response to AMF and AMF+Rhizobium inoculants. Overall, site-specific factors significantly influenced plant nutrient uptake, microbial community dynamics, and functional potential. It underscores the need for tailored management strategies that consider site-specific characteristics to optimize benefits from microbial inoculation. Full article

(This article belongs to the Special Issue Soil Microbial Communities and Ecosystem Functions, 2nd Edition)

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25 pages, 5223 KB

Open AccessEditor’s ChoiceArticle

The Post-Transcriptional Regulatory Protein CsrA Amplifies Its Targetome through Direct Interactions with Stress-Response Regulatory Hubs: The EvgA and AcnA Cases

by Alejandra Matsuri Rojano-Nisimura, Kobe B. Grismore, Josie S. Ruzek, Jacqueline L. Avila and Lydia M. Contreras

Microorganisms 2024, 12(4), 636; https://doi.org/10.3390/microorganisms12040636 - 22 Mar 2024

Cited by 4 | Viewed by 3474

Abstract

Global rewiring of bacterial gene expressions in response to environmental cues is mediated by regulatory proteins such as the CsrA global regulator from E. coli. Several direct mRNA and sRNA targets of this protein have been identified; however, high-throughput studies suggest an expanded [...] Read more.

Global rewiring of bacterial gene expressions in response to environmental cues is mediated by regulatory proteins such as the CsrA global regulator from E. coli. Several direct mRNA and sRNA targets of this protein have been identified; however, high-throughput studies suggest an expanded RNA targetome for this protein. In this work, we demonstrate that CsrA can extend its network by directly binding and regulating the evgA and acnA transcripts, encoding for regulatory proteins. CsrA represses EvgA and AcnA expression and disrupting the CsrA binding sites of evgA and acnA, results in broader gene expression changes to stress response networks. Specifically, altering CsrA-evgA binding impacts the genes related to acidic stress adaptation, and disrupting the CsrA-acnA interaction affects the genes involved in metal-induced oxidative stress responses. We show that these interactions are biologically relevant, as evidenced by the improved tolerance of evgA and acnA genomic mutants depleted of CsrA binding sites when challenged with acid and metal ions, respectively. We conclude that EvgA and AcnA are intermediate regulatory hubs through which CsrA can expand its regulatory role. The indirect CsrA regulation of gene networks coordinated by EvgA and AcnA likely contributes to optimizing cellular resources to promote exponential growth in the absence of stress. Full article

(This article belongs to the Special Issue Bacterial Stress Response Mechanisms: Signal Perception, Transduction and Adaptation 2.0)

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19 pages, 6279 KB

Open AccessEditor’s ChoiceArticle

Dinoflagellate Proton-Pump Rhodopsin Genes in Long Island Sound: Diversity and Spatiotemporal Distribution

by Huan Zhang, Kelly J. Nulick, Zair Burris, Melissa Pierce, Minglei Ma and Senjie Lin

Microorganisms 2024, 12(3), 628; https://doi.org/10.3390/microorganisms12030628 - 21 Mar 2024

Cited by 2 | Viewed by 2562

Abstract

Microbial proton-pump rhodopsin (PPR)-based phototrophy, a light-harvesting mechanism different from chlorophyll-based photosystems, may contribute significantly to solar energy entry into the marine ecosystem. PPR transforms solar energy into cellular energy that is used for various metabolic processes in the cells or flagellar movement. [...] Read more.

Microbial proton-pump rhodopsin (PPR)-based phototrophy, a light-harvesting mechanism different from chlorophyll-based photosystems, may contribute significantly to solar energy entry into the marine ecosystem. PPR transforms solar energy into cellular energy that is used for various metabolic processes in the cells or flagellar movement. Although rhodopsins or their encoding genes have been documented in a wide phylogenetic range of cultured dinoflagellates, information is limited about how widespread and how spatiotemporally dynamical dinoflagellate PPR (DiPPR) are in natural marine ecosystems. In this study, we investigated DiPPR in Long Island Sound (LIS), a temperate estuary of the Atlantic Ocean between Connecticut and Long Island, New York, USA. We isolated six novel full-length dinoflagellate proton-pump rhodopsin cDNAs, expanding the DiPPR database that is crucial to PPR research. Based on these new sequences and existing sequences of DiPPR, we designed primers and conducted quantitative PCR and sequencing to determine the abundance and diversity of DiPPR genes spatially and temporally throughout a year in the water samples collected from LIS. DiPPR genes were found year-round and throughout LIS but with higher abundances in the eutrophic Western Sound and in April and July. The gene diversity data suggest that there are at least five distinct rhodopsin-harboring groups of dinoflagellates throughout the year. The abundance of DiPPR genes, measured as copy number per mL of seawater, appeared not to be influenced by water temperature or nitrogen nutrient concentration but exhibited weak negative correlations with orthophosphate concentration and salinity and a positive correlation with the abundance of DiPPR-harboring dinoflagellates. This first quantitative profiling of PPR in natural plankton reveals the prevalence and dynamics of this plastid-independent photoenergy harvesting mechanism in a temperate estuary and provides efficient DiPPR primers potentially useful for future research. Furthermore, this study shed light on the potential role of DiPPR in phosphor nutrition and dinoflagellate population, which warrants further studies. Full article

(This article belongs to the Special Issue Research on Biology of Dinoflagellates)

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22 pages, 3266 KB

Open AccessEditor’s ChoiceArticle

Evaluation of Pyrophosphate-Driven Proton Pumps in Saccharomyces cerevisiae under Stress Conditions

by Krishnan Sreenivas, Leon Eisentraut, Daniel P. Brink, Viktor C. Persson, Magnus Carlquist, Marie F. Gorwa-Grauslund and Ed W. J. van Niel

Microorganisms 2024, 12(3), 625; https://doi.org/10.3390/microorganisms12030625 - 20 Mar 2024

Cited by 2 | Viewed by 2484

Abstract

In Saccharomyces cerevisiae, pH homeostasis is reliant on ATP due to the use of proton-translocating ATPase (H⁺-ATPase) which constitutes a major drain within cellular ATP supply. Here, an exogenous proton-translocating pyrophosphatase (H⁺-PPase) from Arabidopsis thaliana, which uses inorganic [...] Read more.

In Saccharomyces cerevisiae, pH homeostasis is reliant on ATP due to the use of proton-translocating ATPase (H⁺-ATPase) which constitutes a major drain within cellular ATP supply. Here, an exogenous proton-translocating pyrophosphatase (H⁺-PPase) from Arabidopsis thaliana, which uses inorganic pyrophosphate (PP_i) rather than ATP, was evaluated for its effect on reducing the ATP burden. The H⁺-Ppase was localized to the vacuolar membrane or to the cell membrane, and their impact was studied under acetate stress at a low pH. Biosensors (pHluorin and mQueen-2m) were used to observe changes in intracellular pH (pH_i) and ATP levels during growth on either glucose or xylose. A significant improvement of 35% in the growth rate at a pH of 3.7 and 6 g·L⁻¹ acetic acid stress was observed in the vacuolar membrane H⁺-PPase strain compared to the parent strain. ATP levels were elevated in the same strain during anaerobic glucose and xylose fermentations. During anaerobic xylose fermentations, co-expression of pHluorin and a vacuolar membrane H⁺-PPase improved the growth characteristics by means of an improved growth rate (11.4%) and elongated logarithmic growth duration. Our study identified a potential method for improving productivity in the use of S. cerevisiae as a cell factory under the harsh conditions present in industry. Full article

(This article belongs to the Section Microbial Biotechnology)

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11 pages, 1306 KB

Open AccessEditor’s ChoiceArticle

Characterization of the 3,4-Dichloroaniline Degradation Gene Cluster in Acinetobacter soli GFJ2

by Namiko Gibu, Daisuke Kasai, Saki Sato, Michiro Tabata, Alisa Vangnai and Masao Fukuda

Microorganisms 2024, 12(3), 613; https://doi.org/10.3390/microorganisms12030613 - 19 Mar 2024

Cited by 5 | Viewed by 2347

Abstract

3,4-Dichloroaniline (34DCA), a major metabolite of phenylurea herbicides, causes environmental contamination owing to its toxicity and recalcitrant properties. Acinetobacter soli strain GFJ2, isolated from soil potentially contaminated with herbicides, can degrade 34DCA. This study aimed to identify and characterize the 34DCA degradation gene [...] Read more.

3,4-Dichloroaniline (34DCA), a major metabolite of phenylurea herbicides, causes environmental contamination owing to its toxicity and recalcitrant properties. Acinetobacter soli strain GFJ2, isolated from soil potentially contaminated with herbicides, can degrade 34DCA. This study aimed to identify and characterize the 34DCA degradation gene cluster responsible for the conversion of 34DCA to 4,5-dichlorocatechol in the strain GFJ2. Genome analysis revealed one chromosome and seven plasmids in GFJ2, comprising 21, 75, and 3309 copies of rRNA, 75 tRNA, and protein-encoding genes, respectively. A gene cluster responsible for 34DCA degradation was identified, comprising dcdA, dcdB, and dcdC, which encode dioxygenase, flavin reductase, and aldehyde dehydrogenase, respectively. Transcriptional analysis indicated that this gene cluster is constructed as an operon, induced during 34DCA utilization. The heterologous expression of dcdA and dcdB in Escherichia coli confirmed their activity in degrading 34DCA to an intermediate metabolite, converted to 4,5-dichlorocatechol via a reaction involving the dcdC gene product, suggesting their involvement in 34DCA conversion to 4,5-dichlorocatechol. Deletion mutants of dcdA and dcdB lost 34DCA degradation ability, confirming their importance in 34DCA utilization in GFJ2. This study provides insights into the genetic mechanisms of 34DCA degradation by GFJ2, with potential applications in the bioremediation of environments contaminated by phenylurea herbicides. Full article

(This article belongs to the Special Issue Microbial Biocatalysis and Biodegradation 2.0)

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17 pages, 4609 KB

Open AccessEditor’s ChoiceArticle

Recombinant Production of Pseudomonas aeruginosa Rhamnolipids in P. putida KT2440 on Acetobacterium woodii Cultures Grown Chemo-Autotrophically with Carbon Dioxide and Hydrogen

by Jonas Widberger, Andreas Wittgens, Sebastian Klaunig, Markus Krämer, Ann-Kathrin Kissmann, Franziska Höfele, Tina Baur, Tanja Weil, Marius Henkel, Rudolf Hausmann, Frank R. Bengelsdorf, Bernhard J. Eikmanns, Peter Dürre and Frank Rosenau

Microorganisms 2024, 12(3), 529; https://doi.org/10.3390/microorganisms12030529 - 6 Mar 2024

Cited by 5 | Viewed by 3080

Abstract

The establishment of sustainable processes for the production of commodity chemicals is one of today’s central challenges for biotechnological industries. The chemo-autotrophic fixation of CO₂ and the subsequent production of acetate by acetogenic bacteria via anaerobic gas fermentation represents a promising platform [...] Read more.

The establishment of sustainable processes for the production of commodity chemicals is one of today’s central challenges for biotechnological industries. The chemo-autotrophic fixation of CO₂ and the subsequent production of acetate by acetogenic bacteria via anaerobic gas fermentation represents a promising platform for the ecologically sustainable production of high-value biocommodities via sequential fermentation processes. In this study, the applicability of acetate-containing cell-free spent medium of the gas-fermenting acetogenic bacterium A. woodii WP1 as the feeder strain for growth and the recombinant production of P. aeruginosa PAO1 mono-rhamnolipids in the well-established nonpathogenic producer strain P. putida KT2440 were investigated. Additionally, the potential possibility of a simplified production process without the necessary separation of feeder strain cells was elucidated via the cultivation of P. putida in cell-containing A. woodii culture broth. For these cultures, the content of both strains was investigated by examining the relative quantification of strain-exclusive genes via qPCR. The recombinant production of mono-rhamnolipids was successfully achieved with maximum titers of approximately 360–400 mg/L for both cell-free and cell-containing A. woodii spent medium. The reported processes therefore represent a successful proof of principle for gas fermentation-derived acetate as a potential sustainable carbon source for future recombinant rhamnolipid production processes by P. putida KT2440. Full article

(This article belongs to the Special Issue Advances in Microbial Surfactants: Production and Applications)

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15 pages, 2893 KB

Open AccessEditor’s ChoiceArticle

A Retrospective Analysis of Salmonella Isolates across 11 Animal Species (1982–1999) Led to the First Identification of Chromosomally Encoded bla_SCO-1 in the USA

by Nneka Vivian Iduu, Donna Raiford, Austin Conley, Joy Scaria, Julie Nelson, Laura Ruesch, Stuart Price, Min Yue, Jiansen Gong, Lanjing Wei and Chengming Wang

Microorganisms 2024, 12(3), 528; https://doi.org/10.3390/microorganisms12030528 - 6 Mar 2024

Cited by 3 | Viewed by 2436

Abstract

Antimicrobial resistance (AMR) in non-typhoidal Salmonella is a pressing public health concern in the United States, necessitating continuous surveillance. We conducted a retrospective analysis of 251 Salmonella isolates from 11 animal species recovered between 1982 and 1999, utilizing serotyping, antimicrobial susceptibility testing, and [...] Read more.

Antimicrobial resistance (AMR) in non-typhoidal Salmonella is a pressing public health concern in the United States, necessitating continuous surveillance. We conducted a retrospective analysis of 251 Salmonella isolates from 11 animal species recovered between 1982 and 1999, utilizing serotyping, antimicrobial susceptibility testing, and whole-genome sequencing (WGS). Phenotypic resistance was observed in 101 isolates, with S. Typhimurium, S. Dublin, S. Agona, and S. Muenster prevailing among 36 identified serovars. Notably, resistance to 12 of 17 antibiotics was detected, with ampicillin being most prevalent (79/251). We identified 38 resistance genes, primarily mediating aminoglycoside (n = 13) and β-lactamase (n = 6) resistance. Plasmid analysis unveiled nine distinct plasmids associated with AMR genes in these isolates. Chromosomally encoded bla_SCO-1 was present in three S. Typhimurium and two S. Muenster isolates from equine samples, conferring resistance to amoxicillin/clavulanic acid. Phylogenetic analysis revealed three distinct clusters for these five isolates, indicating evolutionary divergence. This study represents the first report of bla_SCO-1 in the USA, and our recovered isolates harboring this gene as early as 1989 precede those of all other reports. The enigmatic nature of bla_SCO-1 prompts further research into its function. Our findings highlight the urgency of addressing antimicrobial resistance in Salmonella for effective public health interventions. Full article

(This article belongs to the Special Issue Antimicrobial Resistance and Genetic Elements in Bacteria, 2nd Edition)

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14 pages, 3411 KB

Open AccessEditor’s ChoiceArticle

A Pseudovirus-Based Neutralization Assay for SARS-CoV-2 Variants: A Rapid, Cost-Effective, BSL-2–Based High-Throughput Assay Useful for Vaccine Immunogenicity Evaluation

by Zhaohui Cai, Raj Kalkeri, Mingzhu Zhu, Shane Cloney-Clark, Benjamin Haner, Mi Wang, Bahar Osman, Dominic Dent, Sheau-Line Feng, Zach Longacre, Greg Glenn and Joyce S. Plested

Microorganisms 2024, 12(3), 501; https://doi.org/10.3390/microorganisms12030501 - 29 Feb 2024

Cited by 11 | Viewed by 5760

Abstract

Neutralizing antibody responses from COVID-19 vaccines are pivotal in conferring protection against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Effective COVID-19 vaccines and assays measuring neutralizing antibodies against emerging variants (i.e., XBB.1.5, XBB.1.16, and XBB.2.3) are needed. The use of biosafety level (BSL)-3 [...] Read more.

Neutralizing antibody responses from COVID-19 vaccines are pivotal in conferring protection against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Effective COVID-19 vaccines and assays measuring neutralizing antibodies against emerging variants (i.e., XBB.1.5, XBB.1.16, and XBB.2.3) are needed. The use of biosafety level (BSL)-3 laboratories for live virus assays results in higher costs and a longer turnaround time; therefore, a BSL-2–based pseudovirus neutralization assay (PNT) was developed. The pseudoviruses were produced by cotransfecting cells with plasmids encoding a lentiviral backbone-expressing luciferase reporter; non-surface proteins for lentiviral production; and ancestral or Omicron (BA.1 and BA.5) SARS-CoV-2 spike (S) proteins. The PNT was developed and optimized in dose and kinetics experiments. The representative serum samples (COVID-19–convalescent or NVX-CoV2373–vaccinated participants enrolled in the 2019nCoV-101 trial) demonstrated a wide dynamic range. The neutralization data showed robust correlation with validated anti-recombinant spike IgG levels and angiotensin-converting enzyme 2 inhibition titers (ancestral). This assay is suitable for measurement of the neutralization ability in clinical samples from individuals infected with SARS-CoV-2 or immunized with a COVID-19 vaccine. The results suggest that this PNT provides a lower cost, high-throughput, rapid turnaround alternative to BSL-3–based microneutralization assays and enables the discovery and development of effective vaccines against emerging variants. Full article

(This article belongs to the Section Virology)

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12 pages, 1121 KB

Open AccessEditor’s ChoiceArticle

Altered Plasma microRNA Signature in Hospitalized COVID-19 Patients Requiring Oxygen Support

by Sandra Franco, Lourdes Mateu, Raquel Pluvinet, Jose Francisco Sanchez-Herrero, Ruth Toledo, Lauro Sumoy, Marta Massanella and Miguel Angel Martinez

Microorganisms 2024, 12(3), 440; https://doi.org/10.3390/microorganisms12030440 - 21 Feb 2024

Cited by 4 | Viewed by 2538

Abstract

To discover potential micro(mi)RNA biomarkers of SARS-CoV-2 infection and disease progression, large-scale deep-sequencing analysis of small RNA expression was performed on plasma samples from 40 patients hospitalized for SARS-CoV-2 infection (median 13.50 [IQR 9–24] days since symptoms initiation) and 21 healthy noninfected individuals. [...] Read more.

To discover potential micro(mi)RNA biomarkers of SARS-CoV-2 infection and disease progression, large-scale deep-sequencing analysis of small RNA expression was performed on plasma samples from 40 patients hospitalized for SARS-CoV-2 infection (median 13.50 [IQR 9–24] days since symptoms initiation) and 21 healthy noninfected individuals. A total of 1218 different miRNAs were identified. When compared with healthy noninfected donors, SARS-CoV-2-infected patients showed significantly (fold change [FC] > 1.2 and adjusted p [padj] < 0.05) altered expression of 190 miRNAs. The top-10 differentially expressed (DE) miRNAs were miR-122-5p, let-7b-5p, miR-146a-5p, miR-342-3p, miR-146b-5p, miR-629-5p, miR-24-3p, miR-12136, let-7a-5p, and miR-191-5p, which displayed FC and padj values ranging from 153 to 5 and 2.51 × 10⁻³² to 2.21 × 10⁻²¹, respectively, which unequivocally diagnosed SARS-CoV-2 infection. No differences in blood cell counts and biochemical plasma parameters, including interleukin 6, ferritin, and D-dimer, were observed between COVID-19 patients on high-flow oxygen therapy, low-flow oxygen therapy, or not requiring oxygen therapy. Notably, 31 significantly deregulated miRNAs were found, when patients on high- and low-flow oxygen therapy were compared. SARS-CoV-2 infection generates a specific miRNA signature in hospitalized patients. Specific miRNA profiles are associated with COVID-19 prognosis in patients requiring oxygen flow. Full article

(This article belongs to the Special Issue Molecular Mechanisms of Viral Persistence and Immune Evasion)

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17 pages, 1616 KB

Open AccessEditor’s ChoiceArticle

Vine-Winery Byproducts as Precious Resource of Natural Antimicrobials: In Vitro Antibacterial and Antibiofilm Activity of Grape Pomace Extracts against Foodborne Pathogens

by Daniela Sateriale, Giuseppina Forgione, Martina Di Rosario, Chiara Pagliuca, Roberta Colicchio, Paola Salvatore, Marina Paolucci and Caterina Pagliarulo

Microorganisms 2024, 12(3), 437; https://doi.org/10.3390/microorganisms12030437 - 21 Feb 2024

Cited by 25 | Viewed by 3406

Abstract

Grape pomace is the main by-product of vine-winery chains. It requires adequate treatment and disposal but is also an economically underused source of bioactive plant secondary metabolites. This study aimed to investigate the antibacterial effects of polyphenolic extracts from Aglianico (Vitis vinifera [...] Read more.

Grape pomace is the main by-product of vine-winery chains. It requires adequate treatment and disposal but is also an economically underused source of bioactive plant secondary metabolites. This study aimed to investigate the antibacterial effects of polyphenolic extracts from Aglianico (Vitis vinifera L.) grape pomace. In particular, hydroethanolic extracts obtained via an ultrasonic-assisted extraction technique were selected for antimicrobial tests. The extracts were screened for their antibacterial effects against foodborne pathogens that were both Gram-positive, in the case of Staphylococcus aureus and Bacillus cereus, and Gram-negative, in the case of Escherichia coli and Salmonella enterica subsp. enterica serovar Typhimurium, showing variable bacteriostatic and bactericidal effects. In addition, our results demonstrated that the tested grape pomace extracts can reduce the inhibitory concentration of standard antibiotics. Interestingly, selected extracts inhibited biofilm development by S. aureus and B. cereus. Overall, these new insights into the antibacterial properties of grape pomace extracts may represent a relevant step in the design of novel therapeutic tools to tackle foodborne diseases, and in the management of resistant biofilm-related infections. Full article

(This article belongs to the Special Issue Antimicrobial Effects of Bioactive Compounds: Phytoproducts-Based Fight against Pathogens)

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13 pages, 3215 KB

Open AccessEditor’s ChoiceArticle

Identification of Genes Associated with Resistance to Persulcatusin, a Tick Defensin from Ixodes persulcatus

by So Shimoda, Junya Ito, Tasuke Ando, Ryuta Tobe, Kiyotaka Nakagawa and Hiroshi Yoneyama

Microorganisms 2024, 12(2), 412; https://doi.org/10.3390/microorganisms12020412 - 19 Feb 2024

Cited by 2 | Viewed by 2394

Abstract

Antimicrobial peptides (AMPs) are present in a wide range of plants, animals, and microorganisms. Since AMPs are characterized by their effectiveness against emergent antibiotic-resistant bacteria, they are attracting attention as next-generation antimicrobial compounds that could solve the problem of drug-resistant bacteria. Persulcatusin (IP), [...] Read more.

Antimicrobial peptides (AMPs) are present in a wide range of plants, animals, and microorganisms. Since AMPs are characterized by their effectiveness against emergent antibiotic-resistant bacteria, they are attracting attention as next-generation antimicrobial compounds that could solve the problem of drug-resistant bacteria. Persulcatusin (IP), an antibacterial peptide derived from the hard tick Ixodes persulcatus, shows high antibacterial activity against various Gram- positive bacteria as well as multidrug-resistant bacteria. However, reports on the antibacterial action and resistance mechanisms of IP are scarce. In this study, we spontaneously generated mutants showing increased a minimum inhibitory concentration (MIC) of IP and analyzed their cross-resistance to other AMPs and antibiotics. We also used fluorescent probes to investigate the target of IP activity by evaluating IP-induced damage to the bacterial cytoplasmic membrane. Our findings suggest that the antimicrobial activity of IP on bacterial cytoplasmic membranes occurs via a mechanism of action different from that of known AMPs. Furthermore, we screened for mutants with high susceptibility to IP using a transposon mutant library and identified 16 genes involved in IP resistance. Our results indicate that IP, like other AMPs, depolarizes the bacterial cytoplasmic membrane, but it may also alter membrane structure and inhibit cell-wall synthesis. Full article

(This article belongs to the Special Issue Antimicrobial Peptides: Therapeutic Potentials 2.0)

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16 pages, 4046 KB

Open AccessEditor’s ChoiceArticle

Harnessing Phosphorous (P) Fertilizer-Insensitive Bacteria to Enhance Rhizosphere P Bioavailability in Legumes

by Antisar Afkairin, Mary M. Dixon, Cassidy Buchanan, James A. Ippolito, Daniel K. Manter, Jessica G. Davis and Jorge M. Vivanco

Microorganisms 2024, 12(2), 353; https://doi.org/10.3390/microorganisms12020353 - 8 Feb 2024

Cited by 11 | Viewed by 2389

Abstract

Phosphorous (P) is widely used in agriculture; yet, P fertilizers are a nonrenewable resource. Thus, mechanisms to improve soil P bioavailability need to be found. Legumes are efficient in P acquisition and, therefore, could be used to develop new technologies to improve soil [...] Read more.

Phosphorous (P) is widely used in agriculture; yet, P fertilizers are a nonrenewable resource. Thus, mechanisms to improve soil P bioavailability need to be found. Legumes are efficient in P acquisition and, therefore, could be used to develop new technologies to improve soil P bioavailability. Here, we studied different species and varieties of legumes and their rhizosphere microbiome responses to low-P stress. Some varieties of common beans, cowpeas, and peas displayed a similar biomass with and without P fertilization. The rhizosphere microbiome of those varieties grown without P was composed of unique microbes displaying different levels of P solubilization and mineralization. When those varieties were amended with P, some of the microbes involved in P solubilization and mineralization decreased in abundance, but other microbes were insensitive to P fertilization. The microbes that decreased in abundance upon P fertilization belonged to groups that are commonly used as biofertilizers such as Pseudomonas and Azospirillum. The microbes that were not affected by P fertilization constitute unique species involved in P mineralization such as Arenimonas daejeonensis, Hyphomicrobium hollandicum, Paenibacillus oenotherae, and Microlunatus speluncae. These P-insensitive microbes could be used to optimize P utilization and drive future sustainable agricultural practices to reduce human dependency on a nonrenewable resource. Full article

(This article belongs to the Section Plant Microbe Interactions)

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15 pages, 2548 KB

Open AccessEditor’s ChoiceArticle

The Antimicrobial Resistance of Enterotoxigenic Escherichia coli from Diarrheal Patients and the Environment in Two Geographically Distinct Rural Areas in Bangladesh over the Years

by Fatema-Tuz Johura, Marzia Sultana, Abdus Sadique, Shirajum Monira, David A. Sack, Richard Bradley Sack, Munirul Alam and Subhra Chakraborty

Microorganisms 2024, 12(2), 301; https://doi.org/10.3390/microorganisms12020301 - 31 Jan 2024

Cited by 13 | Viewed by 3739

Abstract

Antimicrobial resistance (AMR) is an unprecedented global health challenge, involving the transfer of bacteria and genes between humans and the environment. We simultaneously and longitudinally determined the AMR of enterotoxigenic Escherichia coli (ETEC) strains isolated from diarrheal patients and an aquatic environment over [...] Read more.

Antimicrobial resistance (AMR) is an unprecedented global health challenge, involving the transfer of bacteria and genes between humans and the environment. We simultaneously and longitudinally determined the AMR of enterotoxigenic Escherichia coli (ETEC) strains isolated from diarrheal patients and an aquatic environment over two years from two geographically distinct locations, Coastal Mathbaria and Northern Chhatak in Bangladesh. A total of 60% and 72% of ETEC strains from the patients in Mathbaria and Chhatak, respectively, were multi-drug resistant (MDR) with a high proportion of ETEC resistant to nalidixic acid (80.7%), macrolides (49.1–89.7%), ampicillin (57.9–69%), and trimethoprim/sulfamethoxazole (55.2%). From the surface water, 68.8% and 30% of ETEC were MDR in Mathbaria and Chhatak, respectively, with a high proportion of ETEC strains resistant to macrolides (87.5–100%), ampicillin (50–75%), ceftriaxone (62.5%), and nalidixic acid (40%). Notably, 80–100% of the ETEC strains were susceptible to tetracycline and quinolones (ciprofloxacin and norfloxacin), both in clinical and aquatic ETEC. The AMR varied by the ETEC toxin types. The patterns of excessive or limited consumption of drugs to treat diarrhea over time in Bangladesh were reflected in the ETEC AMR from the patients and the environment. The high prevalence of MDR-ETEC strains in humans and the environment is of concern, which calls for vaccines and other preventative measures against ETEC. Full article

(This article belongs to the Special Issue Enterotoxigenic Escherichia coli Infection and Vaccine-Mediated Immunity)

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21 pages, 3473 KB

Open AccessEditor’s ChoiceArticle

Investigation of Staphylococcus aureus Biofilm-Associated Toxin as a Potential Squamous Cell Carcinoma Therapeutic

by Zi Xin Ong, Bavani Kannan, Anthony R. J. Phillips and David L. Becker

Microorganisms 2024, 12(2), 293; https://doi.org/10.3390/microorganisms12020293 - 30 Jan 2024

Cited by 3 | Viewed by 2906

Abstract

Cancer therapies developed using bacteria and their components have been around since the 19th century. Compared to traditional cancer treatments, the use of bacteria-derived compounds as cancer therapeutics could offer a higher degree of specificity, with minimal off-target effects. Here, we explored the [...] Read more.

Cancer therapies developed using bacteria and their components have been around since the 19th century. Compared to traditional cancer treatments, the use of bacteria-derived compounds as cancer therapeutics could offer a higher degree of specificity, with minimal off-target effects. Here, we explored the use of soluble bacteria-derived toxins as a potential squamous cell carcinoma (SCC) therapeutic. We optimized a protocol to generate Staphylococcus aureus biofilm-conditioned media (BCM), where soluble bacterial products enriched in the development of biofilms were isolated from a bacterial culture and applied to SCC cell lines. Bioactive components of S. aureus ATCC 29213 (SA29213) BCM display selective toxicity towards cancerous human skin SCC-12 at low doses, while non-cancerous human keratinocyte HaCaT and fibroblast BJ-5ta are minimally affected. SA29213 BCM treatment causes DNA damage to SCC-12 and initiates Caspase 3-dependent-regulated cell death. The use of the novel SA29213 bursa aurealis transposon mutant library led to the identification of S. aureus alpha hemolysin as the main bioactive compound responsible for the observed SCC-12-specific toxicity. The antibody neutralisation of Hla eradicates the cytotoxicity of SA29213 BCM towards SCC-12. Hla displays high SCC-12-specific toxicity, which is exerted primarily through Hla-ADAM10 interaction, Hla oligomerisation, and pore formation. The high target specificity and potential to cause cell death in a controlled manner highlight SA29213 Hla as a good candidate as an alternative SCC therapeutic. Full article

(This article belongs to the Section Biofilm)

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24 pages, 11828 KB

Open AccessEditor’s ChoiceArticle

The Citizen Phage Library: Rapid Isolation of Phages for the Treatment of Antibiotic Resistant Infections in the UK

by Julie Fletcher, Robyn Manley, Christian Fitch, Christina Bugert, Karen Moore, Audrey Farbos, Michelle Michelsen, Shayma Alathari, Nicola Senior, Alice Mills, Natalie Whitehead, James Soothill, Stephen Michell and Ben Temperton

Microorganisms 2024, 12(2), 253; https://doi.org/10.3390/microorganisms12020253 - 25 Jan 2024

Cited by 11 | Viewed by 8572

Abstract

Antimicrobial resistance poses one of the greatest threats to global health and there is an urgent need for new therapeutic options. Phages are viruses that infect and kill bacteria and phage therapy could provide a valuable tool for the treatment of multidrug-resistant infections. [...] Read more.

Antimicrobial resistance poses one of the greatest threats to global health and there is an urgent need for new therapeutic options. Phages are viruses that infect and kill bacteria and phage therapy could provide a valuable tool for the treatment of multidrug-resistant infections. In this study, water samples collected by citizen scientists as part of the Citizen Phage Library (CPL) project, and wastewater samples from the Environment Agency yielded phages with activity against clinical strains Klebsiella pneumoniae BPRG1484 and Enterobacter cloacae BPRG1482. A total of 169 and 163 phages were found for K. pneumoniae and E. cloacae, respectively, within four days of receiving the strains. A third strain (Escherichia coli BPRG1486) demonstrated cross-reactivity with 42 E. coli phages already held in the CPL collection. Seed lots were prepared for four K. pneumoniae phages and a cocktail combining these phages was found to reduce melanisation in a Galleria mellonella infection model. The resources and protocols utilised by the Citizen Phage Library enabled the rapid isolation and characterisation of phages targeted against multiple strains. In the future, within a clearly defined regulatory framework, phage therapy could be made available on a named-patient basis within the UK. Full article

(This article belongs to the Special Issue The Phage: Biology and Therapeutic Use)

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17 pages, 4259 KB

Open AccessEditor’s ChoiceArticle

Genetic Insights into Biofilm Formation by a Pathogenic Strain of Vibrio harveyi

by Amandine Morot, François Delavat, Alexis Bazire, Christine Paillard, Alain Dufour and Sophie Rodrigues

Microorganisms 2024, 12(1), 186; https://doi.org/10.3390/microorganisms12010186 - 17 Jan 2024

Cited by 3 | Viewed by 3657

Abstract

The Vibrio genus includes bacteria widely distributed in aquatic habitats and the infections caused by these bacteria can affect a wide range of hosts. They are able to adhere to numerous surfaces, which can result in biofilm formation that helps maintain them in [...] Read more.

The Vibrio genus includes bacteria widely distributed in aquatic habitats and the infections caused by these bacteria can affect a wide range of hosts. They are able to adhere to numerous surfaces, which can result in biofilm formation that helps maintain them in the environment. The involvement of the biofilm lifestyle in the virulence of Vibrio pathogens of aquatic organisms remains to be investigated. Vibrio harveyi ORM4 is a pathogen responsible for an outbreak in European abalone Haliotis tuberculata populations. In the present study, we used a dynamic biofilm culture technique coupled with laser scanning microscopy to characterize the biofilm formed by V. harveyi ORM4. We furthermore used RNA-seq analysis to examine the global changes in gene expression in biofilm cells compared to planktonic bacteria, and to identify biofilm- and virulence-related genes showing altered expression. A total of 1565 genes were differentially expressed, including genes associated with motility, polysaccharide synthesis, and quorum sensing. The up-regulation of 18 genes associated with the synthesis of the type III secretion system suggests that this virulence factor is induced in V. harveyi ORM4 biofilms, providing indirect evidence of a relationship between biofilm and virulence. Full article

(This article belongs to the Special Issue Advances in Microbial Biofilm Formation)

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13 pages, 2628 KB

Open AccessEditor’s ChoiceArticle

Diversity of Microbial Mats in the Makgadikgadi Salt Pans, Botswana

by Sevasti Filippidou, Alex Price, Charlotte Spencer-Jones, Anthony Scales, Michael C. Macey, Fulvio Franchi, Lesedi Lebogang, Barbara Cavalazzi, Susanne P. Schwenzer and Karen Olsson-Francis

Microorganisms 2024, 12(1), 147; https://doi.org/10.3390/microorganisms12010147 - 11 Jan 2024

Cited by 6 | Viewed by 2898

Abstract

The Makgadikgadi Salt Pans are the remnants of a mega paleo-lake system in the central Kalahari, Botswana. Today, the Makgadikgadi Basin is an arid to semi-arid area receiving water of meteoric origin during the short, wet season. Large microbial mats, which support primary [...] Read more.

The Makgadikgadi Salt Pans are the remnants of a mega paleo-lake system in the central Kalahari, Botswana. Today, the Makgadikgadi Basin is an arid to semi-arid area receiving water of meteoric origin during the short, wet season. Large microbial mats, which support primary production, are formed due to desiccation during the dry season. This study aimed to characterise the microbial diversity of the microbial mats and the underlying sediment. The focus was the Ntwetwe Pan, located west of the Makgadikgadi Basin. Metagenomic analyses demonstrated that the mats consisted of a high relative abundance of Cyanobacteriota (synonym Cyanobacteria) (20.50–41.47%), Pseudomonadota (synonym Proteobacteria) (15.71 to 32.18%), and Actinomycetota (synonym Actinobacteria) (8.53–32.56%). In the underlying sediments, Pseudomonadota, Actinomycetota, and Euryarchaeota represented over 70% of the community. Localised fluctuations in water content and pH did not significantly affect the microbial diversity of the sediment or the mats. Full article

(This article belongs to the Special Issue New Insights into the Diversity and Characterization of Extremophiles)

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16 pages, 2359 KB

Open AccessEditor’s ChoiceArticle

Targeted Metagenomic Databases Provide Improved Analysis of Microbiota Samples

by Agnes Baud and Sean P. Kennedy

Microorganisms 2024, 12(1), 135; https://doi.org/10.3390/microorganisms12010135 - 10 Jan 2024

Cited by 3 | Viewed by 2989

Abstract

We report on Moonbase, an innovative pipeline that builds upon the established tools of MetaPhlAn and Kraken2, enhancing their capabilities for more precise taxonomic detection and quantification in diverse microbial communities. Moonbase enhances the performance of Kraken2 mapping by providing an efficient method [...] Read more.

We report on Moonbase, an innovative pipeline that builds upon the established tools of MetaPhlAn and Kraken2, enhancing their capabilities for more precise taxonomic detection and quantification in diverse microbial communities. Moonbase enhances the performance of Kraken2 mapping by providing an efficient method for constructing project-specific databases. Moonbase was evaluated using synthetic metagenomic samples and compared against MetaPhlAn3 and generalized Kraken2 databases. Moonbase significantly improved species precision and quantification, outperforming marker genes and generalized databases. Construction of a phylogenetic tree from 16S genome data in Moonbase allowed for the incorporation of UniFrac-type phylogenetic information into diversity calculations of samples. We demonstrated that the resulting analysis increased statistical power in distinguishing microbial communities. This study highlights the continual evolution of metagenomic tools with the goal of improving metagenomic analysis and highlighting the potential of the Moonbase pipeline. Full article

(This article belongs to the Section Microbiomes)

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15 pages, 3265 KB

Open AccessEditor’s ChoiceArticle

Solitary Bees Host More Bacteria and Fungi on Their Cuticle than Social Bees

by Markus Thamm, Fabienne Reiß, Leon Sohl, Martin Gabel, Matthias Noll and Ricarda Scheiner

Microorganisms 2023, 11(11), 2780; https://doi.org/10.3390/microorganisms11112780 - 16 Nov 2023

Cited by 9 | Viewed by 3342

Abstract

Bees come into contact with bacteria and fungi from flowering plants during their foraging trips. The Western honeybee (Apis mellifera) shows a pronounced hygienic behavior with social interactions, while the solitary red mason bee (Osmia bicornis) lacks a social [...] Read more.

Bees come into contact with bacteria and fungi from flowering plants during their foraging trips. The Western honeybee (Apis mellifera) shows a pronounced hygienic behavior with social interactions, while the solitary red mason bee (Osmia bicornis) lacks a social immune system. Since both visit the same floral resources, it is intriguing to speculate that the body surface of a solitary bee should harbor a more complex microbiome than that of the social honeybee. We compared the cuticular microbiomes of A. mellifera (including three European subspecies) and O. bicornis for the first time by bacterial 16S rRNA and fungal ITS gene-based high-throughput amplicon sequencing. The cuticular microbiome of the solitary O. bicornis was significantly more complex than that of the social A. mellifera. The microbiome composition of A. mellifera subspecies was very similar. However, we counted significantly different numbers of fungi and a higher diversity in the honeybee subspecies adapted to warmer climates. Our results suggest that the cuticular microbiome of bees is strongly affected by visited plants, lifestyle and adaptation to temperature, which have important implications for the maintenance of the health of bees under conditions of global change. Full article

(This article belongs to the Section Veterinary Microbiology)

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22 pages, 2387 KB

Open AccessEditor’s ChoiceArticle

Monophasic Variant of Salmonella Typhimurium Infection Affects the Serum Metabolome in Swine

by Guillaume Larivière-Gauthier, Annaëlle Kerouanton, Sophie Mompelat, Stéphanie Bougeard, Martine Denis and Philippe Fravalo

Microorganisms 2023, 11(10), 2565; https://doi.org/10.3390/microorganisms11102565 - 15 Oct 2023

Cited by 3 | Viewed by 2260

Abstract

Salmonella is the most relevant foodborne zoonotic agent found in swine, and its presence in French herds is significant. Its carriage is asymptomatic, which makes it difficult to detect during rearing, thus increasing the risk of its presence on pork meat. Studies have [...] Read more.

Salmonella is the most relevant foodborne zoonotic agent found in swine, and its presence in French herds is significant. Its carriage is asymptomatic, which makes it difficult to detect during rearing, thus increasing the risk of its presence on pork meat. Studies have shown that enteric infection in animals could be associated with changes in the serum metabolome composition, through the immune response or changes in the digestive microbiota composition. We hypothesized that these changes in the serum metabolome composition could be used as markers for the detection of asymptomatic animals infected by Salmonella. Using untargeted analysis by liquid chromatography coupled with mass spectrometry, we showed that significant differences in the composition of the serum metabolome could be detected between infected or noninfected animals both 1 and 21 days after experimental infection. This serum metabolome composition significantly changed during the 21 days postinfection in the infected animal groups, suggesting an evolution of the impact of infection with time. Despite this evolution, differences in the serum metabolome composition persisted between infected and noninfected animals 21 days after the initial infection. We also showed a possible difference between high-shedding and low-shedding animals 21 days postinfection. Finally, some of the variations in the metabolome were found to be significantly associated with variations of specific members of the fecal microbiota. Thus, excreting and asymptomatic animals, but also high-shedding animals, could be identified on the basis of their serum metabolome composition. Full article

(This article belongs to the Section Veterinary Microbiology)

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14 pages, 1894 KB

Open AccessEditor’s ChoiceArticle

Heat-Labile Enterotoxin Decreases Macrophage Phagocytosis of Enterotoxigenic Escherichia coli

by Ian E. Hollifield, Natalya I. Motyka, Kaylynn A. Fernando and Jacob P. Bitoun

Microorganisms 2023, 11(8), 2121; https://doi.org/10.3390/microorganisms11082121 - 21 Aug 2023

Cited by 6 | Viewed by 3168

Abstract

Enterotoxigenic E. coli (ETEC) are endemic in low-resource settings and cause robust secretory diarrheal disease in children less than five years of age. ETEC cause secretory diarrhea by producing the heat-stable (ST) and/or heat-labile (LT) enterotoxins. Recent studies have shown that ETEC can [...] Read more.

Enterotoxigenic E. coli (ETEC) are endemic in low-resource settings and cause robust secretory diarrheal disease in children less than five years of age. ETEC cause secretory diarrhea by producing the heat-stable (ST) and/or heat-labile (LT) enterotoxins. Recent studies have shown that ETEC can be carried asymptomatically in children and adults, but how ETEC subvert mucosal immunity to establish intestinal residency remains unclear. Macrophages are innate immune cells that can be exploited by enteric pathogens to evade mucosal immunity, so we interrogated the ability of ETEC and other E. coli pathovars to survive within macrophages. Using gentamicin protection assays, we show that ETEC H10407 is phagocytosed more readily than other ETEC and non-ETEC isolates. Furthermore, we demonstrate that ETEC H10407, at high bacterial burdens, causes nitrite accumulation in macrophages, which is indicative of a proinflammatory macrophage nitric oxide killing response. However, at low bacterial burdens, ETEC H10407 remains viable within macrophages for an extended period without nitrite accumulation. We demonstrate that LT, but not ST, intoxication decreases the number of ETEC phagocytosed by macrophages. Furthermore, we now show that macrophages exposed simultaneously to LPS and LT produce IL-33, which is a cytokine implicated in promoting macrophage alternative activation, iron recycling, and intestinal repair. Lastly, iron restriction using deferoxamine induces IL-33 receptor (IL-33R) expression and allows ETEC to escape macrophages. Altogether, these data demonstrate that LT provides ETEC with the ability to decrease the perceived ETEC burden and suppresses the initiation of inflammation. Furthermore, these data suggest that host IL-33/IL-33R signaling may augment pathways that promote iron restriction to facilitate ETEC escape from macrophages. These data could help explain novel mechanisms of immune subversion that may contribute to asymptomatic ETEC carriage. Full article

(This article belongs to the Special Issue Enterotoxigenic Escherichia coli Infection and Vaccine-Mediated Immunity)

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17 pages, 3283 KB

Open AccessEditor’s ChoiceArticle

Alpha-Gal Bound Aptamer and Vancomycin Synergistically Reduce Staphylococcus aureus Infection In Vivo

by Matthew K. Doherty, Claire Shaw, Leslie Woods and Bart C. Weimer

Microorganisms 2023, 11(7), 1776; https://doi.org/10.3390/microorganisms11071776 - 8 Jul 2023

Cited by 6 | Viewed by 3428

Abstract

Methicillin-resistant Staphylococcus aureus (MRSA) is a pervasive and persistent threat that requires the development of novel therapies or adjuvants for existing ones. Aptamers, small single-stranded oligonucleotides that form 3D structures and can bind to target molecules, provide one possible therapeutic route, especially when [...] Read more.

Methicillin-resistant Staphylococcus aureus (MRSA) is a pervasive and persistent threat that requires the development of novel therapies or adjuvants for existing ones. Aptamers, small single-stranded oligonucleotides that form 3D structures and can bind to target molecules, provide one possible therapeutic route, especially when presented in combination with current antibiotic applications. BALB/c α-1, 3-galactosyltransferase (−/−) knockout (GTKO) mice were infected with MRSA via tail vein IV and subsequently treated with the αSA31 aptamer (n = 4), vancomycin (n = 12), or αSA31 plus vancomycin (n = 12), with split doses in the morning and evening. The heart, lungs, liver, spleen, and kidneys were harvested upon necropsy for histological and qPCR analysis. All mice treated with αSA31 alone died, whereas 5/12 mice treated with vancomycin alone and 7/12 mice treated with vancomycin plus αSA31 survived the course of the experiment. The treatment of MRSA-infected mice with Vancomycin and an adjuvant aptamer αSA31 reduced disease persistence and dispersion as compared to treatment with either vancomycin SA31 alone, indicating the combination of antibiotic and specifically targeted αSA31 aptamer could be a novel way to control MRSA infection. The data further indicate that aptamers may serve as a potential therapeutic option for other emerging antibiotic resistant pathogens. Full article

(This article belongs to the Special Issue Staphylococcal Infections (Host and Pathogenic Factors) 3.0)

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21 pages, 9137 KB

Open AccessEditor’s ChoiceArticle

Comparative Genomic Analysis of a Novel Vibrio sp. Isolated from an Ulcer Disease Event in Atlantic Salmon (Salmo salar)

by Maryam Ghasemieshkaftaki, Ignacio Vasquez, Aria Eshraghi, Anthony Kurt Gamperl and Javier Santander

Microorganisms 2023, 11(7), 1736; https://doi.org/10.3390/microorganisms11071736 - 2 Jul 2023

Cited by 5 | Viewed by 3667

Abstract

Ulcer diseases are a recalcitrant issue at Atlantic salmon (Salmo salar) aquaculture cage-sites across the North Atlantic region. Classical ulcerative outbreaks (also called winter ulcer disease) refer to a skin infection caused by Moritella viscosa. However, several bacterial species are [...] Read more.

Ulcer diseases are a recalcitrant issue at Atlantic salmon (Salmo salar) aquaculture cage-sites across the North Atlantic region. Classical ulcerative outbreaks (also called winter ulcer disease) refer to a skin infection caused by Moritella viscosa. However, several bacterial species are frequently isolated from ulcer disease events, and it is unclear if other undescribed pathogens are implicated in ulcer disease in Atlantic salmon. Although different polyvalent vaccines are used against M. viscosa, ulcerative outbreaks are continuously reported in Atlantic salmon in Canada. This study analyzed the phenotypical and genomic characteristics of Vibrio sp. J383 isolated from internal organs of vaccinated farmed Atlantic salmon displaying clinical signs of ulcer disease. Infection assays conducted on vaccinated farmed Atlantic salmon and revealed that Vibrio sp. J383 causes a low level of mortalities when administered intracelomic at doses ranging from 10⁷–10⁸ CFU/dose. Vibrio sp. J383 persisted in the blood of infected fish for at least 8 weeks at 10 and 12 °C. Clinical signs of this disease were greatest 12 °C, but no mortality and bacteremia were observed at 16 °C. The Vibrio sp. J383 genome (5,902,734 bp) has two chromosomes of 3,633,265 bp and 2,068,312 bp, respectively, and one large plasmid of 201,166 bp. Phylogenetic and comparative analyses indicated that Vibrio sp. J383 is related to V. splendidus, with 93% identity. Furthermore, the phenotypic analysis showed that there were significant differences between Vibrio sp. J383 and other Vibrio spp, suggesting J383 is a novel Vibrio species adapted to cold temperatures. Full article

(This article belongs to the Special Issue Bacterial Infections in Aquaculture of Farmed Fish: Pathogenesis, Pathophysiology, Molecular Diagnosis, and Control)

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12 pages, 1339 KB

Open AccessEditor’s ChoiceArticle

Diversification of Bourbon Virus in New York State

by Rachel E. Lange, Alan P. Dupuis II and Alexander T. Ciota

Microorganisms 2023, 11(6), 1590; https://doi.org/10.3390/microorganisms11061590 - 15 Jun 2023

Cited by 4 | Viewed by 4276

Abstract

Bourbon virus (BRBV, family Orthomyxoviridae) is a tickborne virus recently detected in the United States (US). BRBV was first identified from a fatal human case in 2014 in Bourbon County, Kansas. Enhanced surveillance in Kansas and Missouri implicated Amblyomma americanum as the [...] Read more.

Bourbon virus (BRBV, family Orthomyxoviridae) is a tickborne virus recently detected in the United States (US). BRBV was first identified from a fatal human case in 2014 in Bourbon County, Kansas. Enhanced surveillance in Kansas and Missouri implicated Amblyomma americanum as the primary vector for BRBV. Historically, BRBV was only detected in the lower midwestern US, but since 2020 it has been reported in North Carolina, Virginia, New Jersey, and New York State (NYS). This study aimed to elucidate genetic and phenotypic characteristics of BRBV strains from NYS through whole genome sequencing and the assessment of replication kinetics in mammalian cultures and A. americanum nymphs. Sequence analysis revealed the existence of two divergent BRBV clades circulating in NYS. BRBV NY21-2143 is closely related to the midwestern BRBV strains but has unique substitutions in the glycoprotein. Two other NYS BRBV strains, BRBV NY21-1814 and BRBV NY21-2666, form a distinct clade unique from previously sequenced BRBV strains. Phenotypic diversification was also detected in NYS BRBV strains compared to each other and midwestern BRBV strains, with BRBV NY21-2143 displaying attenuation in rodent-derived cell culture and a fitness advantage in experimentally infected A. americanum. These data suggest genetic and phenotypic diversification of emergent BRBV strains circulating in NYS that could contribute to increased spread of BRBV in the northeastern US. Full article

(This article belongs to the Special Issue Emerging Research on Tick-Borne Pathogens and Diseases)

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16 pages, 2614 KB

Open AccessEditor’s ChoiceArticle

Propidium Monoazide (PMAxx)-Recombinase Polymerase Amplification Exo (RPA Exo) Assay for Rapid Detection of Burkholderia cepacia Complex in Chlorhexidine Gluconate (CHX) and Benzalkonium Chloride (BZK) Solutions

by Soumana Daddy Gaoh, Ohgew Kweon and Youngbeom Ahn

Microorganisms 2023, 11(6), 1401; https://doi.org/10.3390/microorganisms11061401 - 26 May 2023

Cited by 6 | Viewed by 3180

Abstract

Both sterile and non-sterile pharmaceutical products, which include antiseptics, have been recalled due to Burkholderia cepacia complex (BCC) contamination. Therefore, minimizing the frequency of outbreaks may be conducive to the development of a quick and sensitive approach that can distinguish between live and [...] Read more.

Both sterile and non-sterile pharmaceutical products, which include antiseptics, have been recalled due to Burkholderia cepacia complex (BCC) contamination. Therefore, minimizing the frequency of outbreaks may be conducive to the development of a quick and sensitive approach that can distinguish between live and dead loads of BCC. We have assessed an exo probe-based recombinase polymerase amplification (RPA) with 10 µM propidium monoazide (PMAxx) for selective detection of live/dead BCC cells in various concentrations of antiseptics (i.e., chlorhexidine gluconate (CHX) and benzalkonium chloride (BZK) solutions) after 24 h. The optimized assay conducted using a set of primer–probes targeting gbpT was performed at 40 °C for 20 min and shows a detection limit of 10 pg/µL of genomic DNA from B. cenocepacia J2315, equivalent to 10⁴ colony-forming units (CFU/mL). The specificity of a newly designed primer and probe was 80% (20 negatives out of 25). The readings for total cells (i.e., without PMAxx) from 200 µg/mL CHX using PMAxx-RPA exo assay was 310 relative fluorescence units (RFU), compared to 129 RFU with PMAxx (i.e., live cells). Furthermore, in 50–500 µg/mL BZK-treated cells, a difference in the detection rate was observed between the PMAxx-RPA exo assay in live cells (130.4–459.3 RFU) and total cells (207.82–684.5 RFU). This study shows that the PMAxx-RPA exo assay appears to be a valid tool for the simple, rapid and presumptive detection of live BCC cells in antiseptics, thereby ensuring the quality and safety of pharmaceutical products. Full article

(This article belongs to the Special Issue New Insights into Epidemiology, Detection and Characterization of Bacterial Pathogens 2.0)

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15 pages, 1920 KB

Open AccessEditor’s ChoiceArticle

Characterization of Bacterial Transcriptional Regulatory Networks in Escherichia coli through Genome-Wide In Vitro Run-Off Transcription/RNA-seq (ROSE)

by Pascal Schmidt, David Brandt, Tobias Busche and Jörn Kalinowski

Microorganisms 2023, 11(6), 1388; https://doi.org/10.3390/microorganisms11061388 - 25 May 2023

Cited by 4 | Viewed by 4297

Abstract

The global characterization of transcriptional regulatory networks almost exclusively uses in vivo conditions, thereby providing a snapshot on multiple regulatory interactions at the same time. To complement these approaches, we developed and applied a method for characterizing bacterial promoters genome-wide by in vitro [...] Read more.

The global characterization of transcriptional regulatory networks almost exclusively uses in vivo conditions, thereby providing a snapshot on multiple regulatory interactions at the same time. To complement these approaches, we developed and applied a method for characterizing bacterial promoters genome-wide by in vitro transcription coupled to transcriptome sequencing specific for native 5′-ends of transcripts. This method, called ROSE (run-off transcription/RNA-sequencing), only requires chromosomal DNA, ribonucleotides, RNA polymerase (RNAP) core enzyme, and a specific sigma factor, recognizing the corresponding promoters, which have to be analyzed. ROSE was performed on E. coli K-12 MG1655 genomic DNA using Escherichia coli RNAP holoenzyme (including σ70) and yielded 3226 transcription start sites, 2167 of which were also identified in in vivo studies, and 598 were new. Many new promoters not yet identified by in vivo experiments might be repressed under the tested conditions. Complementary in vivo experiments with E. coli K-12 strain BW25113 and isogenic transcription factor gene knockout mutants of fis, fur, and hns were used to test this hypothesis. Comparative transcriptome analysis demonstrated that ROSE could identify bona fide promoters that were apparently repressed in vivo. In this sense, ROSE is well-suited as a bottom-up approach for characterizing transcriptional networks in bacteria and ideally complementary to top-down in vivo transcriptome studies. Full article

(This article belongs to the Special Issue Advances in Bacterial Genetics)

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