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Microorganisms, Volume 8, Issue 5 (May 2020) – 171 articles

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Cover Story (view full-size image) Filamentous bacteriophages are promising immunogenic carriers for antigen delivery. Their use is [...] Read more.
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Open AccessArticle
Processing of Biltong (Dried Beef) to Achieve USDA-FSIS 5-log Reduction of Salmonella without a Heat Lethality Step
Microorganisms 2020, 8(5), 791; https://doi.org/10.3390/microorganisms8050791 - 25 May 2020
Viewed by 510
Abstract
In the US, dried beef products (beef jerky) are a popular snack product in which the manufacture often requires the use of a heat lethality step to provide adequate reduction of pathogens of concern (i.e., 5-log reduction of Salmonella as recommended by the [...] Read more.
In the US, dried beef products (beef jerky) are a popular snack product in which the manufacture often requires the use of a heat lethality step to provide adequate reduction of pathogens of concern (i.e., 5-log reduction of Salmonella as recommended by the United States Department of Agriculture Food Safety and Inspection Service (USDA-FSIS)). Biltong, a South African-style dried beef product, is manufactured with low heat and humidity. Our objectives were to examine processes for the manufacture of biltong that achieves a 5-log reduction of Salmonella without a heat lethality step and with, or without, the use of additional antimicrobials. Beef pieces (1.9 cm × 5.1 cm × 7.6 cm) were inoculated with a 5-serovar mixture of Salmonella (Salmonella Thompson 120, Salmonella Heidelberg F5038BG1, Salmonella Hadar MF60404, Salmonella Enteritidis H3527, and Salmonella Typhimurium H3380), dipped in antimicrobial solutions (lactic acid, acidified calcium sulfate, sodium acid sulfate) or water (no additional antimicrobial), and marinaded while vacuum tumbling and/or while held overnight at 5 °C. After marination, beef pieces were hung in an oven set at 22.2 °C (72 °F), 23.9 °C (75 °F), or 25 °C (77 °F) depending on the process, and maintained at 55% relative humidity. Beef samples were enumerated for Salmonella after inoculation, after dip treatment, after marination, and after 2, 4, 6, and 8 days of drying. Water activity was generally <0.85 by the end of 6–8 days of drying and weight loss was as high as 60%. Trials also examined salt concentration (1.7%, 2.2%, 2.7%) and marinade vinegar composition (2%, 3%, 4%) in the raw formulation. Nearly all approaches achieved 5-log10 reduction of Salmonella and was attributed to the manner of microbial enumeration eliminating the effects of microbial concentration on dried beef due to moisture loss. All trials were run as multiple replications and statistical analysis of treatments were determined by repeated measures analysis of variance (RM-ANOVA) to determine significant differences (p < 0.05). We believe this is the first published report of a biltong process achieving >5.0 log10 reduction of Salmonella which is a process validation requirement of USDA-FSIS for the sale of dried beef in the USA. Full article
(This article belongs to the Special Issue Antimicrobial Interventions for Raw and Processed Foods)
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Open AccessArticle
Evaluation of Indolocarbazoles from Streptomyces sanyensis as a Novel Source of Therapeutic Agents against the Brain-Eating Amoeba Naegleria fowleri
Microorganisms 2020, 8(5), 789; https://doi.org/10.3390/microorganisms8050789 - 25 May 2020
Viewed by 377
Abstract
Naegleria fowleri is an opportunistic pathogenic free-living amoeba which is able to rapidly colonize the central nervous system (CNS) and causes a lethal infection known as primary amoebic meningoencephalitis (PAM). Furthermore, more than 98% of the known cases of PAM are fatal and [...] Read more.
Naegleria fowleri is an opportunistic pathogenic free-living amoeba which is able to rapidly colonize the central nervous system (CNS) and causes a lethal infection known as primary amoebic meningoencephalitis (PAM). Furthermore, more than 98% of the known cases of PAM are fatal and affect mainly children under 12 and young adults. Until now, no fully effective therapeutic agents against N. fowleri are available and hence the urgent need to find novel agents to treat PAM. At present, PAM therapy is based on the combination of amphotericin B, miltefosine, among others, with unwanted toxic effects. Recently, our team isolated various indolocarbazoles (ICZs) from the culture of a mangrove strain of Streptomyces sanyensis which showed activity against kinetoplastids and the Acanthamoeba genus. Hence, in this study, the activity of the previously isolated ICZs, staurosporine (STS), 7-oxostaurosporine (7OSTS), 4′-demethylamino-4′-oxostaurosporine, and streptocarbazole B, was evaluated against two type strains of N. fowleri. Furthermore, the performed activity assays revealed that STS was the most active ICZ presenting an inhibitory concentration 50 (IC50) of 0.08 ± 0.02 µM (SI 109.3). Moreover, STS induced programmed cell death (PCD) in the treated amoebae by triggering DNA condensation, mitochondrial disfunction, cell membrane disruption, and reactive oxygen species (ROS) generation. Therefore, STS could be a promising therapeutic agent against PAM. Full article
(This article belongs to the Special Issue Natural Antimicrobial Compounds)
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Open AccessArticle
Co-Culture with Bifidobacterium catenulatum Improves the Growth, Gut Colonization, and Butyrate Production of Faecalibacterium prausnitzii: In Vitro and In Vivo Studies
Microorganisms 2020, 8(5), 788; https://doi.org/10.3390/microorganisms8050788 - 25 May 2020
Viewed by 472
Abstract
Faecalibacterium prausnitzii is a major commensal bacterium in the human gut. It produces short-chain fatty acids that promote intestinal health. However, the bacterium is extremely oxygen-sensitive, making it difficult to develop as a probiotic. To facilitate practical application of F. prausnitzii, we [...] Read more.
Faecalibacterium prausnitzii is a major commensal bacterium in the human gut. It produces short-chain fatty acids that promote intestinal health. However, the bacterium is extremely oxygen-sensitive, making it difficult to develop as a probiotic. To facilitate practical application of F. prausnitzii, we investigated factors that affect its growth and mammalian gut colonization. We evaluated cross-feeding interactions between F. prausnitzii and seven Bifidobacterium strains, and the anti-inflammatory properties of bacterial metabolites produced in co-culture, in vitro and in vivo. Co-culture of F. prausnitzii and Bifidobacterium catenulatum, with fructooligosaccharides as an energy source, resulted in the greatest viable cell-count and butyrate production increases. Further, the co-culture supernatant reduced the amount of proinflammatory cytokines produced by HT-29 cells and RAW 264.7 macrophages, an effect that was similar to that of butyrate. Furthermore, feeding mice both Faecalibacterium and Bifidobacterium enhanced F. prausnitzii gut colonization. Finally, feeding the co-culture supernatant decreased interleukin 8 levels in the colon and increased butyrate levels in the cecum in the dextran sodium sulfate-induced colitis mouse model. These observations indicate that the Faecalibacterium-Bifidobacterium co-culture exerts an anti-inflammatory effect by promoting F. prausnitzii survival and short-chain fatty acid production, with possible implications for the treatment of inflammatory bowel disease. Full article
(This article belongs to the Section Gut Microbiota)
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Open AccessOpinion
Wine Yeast Terroir: Separating the Wheat from the Chaff—for an Open Debate
Microorganisms 2020, 8(5), 787; https://doi.org/10.3390/microorganisms8050787 - 25 May 2020
Viewed by 339
Abstract
Wine terroir is characterized by a specific taste and style influenced by the cultivar of the fermented grapes, geographical factors such as the vineyard, mesoclimate, topoclimate, and microclimate, soil geology and pedology, and the agronomic approach used. These characteristics together define the concept [...] Read more.
Wine terroir is characterized by a specific taste and style influenced by the cultivar of the fermented grapes, geographical factors such as the vineyard, mesoclimate, topoclimate, and microclimate, soil geology and pedology, and the agronomic approach used. These characteristics together define the concept of “terroir”. Thus, regional distinctive flavors in wine have been the subject of many studies aimed at better understanding the link between the wine and the vineyard. Indeed, the identification of key environmental elements involved in the regional variation of grape and wine quality characteristics is a critical feature for improving wine production in terms of consumer preference and economic appreciation. Many studies have demonstrated the role of abiotic factors in grape composition and consequently in wine style. Biotic factors are also involved such as grape microbial communities. However, the occurrence and effects of region-specific microbiota in defining wine characteristics are more controversial issues. Indeed, several studies using high throughput sequencing technologies have made it possible to describe microbial communities and revealed a link between grape must and soil microbial communities, and the geography of the territory. Based on these observations, the concept of “microbial terroir” emerged. However, this concept has been subject to contradictory studies. The aim of this opinion article is to take a step back and examine in perspective the concept of microbial terroir, by comparing numerous data from different studies and providing arguments in favor of or against this concept to stimulate discussion and point out that experimental research is still needed to study the contribution of this assembly of microorganisms to the final product and to support or refute the concept. Full article
(This article belongs to the Special Issue Yeast in Winemaking)
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Open AccessCommunication
Risk of Symptomatic Infection after Non-Primary Congenital Cytomegalovirus Infection
Microorganisms 2020, 8(5), 786; https://doi.org/10.3390/microorganisms8050786 - 25 May 2020
Viewed by 532
Abstract
Cytomegalovirus (CMV) is the leading cause of congenital infection. Its occurrence is phenotypically heterogeneous. The type of maternal infection, primary or non-primary, is an important factor related to the symptomatic disease, the primary infection was long considered the only cause of severe neonatal [...] Read more.
Cytomegalovirus (CMV) is the leading cause of congenital infection. Its occurrence is phenotypically heterogeneous. The type of maternal infection, primary or non-primary, is an important factor related to the symptomatic disease, the primary infection was long considered the only cause of severe neonatal disease. We aimed to analyze the association of primary and non-primary infection with pathological outcomes in infants and with long-term sequelae at follow-up. This was a monocentric retrospective observational study on a population of 91 infants diagnosed with a CMV infection at the Neonatal Care Unit of Neonatology at the Sant’Anna Hospital of Turin during the period of June 2005 to December 2018. Infants underwent clinical, laboratory, and neuroradiological evaluations at birth. Subsequently, the patients were monitored in an auxological, neurodevelopment, and audiological follow-up. Regarding primary vs. non-primary infection, we found a higher percentage of incidence of symptomatic and neurological localized infection, as well as long-term sequelae in the latter. However, no significant difference between the two populations was found. We underline the possibility of re-infection in previously immunized mothers (non-primary infection) with unfavorable neonatal and long-term outcomes. Full article
(This article belongs to the Special Issue Cytomegalovirus: Biology and Infection)
Open AccessCommunication
Key Bacteria in the Gut Microbiota Network for the Transition between Sedentary and Active Lifestyle
Microorganisms 2020, 8(5), 785; https://doi.org/10.3390/microorganisms8050785 - 24 May 2020
Viewed by 589
Abstract
Physical activity modifies the gut microbiota, exerting health benefits on the host; however, the specific bacteria associated with exercise are not yet known. In this work, we propose a novel method, based on hierarchical topology, to study the differences between the microbiota of [...] Read more.
Physical activity modifies the gut microbiota, exerting health benefits on the host; however, the specific bacteria associated with exercise are not yet known. In this work, we propose a novel method, based on hierarchical topology, to study the differences between the microbiota of active and sedentary lifestyles, and to identify relevant bacterial taxa. Our results show that the microbiota network found in active people has a significantly higher overall efficiency and higher transmissibility rate. We also identified key bacteria in active and sedentary networks that could be involved in the conversion of an active microbial network to a sedentary microbial network and vice versa. Full article
(This article belongs to the Special Issue New Methods in Microbial Research)
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Open AccessReview
The Role of the Microbiota–Gut–Brain Axis and Antibiotics in ALS and Neurodegenerative Diseases
Microorganisms 2020, 8(5), 784; https://doi.org/10.3390/microorganisms8050784 - 23 May 2020
Viewed by 733
Abstract
The human gut hosts a wide and diverse ecosystem of microorganisms termed the microbiota, which line the walls of the digestive tract and colon where they co-metabolize digestible and indigestible food to contribute a plethora of biochemical compounds with diverse biological functions. The [...] Read more.
The human gut hosts a wide and diverse ecosystem of microorganisms termed the microbiota, which line the walls of the digestive tract and colon where they co-metabolize digestible and indigestible food to contribute a plethora of biochemical compounds with diverse biological functions. The influence gut microbes have on neurological processes is largely yet unexplored. However, recent data regarding the so-called leaky gut, leaky brain syndrome suggests a potential link between the gut microbiota, inflammation and host co-metabolism that may affect neuropathology both locally and distally from sites where microorganisms are found. The focus of this manuscript is to draw connection between the microbiota–gut–brain (MGB) axis, antibiotics and the use of “BUGS AS DRUGS” for neurodegenerative diseases, their treatment, diagnoses and management and to compare the effect of current and past pharmaceuticals and antibiotics for alternative mechanisms of action for brain and neuronal disorders, such as Alzheimer disease (AD), Amyotrophic Lateral Sclerosis (ALS), mood disorders, schizophrenia, autism spectrum disorders and others. It is a paradigm shift to suggest these diseases can be largely affected by unknown aspects of the microbiota. Therefore, a future exists for applying microbial, chemobiotic and chemotherapeutic approaches to enhance translational and personalized medical outcomes. Microbial modifying applications, such as CRISPR technology and recombinant DNA technology, among others, echo a theme in shifting paradigms, which involve the gut microbiota (GM) and mycobiota and will lead to potential gut-driven treatments for refractory neurologic diseases. Full article
(This article belongs to the Special Issue Microbiota-Gut-Brain Axis)
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Open AccessArticle
Actinomycetes from the Red Sea Sponge Coscinoderma mathewsi: Isolation, Diversity, and Potential for Bioactive Compounds Discovery
Microorganisms 2020, 8(5), 783; https://doi.org/10.3390/microorganisms8050783 - 23 May 2020
Viewed by 478
Abstract
The diversity of actinomycetes associated with the marine sponge Coscinoderma mathewsi collected from Hurghada (Egypt) was studied. Twenty-three actinomycetes were separated and identified based on the 16S rDNA gene sequence analysis. Out of them, three isolates were classified as novel species of the [...] Read more.
The diversity of actinomycetes associated with the marine sponge Coscinoderma mathewsi collected from Hurghada (Egypt) was studied. Twenty-three actinomycetes were separated and identified based on the 16S rDNA gene sequence analysis. Out of them, three isolates were classified as novel species of the genera Micromonospora, Nocardia, and Gordonia. Genome sequencing of actinomycete strains has revealed many silent biosynthetic gene clusters and has shown their exceptional capacity for the production of secondary metabolites, not observed under classical cultivation conditions. Therefore, the effect of mycolic-acid-containing bacteria or mycolic acid on the biosynthesis of cryptic natural products was investigated. Sponge-derived actinomycete Micromonospora sp. UA17 was co-cultured using liquid fermentation with two mycolic acid-containing actinomycetes (Gordonia sp. UA19 and Nocardia sp. UA 23), or supplemented with pure mycolic acid. LC-HRESIMS data were analyzed to compare natural production across all crude extracts. Micromonospora sp. UA17 was rich with isotetracenone, indolocarbazole, and anthracycline analogs. Some co-culture extracts showed metabolites such as a chlorocardicin, neocopiamycin A, and chicamycin B that were not found in the respective monocultures, suggesting a mycolic acid effect on the induction of cryptic natural product biosynthetic pathways. The antibacterial, antifungal, and antiparasitic activities for the different cultures extracts were also tested. Full article
(This article belongs to the Special Issue Microbial Secondary Metabolites and Biotechnology)
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Open AccessArticle
Is Shiga Toxin-Producing Escherichia coli O45 No Longer a Food Safety Threat? The Danger is Still Out There
Microorganisms 2020, 8(5), 782; https://doi.org/10.3390/microorganisms8050782 - 22 May 2020
Viewed by 479
Abstract
Many Shiga toxin-producing Escherichia coli (STEC) strains, including the serogroups of O157 and most of the top six non-O157 serotypes, are frequently associated with foodborne outbreaks. Therefore, they have been extensively studied using next-generation sequencing technology. However, related information regarding STEC O45 strains [...] Read more.
Many Shiga toxin-producing Escherichia coli (STEC) strains, including the serogroups of O157 and most of the top six non-O157 serotypes, are frequently associated with foodborne outbreaks. Therefore, they have been extensively studied using next-generation sequencing technology. However, related information regarding STEC O45 strains is scarce. In this study, three environmental E. coli O45:H16 strains (RM11911, RM13745, and RM13752) and one clinical E. coli O45:H2 strain (SJ7) were sequenced and used to characterize virulence factors using two reference E. coli O45:H2 strains of clinical origin. Subsequently, whole-genome-based phylogenetic analysis was conducted for the six STEC O45 strains and nine other reference STEC genomes, in order to evaluate their evolutionary relationship. The results show that one locus of enterocyte effacement pathogenicity island was found in all three STEC O45:H2 strains, but not in the STEC O45:H16 strains. Additionally, E. coli O45:H2 strains were evolutionarily close to E. coli O103:H2 strains, sharing high homology in terms of virulence factors, such as Stx prophages, but were distinct from E. coli O45:H16 strains. The findings show that E. coli O45:H2 may be as virulent as E. coli O103:H2, which is frequently associated with severe illness and can provide genomic evidence to facilitate STEC surveillance. Full article
(This article belongs to the Special Issue Shiga Toxin-Producing Escherichia coli)
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Open AccessArticle
Evaluation of Product Distribution in Chemostat and Batch Fermentation in Lactic Acid-Producing Komagataella phaffii Strains Utilizing Glycerol as Substrate
Microorganisms 2020, 8(5), 781; https://doi.org/10.3390/microorganisms8050781 - 22 May 2020
Viewed by 495
Abstract
Lactic acid is the monomeric unit of polylactide (PLA), a bioplastic widely used in the packaging, automotive, food, and pharmaceutical industries. Previously, the yeast Komagataella phaffii was genetically modified for the production of lactate from glycerol. For this, the bovine L-lactate dehydrogenase- (LDH)-encoding [...] Read more.
Lactic acid is the monomeric unit of polylactide (PLA), a bioplastic widely used in the packaging, automotive, food, and pharmaceutical industries. Previously, the yeast Komagataella phaffii was genetically modified for the production of lactate from glycerol. For this, the bovine L-lactate dehydrogenase- (LDH)-encoding gene was inserted and the gene encoding the pyruvate decarboxylase (PDC) was disrupted, resulting in the GLp strain. This showed a yield of 67% L-lactic acid and 20% arabitol as a by-product in batches with oxygen limitation. Following up on these results, the present work endeavored to perform a detailed study of the metabolism of this yeast, as well as perturbing arabitol synthesis in an attempt to increase lactic acid titers. The GLp strain was cultivated in a glycerol-limited chemostat at different dilution rates, confirming that the production of both lactic acid and arabitol is dependent on the specific growth rate (and consequently on the concentration of the limiting carbon source) as well as on the oxygen level. Moreover, disruption of the gene encoding arabitol dehydrogenase (ArDH) was carried out, resulting in an increase of 20% in lactic acid and a 50% reduction in arabitol. This study clarifies the underlying metabolic reasons for arabitol formation in K. phaffii and points to ways for improving production of lactic acid using K. phaffii as a biocatalyst. Full article
(This article belongs to the Special Issue Yeast Fermentation)
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Open AccessArticle
Phage Display Detection of Mimotopes that Are Shared Epitopes of Clinically and Epidemiologically Relevant Enterobacteria
Microorganisms 2020, 8(5), 780; https://doi.org/10.3390/microorganisms8050780 - 22 May 2020
Viewed by 392
Abstract
Background: Escherichia coli and Salmonella are etiologic agents of intestinal infections. A previous study showed the presence of shared epitopes between lipopolysaccharides (LPSs) of E. coli O157 and Salmonella. Aim: Using phage display, the aim of this study is to identify mimotopes [...] Read more.
Background: Escherichia coli and Salmonella are etiologic agents of intestinal infections. A previous study showed the presence of shared epitopes between lipopolysaccharides (LPSs) of E. coli O157 and Salmonella. Aim: Using phage display, the aim of this study is to identify mimotopes of shared epitopes in different enterobacterial LPSs. Methods: We use anti-LPS IgG from E. coli O157 and Salmonella to select peptide mimotopes of the M13 phage. The amino acid sequence of the mimotopes is used to synthesize peptides, which are in turn used to immunize rabbits. The antibody response of the resulting sera against the LPSs and synthetic peptides (SPs) is analyzed by ELISA and by Western blot assays, indicating that LPS sites are recognized by the same antibody. In a complementary test, the reactions of human serum samples obtained from the general population against the SPs and LPSs are also analyzed. Results: From the last biopanning phase, sixty phagotopes are selected. The analysis of the peptide mimotope amino acid sequences shows that in 4 of them the S/N/A/PF motif is a common sequence. Antibodies from the sera of immunized rabbits with SP287/3, SP459/1, SP308/3, and SP073/14 react against both their own peptide and the different LPSs. The Western blot test shows a sera reaction against both the lateral chains and the cores of the LPSs. The analysis of the human sera shows a response against the SPs and LPSs. Conclusion: The designed synthetic peptides are mimotopes of LPS epitopes of Salmonella and E. coli that possess immunogenic capacity. These mimotopes could be considered for use in the design of vaccines against both enterobacteria. Full article
(This article belongs to the Special Issue Phage Display—Perspectives and Translational Applications)
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Open AccessArticle
Pathophysiology of Hyperechogenic Bowel in Congenitally Human Cytomegalovirus Infected Fetuses
Microorganisms 2020, 8(5), 779; https://doi.org/10.3390/microorganisms8050779 - 22 May 2020
Viewed by 356
Abstract
Hyperechogenic bowel (HB) is a nonspecific ultrasound finding that can be associated with human cytomegalovirus (CMV) congenital infection. In this study, we investigated HB pathophysiology in CMV-infected fetuses. We examined small and large intestine as well as pancreas in 8 fetuses at 22 [...] Read more.
Hyperechogenic bowel (HB) is a nonspecific ultrasound finding that can be associated with human cytomegalovirus (CMV) congenital infection. In this study, we investigated HB pathophysiology in CMV-infected fetuses. We examined small and large intestine as well as pancreas in 8 fetuses at 22 weeks of gestation with congenital CMV infection. Ultrasound findings showed 4 fetuses with HB and 4 without. As negative group, 4 fetuses without CMV infection and without HB were studied. Immunohistochemistry for CMV, lymphocytic infiltrate, B-cell leukemia/lymphoma-2 (bcl-2), CD-117, cystic fibrosis transmembrane regulator (CFTR) were performed. HB fetuses showed multiple and sequential CMV-positive ganglion cells of Auerbach’s myenteric plexus. In the ganglia, bcl-2 was weakly expressed representing a reduced neuronal functionality. CD-117 revealed a regular distribution of Cajal cells, the pacemakers of intestinal contractility. Pancreas showed normal CFTR staining, indicating a preserved exocrine secretion, thus unlikely a contributory factor in HB. In CMV-infected fetuses without HB, CMV-positive cells were scatteredly found in ganglion cells and bcl-2 was strongly expressed. Intestinal CD-117 and pancreatic CFTR expression were similar to fetuses with HB. In conclusion, fetal CMV infection of the bowel may lead to peristalsis impairment (paralytic ileus) due to intestinal plexus involvement, which at ultrasound appeared as HB. Full article
(This article belongs to the Special Issue Cytomegalovirus: Biology and Infection)
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Open AccessReview
Evolution and Adaptation of the Avian H7N9 Virus into the Human Host
Microorganisms 2020, 8(5), 778; https://doi.org/10.3390/microorganisms8050778 - 21 May 2020
Viewed by 519
Abstract
Influenza viruses arise from animal reservoirs, and have the potential to cause pandemics. In 2013, low pathogenic novel avian influenza A(H7N9) viruses emerged in China, resulting from the reassortment of avian-origin viruses. Following evolutionary changes, highly pathogenic strains of avian influenza A(H7N9) viruses [...] Read more.
Influenza viruses arise from animal reservoirs, and have the potential to cause pandemics. In 2013, low pathogenic novel avian influenza A(H7N9) viruses emerged in China, resulting from the reassortment of avian-origin viruses. Following evolutionary changes, highly pathogenic strains of avian influenza A(H7N9) viruses emerged in late 2016. Changes in pathogenicity and virulence of H7N9 viruses have been linked to potential mutations in the viral glycoproteins hemagglutinin (HA) and neuraminidase (NA), as well as the viral polymerase basic protein 2 (PB2). Recognizing that effective viral transmission of the influenza A virus (IAV) between humans requires efficient attachment to the upper respiratory tract and replication through the viral polymerase complex, experimental evidence demonstrates the potential H7N9 has for increased binding affinity and replication, following specific amino acid substitutions in HA and PB2. Additionally, the deletion of extended amino acid sequences in the NA stalk length was shown to produce a significant increase in pathogenicity in mice. Research shows that significant changes in transmissibility, pathogenicity and virulence are possible after one or a few amino acid substitutions. This review aims to summarise key findings from that research. To date, all strains of H7N9 viruses remain restricted to avian reservoirs, with no evidence of sustained human-to-human transmission, although mutations in specific viral proteins reveal the efficacy with which these viruses could evolve into a highly virulent and infectious, human-to-human transmitted virus. Full article
(This article belongs to the Special Issue The Biology of Influenza Viruses)
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Open AccessReview
Delivery of Heterologous Proteins, Enzymes, and Antigens via the Bacterial Type III Secretion System
Microorganisms 2020, 8(5), 777; https://doi.org/10.3390/microorganisms8050777 - 21 May 2020
Viewed by 335
Abstract
The Type III Secretion System (T3SS) is a multimeric protein complex composed of over 20 different proteins, utilized by Gram-negative bacteria to infect eukaryotic host cells. The T3SS has been implicated as a virulence factor by which pathogens cause infection and has recently [...] Read more.
The Type III Secretion System (T3SS) is a multimeric protein complex composed of over 20 different proteins, utilized by Gram-negative bacteria to infect eukaryotic host cells. The T3SS has been implicated as a virulence factor by which pathogens cause infection and has recently been characterized as a communication tool between bacteria and plant cells in the rhizosphere. The T3SS has been repurposed to be used as a tool for the delivery of non-native or heterologous proteins to eukaryotic cells or the extracellular space for a variety of purposes, including drug discovery and drug delivery. This review covers the methodology of heterologous protein secretion as well as multiple cases of utilizing the T3SS to deliver heterologous proteins or artificial materials. The research covered in this review will serve to outline the scope and limitations of utilizing the T3SS as a tool for protein delivery. Full article
(This article belongs to the Section Microbial Biotechnology)
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Open AccessArticle
Determination of Loline Alkaloids and Mycelial Biomass in Endophyte-Infected Schedonorus pratensis by Near-Infrared Spectroscopy and Chemometrics
Microorganisms 2020, 8(5), 776; https://doi.org/10.3390/microorganisms8050776 - 21 May 2020
Viewed by 543
Abstract
Near infrared spectroscopy (NIRS) is an accurate, fast and nondestructive technique whose use in predicting forage quality has become increasingly relevant in recent decades. Epichloë-infected grass varieties are commonly used in areas with high pest pressure due to their better performances compared [...] Read more.
Near infrared spectroscopy (NIRS) is an accurate, fast and nondestructive technique whose use in predicting forage quality has become increasingly relevant in recent decades. Epichloë-infected grass varieties are commonly used in areas with high pest pressure due to their better performances compared to endophyte-free varieties. The insect resistance of Epichloë-infected grasses has been associated with four main groups of endophyte secondary metabolites: ergot alkaloids, indole-diterpenes, lolines and peramine. Concentrations of these alkaloids are usually measured with high performance liquid chromatography or gas chromatography analysis, which are accurate methods but relatively expensive and laborious. In this paper, we developed a rapid method based on NIRS to detect and quantify loline alkaloids in wild accessions of Schedonorus pratensis infected with the fungal endophyte Epichloë uncinata. The quantitative NIR equations obtained by modified partial least squares algorithm had coefficients of correlation of 0.90, 0.78, 0.85, 0.90 for N-acetylloline, N-acetylnorloline and N-formylloline and the sum of the three, respectively. The acquired NIR spectra were also used for developing an equation to predict in planta fungal biomass with a coefficient of correlation of 0.75. These results showed that the use of NIRS and chemometrics allows the quantification of loline alkaloids and mycelial biomass in a heterogeneous set of endophyte-infected meadow fescue samples. Full article
(This article belongs to the Special Issue Fungal Endophytes and Their Interactions with Plants)
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Open AccessArticle
The Crimean-Congo Hemorrhagic Fever Virus NSm Protein Is Dispensable for Growth In Vitro and Disease in Ifnar-/- Mice
Microorganisms 2020, 8(5), 775; https://doi.org/10.3390/microorganisms8050775 - 21 May 2020
Cited by 1 | Viewed by 419
Abstract
Crimean-Congo hemorrhagic fever virus (CCHFV) is a tri-segmented, tick-borne nairovirus that causes disease of ranging severity in humans. The CCHFV M segment encodes a complex glycoprotein precursor (GPC) that undergoes extensive endoproteolytic cleavage, giving rise to two structural proteins (Gn and Gc) required [...] Read more.
Crimean-Congo hemorrhagic fever virus (CCHFV) is a tri-segmented, tick-borne nairovirus that causes disease of ranging severity in humans. The CCHFV M segment encodes a complex glycoprotein precursor (GPC) that undergoes extensive endoproteolytic cleavage, giving rise to two structural proteins (Gn and Gc) required for virus attachment and entry, and to multiple non-structural proteins (NSm, GP160, GP85, and GP38). The functions of these non-structural proteins remain largely unclear. Here, we investigate the role of NSm during infection by generating a recombinant CCHFV lacking the complete NSm domain (10200∆NSm) and observing CCHFV ∆NSm replication in cell lines and pathogenicity in Ifnar-/- mice. Our data demonstrate that the NSm domain is dispensable for viral replication in vitro, and, despite the delayed onset of clinical signs, CCHFV lacking this domain caused severe or lethal disease in infected mice. Full article
(This article belongs to the Special Issue Hemorrhagic Fever Viruses: Pathogenesis and Countermeasures)
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Open AccessArticle
Comparative Genomics of the Rhodococcus Genus Shows Wide Distribution of Biodegradation Traits
Microorganisms 2020, 8(5), 774; https://doi.org/10.3390/microorganisms8050774 - 21 May 2020
Viewed by 713
Abstract
The genus Rhodococcus exhibits great potential for bioremediation applications due to its huge metabolic diversity, including biotransformation of aromatic and aliphatic compounds. Comparative genomic studies of this genus are limited to a small number of genomes, while the high number of sequenced strains [...] Read more.
The genus Rhodococcus exhibits great potential for bioremediation applications due to its huge metabolic diversity, including biotransformation of aromatic and aliphatic compounds. Comparative genomic studies of this genus are limited to a small number of genomes, while the high number of sequenced strains to date could provide more information about the Rhodococcus diversity. Phylogenomic analysis of 327 Rhodococcus genomes and clustering of intergenomic distances identified 42 phylogenomic groups and 83 species-level clusters. Rarefaction models show that these numbers are likely to increase as new Rhodococcus strains are sequenced. The Rhodococcus genus possesses a small “hard” core genome consisting of 381 orthologous groups (OGs), while a “soft” core genome of 1253 OGs is reached with 99.16% of the genomes. Models of sequentially randomly added genomes show that a small number of genomes are enough to explain most of the shared diversity of the Rhodococcus strains, while the “open” pangenome and strain-specific genome evidence that the diversity of the genus will increase, as new genomes still add more OGs to the whole genomic set. Most rhodococci possess genes involved in the degradation of aliphatic and aromatic compounds, while short-chain alkane degradation is restricted to a certain number of groups, among which a specific particulate methane monooxygenase (pMMO) is only found in Rhodococcus sp. WAY2. The analysis of Rieske 2Fe-2S dioxygenases among rhodococci genomes revealed that most of these enzymes remain uncharacterized. Full article
(This article belongs to the Section Systems Microbiology)
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Open AccessBrief Report
Manipulation of Alcohol and Short-Chain Fatty Acids in the Metabolome of Commensal and Virulent Klebsiella pneumoniae by Linolenic Acid
Microorganisms 2020, 8(5), 773; https://doi.org/10.3390/microorganisms8050773 - 21 May 2020
Viewed by 310
Abstract
Endogenous alcohol produced by the gut microbiome is transported via the bloodstream to the liver for detoxification. Gut dysbiosis can result in chronic excess alcohol production that contributes to the development of hepatic steatosis. The aim of this study was to examine whether [...] Read more.
Endogenous alcohol produced by the gut microbiome is transported via the bloodstream to the liver for detoxification. Gut dysbiosis can result in chronic excess alcohol production that contributes to the development of hepatic steatosis. The aim of this study was to examine whether linolenic acid can manipulate the production of harmful alcohol and beneficial short-chain fatty acids (SCFAs) in the metabolome of commensal Klebsiella pneumoniae (K. pneumoniae) and the virulent K. pneumoniae K1 serotype. Glucose fermentation by the K. pneumoniae K1 serotype yielded increased production of alcohol and decreased SCFAs (especially acetate and propionate) compared to those of commensal K. pneumoniae. However, the use of linolenic acid instead of glucose significantly reduced alcohol and increased SCFAs in the fermentation media of the K. pneumoniae K1 serotype. The work highlights the value of shaping the microbial metabolome using linolenic acid, which can potentially regulate the gut–liver axis for the prevention and treatment of alcohol-induced liver diseases. Full article
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Open AccessArticle
Microbial Consortiums of Hydrogenotrophic Methanogenic Mixed Cultures in Lab-Scale Ex-Situ Biogas Upgrading Systems under Different Conditions of Temperature, pH and CO
Microorganisms 2020, 8(5), 772; https://doi.org/10.3390/microorganisms8050772 - 21 May 2020
Viewed by 441
Abstract
In this study, hydrogenotrophic methanogenic mixed cultures taken from 13 lab-scale ex-situ biogas upgrading systems under different temperature (20–70 °C), pH (6.0–8.5), and CO (0–10%, v/v) variables were systematically investigated. High-throughput 16S rRNA gene sequencing was used to identify the [...] Read more.
In this study, hydrogenotrophic methanogenic mixed cultures taken from 13 lab-scale ex-situ biogas upgrading systems under different temperature (20–70 °C), pH (6.0–8.5), and CO (0–10%, v/v) variables were systematically investigated. High-throughput 16S rRNA gene sequencing was used to identify the microbial consortia, and statistical analyses were conducted to reveal the microbial diversity, the core functional microbes, and their correlative relationships with tested variables. Overall, bacterial community was more complex than the archaea community in all mixed cultures. Hydrogenotrophic methanogens Methanothermobacter, Methanobacterium, and Methanomassiliicoccus, and putative syntrophic acetate-oxidizing bacterium Coprothermobacter and Caldanaerobacter were found to predominate, but the core functional microbes varied under different conditions. Multivariable sensitivity analysis indicated that temperature (p < 0.01) was the crucial variable to determine the microbial consortium structures in hydrogenotrophic methanogenic mixed cultures. pH (0.01 < p < 0.05) significantly interfered with the relative abundance of dominant archaea. Although CO did not affect community (p > 0.1), some potential CO-utilizing syntrophic metabolisms might be enhanced. Understanding of microbial consortia in the hydrogenotrophic methanogenic mixed cultures related to environmental variables was a great advance to reveal the microbial ecology in microbial biogas upgrading process. Full article
(This article belongs to the Special Issue Anaerobic Digestion)
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Open AccessArticle
Insight into the Lifestyle of Amoeba Willaertia magna during Bioreactor Growth Using Transcriptomics and Proteomics
Microorganisms 2020, 8(5), 771; https://doi.org/10.3390/microorganisms8050771 - 21 May 2020
Viewed by 945
Abstract
Willaertia magna C2c maky is a thermophilic free-living amoeba strain that showed ability to eliminate Legionella pneumophila, a pathogenic bacterium living in the aquatic environment. The amoeba industry has proposed the use of Willaertia magna as a natural biocide to control L. [...] Read more.
Willaertia magna C2c maky is a thermophilic free-living amoeba strain that showed ability to eliminate Legionella pneumophila, a pathogenic bacterium living in the aquatic environment. The amoeba industry has proposed the use of Willaertia magna as a natural biocide to control L. pneumophila proliferation in cooling towers. Here, transcriptomic and proteomic studies were carried out in order to expand knowledge on W. magna produced in a bioreactor. Illumina RNA-seq generated 217 million raw reads. A total of 8790 transcripts were identified, of which 6179 and 5341 were assigned a function through comparisons with National Center of Biotechnology Information (NCBI) reference sequence and the Clusters of Orthologous Groups of proteins (COG) databases, respectively. To corroborate these transcriptomic data, we analyzed the W. magna proteome using LC–MS/MS. A total of 3561 proteins were identified. The results of transcriptome and proteome analyses were highly congruent. Metabolism study showed that W. magna preferentially consumed carbohydrates and fatty acids to grow. Finally, an in-depth analysis has shown that W. magna produces several enzymes that are probably involved in the metabolism of secondary metabolites. Overall, our multi-omic study of W. magna opens the way to a better understanding of the genetics and biology of this amoeba. Full article
(This article belongs to the Section Microbial Biotechnology)
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Open AccessArticle
Detection of Canine Vector-Borne Filariasis and Their Wolbachia Endosymbionts in French Guiana
Microorganisms 2020, 8(5), 770; https://doi.org/10.3390/microorganisms8050770 - 21 May 2020
Viewed by 447
Abstract
In French Guiana, canine heartworm disease is well known, but the diversity of filarial parasites of dogs remains largely unknown. A total of 98 canine blood samples from Cayenne and Kourou were assessed by a blood wet mount preparation, heartworm antigen test and [...] Read more.
In French Guiana, canine heartworm disease is well known, but the diversity of filarial parasites of dogs remains largely unknown. A total of 98 canine blood samples from Cayenne and Kourou were assessed by a blood wet mount preparation, heartworm antigen test and molecular exploration of filarioid and Wolbachia DNAs, followed by a multiplex species-specific qPCR’s identification and a subsequent sequencing analysis. Thereafter, a phylogeny based on maximum likelihood was carried out to facilitate specific identification. Five dogs were microfilaremic. Heartworm antigens were detected in 15 (15.3%) dogs. Of these, six (6.1%) were considered as occult infections as neither microfilariae nor Dirofilaria immitis DNA were detected. The 11 (11.2%) D. immitis isolates corresponded to a low virulent strain. Six of the D. immitis isolates were positive for Wolbachia endosymbionts of D. immitis belonging to the clade C DNA. Acanthocheilonema reconditum DNA was detected in 3 (3.1%) samples. Of these latter, one was found co-infected with the Brugia sp. genotype and the DNA of the clade D of the Wolbachia endosymbiont of Brugia species. This latter was also detected in two filarioid DNA-free samples. Finally, two samples were positive for Cercopithifilaria bainae genotype, which is distinct from those identified in Europe. The present study highlights the urgent need to implement chemoprophylaxis associated with anti-Wolbachia drugs to control these potential zoonoses. Full article
(This article belongs to the Special Issue Canine Vector Borne Diseases)
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Open AccessCorrection
Correction: Wang, B., et al. Pectin Degradation Is an Important Determinant for Alfalfa Silage Fermentation through the Rescheduling of the Bacterial Community. Microorganisms 2020, 8, 488
Microorganisms 2020, 8(5), 769; https://doi.org/10.3390/microorganisms8050769 - 20 May 2020
Viewed by 337
Abstract
The authors wish to make the following correction to this paper [...] Full article
(This article belongs to the Special Issue Lactic Acid Bacteria, Biopreservation Agents for Fruit and Vegetables)
Open AccessArticle
Pan-Genome-Based Analysis as a Framework for Demarcating Two Closely Related Methanotroph Genera Methylocystis and Methylosinus
Microorganisms 2020, 8(5), 768; https://doi.org/10.3390/microorganisms8050768 - 20 May 2020
Viewed by 407
Abstract
The Methylocystis and Methylosinus are two of the five genera that were included in the first taxonomic framework of methanotrophic bacteria created half a century ago. Members of both genera are widely distributed in various environments and play a key role in reducing [...] Read more.
The Methylocystis and Methylosinus are two of the five genera that were included in the first taxonomic framework of methanotrophic bacteria created half a century ago. Members of both genera are widely distributed in various environments and play a key role in reducing methane fluxes from soils and wetlands. The original separation of these methanotrophs in two distinct genera was based mainly on their differences in cell morphology. Further comparative studies that explored various single-gene-based phylogenies suggested the monophyletic nature of each of these genera. Current availability of genome sequences from members of the Methylocystis/Methylosinus clade opens the possibility for in-depth comparison of the genomic potentials of these methanotrophs. Here, we report the finished genome sequence of Methylocystis heyeri H2T and compare it to 23 currently available genomes of Methylocystis and Methylosinus species. The phylogenomic analysis confirmed that members of these genera form two separate clades. The Methylocystis/Methylosinus pan-genome core comprised 1173 genes, with the accessory genome containing 4941 and 11,192 genes in the shell and the cloud, respectively. Major differences between the genome-encoded environmental traits of these methanotrophs include a variety of enzymes for methane oxidation and dinitrogen fixation as well as genomic determinants for cell motility and photosynthesis. Full article
(This article belongs to the Special Issue Biology, Diversity, and Ecology of Methanotrophic Bacteria)
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Open AccessArticle
Environmental Impact on Differential Composition of Gut Microbiota in Indoor Chickens in Commercial Production and Outdoor, Backyard Chickens
Microorganisms 2020, 8(5), 767; https://doi.org/10.3390/microorganisms8050767 - 20 May 2020
Viewed by 387
Abstract
In this study, we compared the caecal microbiota composition of egg-laying hens from commercial production that are kept indoors throughout their whole life with microbiota of hens kept outdoors. The microbiota of outdoor hens consisted of lower numbers of bacterial species than the [...] Read more.
In this study, we compared the caecal microbiota composition of egg-laying hens from commercial production that are kept indoors throughout their whole life with microbiota of hens kept outdoors. The microbiota of outdoor hens consisted of lower numbers of bacterial species than the microbiota of indoor hens. At the phylum level, microbiota of outdoor hens was enriched for Bacteroidetes (62.41 ± 4.47% of total microbiota in outdoor hens and 52.01 ± 6.27% in indoor hens) and Proteobacteria (9.33 ± 4.99% in outdoor and 5.47 ± 2.24% in indoor hens). On the other hand, Firmicutes were more abundant in the microbiota of indoor hens (33.28 ± 5.11% in indoor and 20.66 ± 4.41% in outdoor hens). Horizontally transferrable antibiotic resistance genes tetO, tet(32), tet(44), and tetW were also less abundant in the microbiota of outdoor hens than indoor hens. A comparison of the microbiota composition at the genus and species levels pointed toward isolates specifically adapted to the two extreme environments. However, genera and species recorded as being similarly abundant in the microbiota of indoor and outdoor hens are equally as noteworthy because these represent microbiota members that are highly adapted to chickens, irrespective of their genetics, feed composition, and living environment. Full article
(This article belongs to the Special Issue Gut Microbiota Development in Farm Animals)
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Open AccessPerspective
Proteomics of Brucella: Technologies and Their Applications for Basic Research and Medical Microbiology
Microorganisms 2020, 8(5), 766; https://doi.org/10.3390/microorganisms8050766 - 20 May 2020
Viewed by 414
Abstract
Brucellosis is a global zoonosis caused by Gram-negative, facultative intracellular bacteria of the genus Brucella (B.). Proteomics has been used to investigate a few B. melitensis and B. abortus strains, but data for other species and biovars are limited. Hence, a [...] Read more.
Brucellosis is a global zoonosis caused by Gram-negative, facultative intracellular bacteria of the genus Brucella (B.). Proteomics has been used to investigate a few B. melitensis and B. abortus strains, but data for other species and biovars are limited. Hence, a comprehensive analysis of proteomes will significantly contribute to understanding the enigmatic biology of brucellae. For direct identification and typing of Brucella, matrix-assisted laser desorption ionization—time of flight mass spectrometry (MALDI—TOF MS) has become a reliable tool for routine diagnosis due to its ease of handling, price and sensitivity highlighting the potential of proteome-based techniques. Proteome analysis will also help to overcome the historic but still notorious Brucella obstacles of infection medicine, the lack of safe and protective vaccines and sensitive serologic diagnostic tools by identifying the most efficient protein antigens. This perspective summarizes past and recent developments in Brucella proteomics with a focus on species identification and serodiagnosis. Future applications of proteomics in these fields are discussed. Full article
(This article belongs to the Special Issue Updates on Brucellosis)
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Open AccessArticle
Dual-Purpose Inoculants and Their Effects on Corn Silage
Microorganisms 2020, 8(5), 765; https://doi.org/10.3390/microorganisms8050765 - 20 May 2020
Viewed by 358
Abstract
This study was conducted to screen dual-purpose lactic acid bacteria (LAB) from uncontrolled farm-scale silage, and then we confirmed their effects on corn silage. The LAB were isolated from eight farm-scale corn silages, and then we screened the antifungal activity against Fusarium graminearum [...] Read more.
This study was conducted to screen dual-purpose lactic acid bacteria (LAB) from uncontrolled farm-scale silage, and then we confirmed their effects on corn silage. The LAB were isolated from eight farm-scale corn silages, and then we screened the antifungal activity against Fusarium graminearum and the carboxylesterase activity using spectrophotometer with p-nitrophenyl octanoate as substrate and McIlvane solution as buffer. From a total of 25 isolates, 5M2 and 6M1 isolates were selected as silage inoculants because presented both activities of antifungal and carboxylesterase. According 16S rRNA gene sequencing method, 5M2 isolate had 100.0% similarity with Lactobacillus brevis, and 6M1 isolate had 99.7% similarity with L. buchneri. Corn forage was ensiled in bale silo (500 kg) for 72 d without inoculant (CON) or with mixture of selected isolates at 1:1 ratio (INO). The INO silage had higher nutrient digestibility in the rumen than CON silage. Acetate was higher and yeasts were lower in INO silage than in CON silage on the day of silo opening. In all days of aerobic exposure, yeasts were lower in INO silage than CON silage. The present study concluded that Lactobacillus brevis 5M2 and L. buchneri 6M1 confirmed antifungal and carboxylesterase activities on farm-scale corn silage. Full article
(This article belongs to the Special Issue Lactic Acid Bacteria, Biopreservation Agents for Fruit and Vegetables)
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Open AccessArticle
Entrapped Psychrotolerant Yeast Cells within Pine Sawdust for Low Temperature Wine Making: Impact on Wine Quality
Microorganisms 2020, 8(5), 764; https://doi.org/10.3390/microorganisms8050764 - 20 May 2020
Viewed by 315
Abstract
An alternative methodology is proposed for low temperature winemaking using freeze-dried raw materials. Pine sawdust was delignified and the received porous cellulosic material was applied as immobilization carrier of the psychrotolerant yeast strain Saccharomyces cerevisiae AXAZ-1. The immobilization of yeast cells was examined [...] Read more.
An alternative methodology is proposed for low temperature winemaking using freeze-dried raw materials. Pine sawdust was delignified and the received porous cellulosic material was applied as immobilization carrier of the psychrotolerant yeast strain Saccharomyces cerevisiae AXAZ-1. The immobilization of yeast cells was examined and verified by scanning electron microscopy (SEM). The immobilized biocatalyst and high-gravity grape must were separately freeze-dried without cryoprotectants and stored at room temperature (20–22 °C) for 3 months. The effect of storage on the fermentation efficiency of the immobilized biocatalyst at low temperatures (1–10 °C), as well as on the aromatic characteristics of the produced wines was evaluated. Storage time had no significant effect on the fermentation efficiency of the biocatalyst resulting in most cases in high ethanol production 13.8–14.8% v/v. The volatile fraction of the produced wines was examined using headspace solid-phase microextraction (HS-SPME) followed by gas chromatography mass spectrometry (GC/MS). GC-MS/SPME analysis along with the organoleptic evaluation revealed in all produced wines a plethora of fresh and fruit aromatic notes. To conclude, fermentation kinetics and aromatic profile evaluation encourages the production of high-quality sweet wines at low temperatures using pine sawdust (Pinus halepensis) entrapped yeast cells as a promoter. Full article
(This article belongs to the Special Issue Substrate Tolerant Fermentations)
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Open AccessArticle
Evaluation of a Highly Efficient DNA Extraction Method for Bacillus anthracis Endospores
Microorganisms 2020, 8(5), 763; https://doi.org/10.3390/microorganisms8050763 - 20 May 2020
Viewed by 413
Abstract
A variety of methods have been established in order to optimize the accessibility of DNA originating from Bacillus anthracis cells and endospores to facilitate highly sensitive molecular diagnostics. However, most endospore lysis techniques have not been evaluated in respect to their quantitative proficiencies. [...] Read more.
A variety of methods have been established in order to optimize the accessibility of DNA originating from Bacillus anthracis cells and endospores to facilitate highly sensitive molecular diagnostics. However, most endospore lysis techniques have not been evaluated in respect to their quantitative proficiencies. Here, we started by systematically assessing the efficiencies of 20 DNA extraction kits for vegetative B. anthracis cells. Of these, the Epicentre MasterPure kit gave the best DNA yields and quality suitable for further genomic analysis. Yet, none of the kits tested were able to extract reasonable quantities of DNA from cores of the endospores. Thus, we developed a mechanical endospore lysis protocol, facilitating the extraction of high-quality DNA. Transmission electron microscopy or the labelling of spores with the indicator dye propidium monoazide was utilized to assess lysis efficiency. Finally, the yield and quality of genomic spore DNA were quantified by PCR and they were found to be dependent on lysis matrix composition, instrumental parameters, and the method used for subsequent DNA purification. Our final standardized lysis and DNA extraction protocol allows for the quantitative detection of low levels (<50 CFU/mL) of B. anthracis endospores and it is suitable for direct quantification, even under resource-limited field conditions, where culturing is not an option. Full article
(This article belongs to the Special Issue An Update on Anthrax)
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Open AccessEditorial
Editorial for Special Issue “Natural Alternatives against Bacterial Foodborne Pathogens”
Microorganisms 2020, 8(5), 762; https://doi.org/10.3390/microorganisms8050762 - 20 May 2020
Viewed by 406
Abstract
In recent years, increased resistance to antibiotics and disinfectants from foodborne bacterial pathogens has become a relevant consumer health issue and a growing concern for food safety authorities [...] Full article
(This article belongs to the Special Issue Natural Alternatives against Bacterial Foodborne Pathogens)
Open AccessArticle
Assessment of the Presence of Hepatitis E Virus in Surface Water and Drinking Water in Portugal
Microorganisms 2020, 8(5), 761; https://doi.org/10.3390/microorganisms8050761 - 19 May 2020
Viewed by 510
Abstract
Hepatitis E virus (HEV) is a non-enveloped single-stranded positive-sense RNA virus, belonging to the Hepeviridae family, resistant to environmental conditions, and transmitted by the consumption of contaminated water. This virus is responsible for both sporadic and epidemic outbreaks, leading to thousands of infections [...] Read more.
Hepatitis E virus (HEV) is a non-enveloped single-stranded positive-sense RNA virus, belonging to the Hepeviridae family, resistant to environmental conditions, and transmitted by the consumption of contaminated water. This virus is responsible for both sporadic and epidemic outbreaks, leading to thousands of infections per year in several countries, and is thus considered an emerging disease in Europe and Asia. This study refers to a survey in Portugal during 2019, targeting the detection and eventual quantification of enteric viruses in samples from surface and drinking water. Samples positive for HEV RNA were recurrently found by reverse transcription quantitative PCR (RT-qPCR), in both types of matrix. The infectivity of these samples was evaluated in cultured Vero E6 cells and RNA from putative viruses produced in cultures evidencing cytopathic effects and was subjected to RT-qPCR targeting HEV genomic RNA. Our results evidenced the existence of samples positive either for HEV RNA (77.8% in surface water and 66.7% in drinking water) or for infectious HEV (23.0% in surface water and 27.7% in drinking water). These results highlight the need for effective virological control of water for human consumption and activities. Full article
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