Search Results (127)

Pulmonary nocardiosis (PN) is a rare, opportunistic, and potentially life-threatening infection, especially in disseminated cases. This retrospective study aimed to characterize the clinical features of PN and assess the diagnostic utility of metagenomic next-generation sequencing (mNGS). We reviewed data from 19 patients diagnosed with PN between September 2019 and August 2022, including 3 with disseminated disease. Common symptoms included fever, cough, and sputum production, while chest imaging frequently revealed nodules, consolidations, exudates, cavities, and pleural effusions. The sensitivity of mNGS for detecting Nocardia was significantly higher than that of culture (100% vs. 36.84%, p < 0.001). mNGS successfully identified Nocardia species and co-infected pathogens. The most common species was Nocardia farcinica. Four PN cases were co-infected with Rhizomucor pusillus, Cryptococcus neoformans, Lichtheimia ramosa, and Aspergillus spp. Eighteen patients (94.7%) received trimethoprim-sulfamethoxazole (TMP-SMZ). Sixteen cases (84.2%) were improved or cured. Misdiagnosis is common due to the nonspecificity of clinical and imaging presentations of pulmonary nocardiosis. The timely combination of mNGS represents a promising approach to enhance the diagnosis of pulmonary nocardiosis and inform targeted antimicrobial therapy. TMP-SMZ is the first line of treatment. Full article

Human metapneumovirus (hMPV), a member of the Pneumoviridae subfamily, has emerged as a significant etiological agent of acute respiratory tract infections across diverse age groups, particularly affecting infants, the elderly, and immunocompromised individuals. Since its initial identification in 2001, hMPV has been recognized globally for its seasonal circulation pattern, predominantly in late winter and spring. hMPV is a leading etiological agent, accounting for approximately 5% to 10% of hospitalizations among pediatric patients with acute respiratory tract infections. hMPV infection can result in severe bronchiolitis and pneumonia, particularly in young children, with clinical manifestations often indistinguishable from those caused by human RSV. Primary hMPV infection typically occurs during early childhood; however, re-infections are frequent and may occur throughout an individual’s lifetime. hMPV is an enveloped, negative-sense RNA virus transmitted through respiratory droplets and aerosols, with a 3–5-day incubation period. The host immune response is marked by elevated pro-inflammatory cytokines, which contribute to disease severity. Advances in molecular diagnostics, particularly reverse transcription–quantitative polymerase chain reaction (RT-qPCR) and metagenomic next-generation sequencing (mNGS), have improved detection accuracy and efficiency. Despite these advancements, treatment remains largely supportive, as no specific antiviral therapy has yet been approved. Promising developments in vaccine research, including mRNA-based candidates, are currently undergoing clinical evaluation. This review synthesizes current knowledge on hMPV, highlighting its virological, epidemiological, and clinical characteristics, along with diagnostic advancements and emerging therapeutic strategies, while underscoring the critical role of continued research and sustained preventive measures—including vaccines, monoclonal antibodies, and non-pharmaceutical interventions—in mitigating the global burden of hMPV-related disease. Full article

Outbreaks of infectious diseases contribute significantly to morbidity and mortality in resource-limited settings, yet the capacity to identify their etiology remains limited. We aimed to characterize microbes and antimicrobial resistance (AMR) genes in Tanzanian children affected by an acute febrile illness (AFI) outbreak using metagenomic next-generation sequencing (mNGS). A cross-sectional study was conducted on archived blood samples from children who presented with AFI between 2018 and 2019. Total nucleic acids were extracted from 200 µL of blood, and complementary DNA (cDNA), along with enriched pathogenic DNA, was sequenced using the Illumina MiSeq platform. mNGS data were analyzed using CZ-ID Illumina mNGS bioinformatics pipeline v7.0. Results were obtained from 25 participants (mean age: 11.6 years; SD ± 5), of whom 36% had a moderate to high-grade fever. The following five potential microbial causes of AFI were identified: Escherichia coli (n = 19), Paraclostridium bifermentans (n = 2), Pegivirus C (n = 2), Shigella flexneri (n = 1) and Pseudomonas fluorescens (n = 1), with E. coli being the most prevalent. Twelve AMR genes were detected, including mdtC, acrF, mdtF, and emrB. E. coli harbored most of the AMR genes previously associated with resistance to commonly used antibiotics. mNGS offers a promising complementary approach to conventional diagnostics for identifying pathogens and AMR profiles in vulnerable populations. Full article

Background: Whipple’s disease (WD) is a rare infection caused by Tropheryma whipplei. Diagnosis is challenging and requires a combination of several data sets, such as patient history, clinical and laboratory investigations, and endoscopy with histology analyses. While persistent diarrhea is a common symptom, WD can affect multiple organs. Case description: We present the case of a 66-year-old immunocompetent patient with WD and a history of Helicobacter pylori infection who developed chronic diarrhea. Colonoscopy and histopathological analysis revealed the presence of foamy macrophages with periodic acid-Schiff-positive particles. Subsequently, molecular methods confirmed the clinical WD diagnosis and metagenomic analyses further identified a co-infection with Giardia intestinalis. The patient fully recovered after 14 months of antibiotic therapy. During pharmacological treatment, clinical and laboratory follow-ups were conducted at 6 and 12 months, and microbiome profiles were also analyzed to identify the most abundant species in the samples. Conclusion: The metagenomic analyses showed the eradication of the two pathogens and a progressive restoration to a healthy/balanced status after antibiotic therapy. Full article

Background: Infectious uveitis is a potentially sight-threatening condition that requires timely and accurate pathogen identification to guide effective therapy. However, conventional microbiological tests (CMTs) often lack sensitivity and the inclusiveness of pathogen detection. Metagenomic next-generation sequencing (mNGS) offers an unbiased approach to detecting a broad range of pathogens. This review evaluates its diagnostic performance in detecting infectious uveitis. Methods: A systematic search across multiple databases identified studies assessing the use of mNGS for diagnosing infectious uveitis. The included studies compared mNGS to CMTs, including polymerase chain reaction (PCR), culture, serology, and the IGRA (Interferon-Gamma Release Assay). The study characteristics; the detection rates; and the sensitivity, specificity, and predictive values were extracted. The sensitivity and specificity of mNGS were calculated using CMTs as a reference. Results: Twelve studies comprising 859 patients were included. The sensitivity of mNGS compared to that of CMTs ranged from 38.4% to 100%, while specificity varied between 15.8% and 100%. The commonly detected pathogens included varicella-zoster virus, cytomegalovirus, Toxoplasma gondii, and herpes simplex virus. In some cases, mNGS outperformed PCR in viral detection, aiding diagnosis when the standard methods failed. However, contamination risks and inconsistent diagnostic thresholds were noted. Conclusions: mNGS enables the diagnosis of infectious uveitis, particularly for viral causes, but its variable performance and standardization challenges warrant further investigation. Full article

Zoonotic diseases present a growing public health challenge, particularly in sub-Saharan Africa (SSA) due to close interactions between humans and animals and poor diagnostic capacity. This pilot study investigated human exposure to zoonotic pathogens in Zambia among 47 suspected COVID-19 patients from whom nasopharyngeal samples were collected between November 2020 and February 2021 at two major COVID-19 referral centers in Lusaka. Using metagenomic next-generation sequencing (mNGS), the study identified a diverse range of pathogens, including bacterial, fungal, viral, and parasitic species. The prevalence of zoonotic pathogens was 57.4%. Noteworthy zoonoses included Bacillus anthracis, Sporothrix schenckii, Listeria monocytogenes, Yersinia pestis, Streptococcus suis, Vibrio parahaemolyticus, Brucella melitensis, Rickettsia prowazekii, Shewanella algae, Rickettsia japonica, Coxiella burnetii, Leptospira borgpetersenii, Erysipelothrix rhusiopathiae, Brucella abortus, Bartonella quintana, Banna virus, Vibrio alginolyticus, Bartonella clarridgeiae, Rickettsia canadensis, Leishmania braziliensis, Trypanosoma brucei, Pasteurella multocida, and Arcobacter butzleri. Despite moderate diversity in the microbial community, no significant demographic or health-related factors, including age, gender, or comorbidities such as HIV, were found to be statistically associated with zoonotic pathogen infection. The findings provide valuable data on the presence of zoonotic pathogens in humans in Zambia and highlight the need for more comprehensive research into zoonotic diseases in both clinical and non-clinical settings. Full article

Metagenomic next-generation sequencing (mNGS) has emerged as a revolutionary tool for infectious disease diagnostics. The necessity of mNGS in real-world clinical practice for common Lower Respiratory Tract Infections (LRTI) needs further evaluation. A total of 184 bronchoalveolar lavage fluid (BALF) samples and 322 sputa associated with LRTI were fully examined. The detection performance was compared between mNGS and standard microbiology culture, using Sanger sequencing as the reference method. 52.05% (165/317) of sputa showed identical results for all three methods. Compared to Sanger sequencing, the same results obtained by mNGS were 88.20% (284/322). In 2.80% (9/322) of cases, Sanger sequencing detected more microorganisms, while mNGS detected more in 9% (29/322) of cases. For BALF, 49.41% (85/172) of cases showed identical results for all three methods. In 91.30% (168/184) of cases, identical results were produced by both mNGS and Sanger sequencing. mNGS detected more species in 7.61% (14/184) of cases, whereas in 2.80% (2/184) instances, the Sanger sequencing detected more microorganisms than mNGS. In the 184 BALF samples, 66 samples were identified as having co-infections by mNGS, Sanger sequencing identified 64 samples, and cultures identified 22 samples. Our study demonstrates that mNGS offers a significant advantage over conventional culture methods in detecting co-infections. For common bacterial pathogens, conventional culture methods are sufficient for detection. However, mNGS provides comprehensive pathogen detection and is particularly useful for identifying rare and difficult-to-culture pathogens. Full article

Hybrid capture-based target enrichment prior to sequencing has been shown to significantly improve the sensitivity of detection for genetic regions of interest. In the context of One Health relevant pathogen detection, we present a hybrid capture-based sequencing method that employs an optimized probe set consisting of 149,990 probes, targeting 663 viruses associated with humans and animals. The detection performance was initially assessed using viral reference materials in a background of human nucleic acids. Compared to standard metagenomic next-generation sequencing (mNGS), our method achieved substantial read enrichment, with increases ranging from 143- to 1126-fold, and enhanced detection sensitivity by lowering the limit of detection (LoD) from 10³–10⁴ copies to as few as 10 copies based on whole genomes. This method was further validated using infectious samples from both animals and humans, including bovine rectal swabs and throat swabs from SARS-CoV-2 patients across various concentration gradients. In both sample types, our hybrid capture-based sequencing method exhibited heightened sensitivity, increased viral genome coverage, and more comprehensive viral identification and characterization. Our method bridges a critical divide between diagnostic detection and genomic surveillance. These findings illustrate that our hybrid capture-based sequencing method can effectively enhance sensitivity to as few as 10 viral copies and genome coverage to >99% in medium-to-high viral loads. This dual capability is particularly impactful for emerging pathogens like SARS-CoV-2, where early detection and genomic characterization are equally vital, thereby addressing the limitations of metagenomics in the surveillance of emerging infectious diseases in complex samples. Full article

The semen microbiome, once believed to be sterile, is now recognized as a dynamic ecosystem containing a diverse range of microorganisms with potential implications for male fertility and reproductive health. We aimed to examine the relationship between the semen microbiome, male infertility, and reproductive outcomes, highlighting the transformative role of next generation sequencing techniques and bioinformatics in exploring this intricate interaction, and we present a critical review of the published literature on this issue. Current evidence suggests a complex association between the composition of the semen microbiome and male fertility, with certain bacterial genera, such as Lactobacillus and Prevotella that exert opposing effects on sperm quality and DNA integrity. In addition, the influence of the semen microbiome extends beyond natural fertility, affecting assisted reproductive technologies and pregnancy outcomes. Despite considerable progress, challenges remain in standardizing methodologies and interpreting findings. In conclusion, we identify the lack of a definitive management proposal for couples presenting with this phenomenon, and we underline the need for an algorithm and indicate the questions raised that point toward our goal for a strategy. Continued research is essential to clarify the role of the semen microbiome in male reproductive health and to advance the development of personalized fertility management approaches. Full article

Modern agriculture should limit its degrading impact on the soils, the natural environment, and the climate. No-tillage soil cultivation technologies, which have been in use for many years and are constantly being improved, are a good example of these actions; although, in-depth studies on their impact on the soil microbial community are currently scarce. The aim of our study was to evaluate the effect of cultivation technology on the soil bacterial community to assess differences that can be reflected in the environmental and agricultural functionality, identifying possible bacterial species with ecological properties. In this context, the composition of bacterial communities (at the phyla, order, class, and species levels) was evaluated under different conditions, such as conventional tillage (CT) (plophing), reduced tillage (RT) (stubble cultivator), strip tillage (ST), and no-tillage (direct sowing on stubble and fallow buffer zone of the experimental field), in a horse bean plantation. Metagenomic methods (next generation sequencing technology, NGS) were used to determine the percentage of individual operational taxonomic units (OTUs). Our study showed that no-tillage cultivation technologies, mainly strip and no-tillage methods, had a positive effect on microbiological communities. In fact, key species related to soil fertility and crop yield, such as Gemmatimonas aurantiaca (a microorganism that reduce nitrous oxide, N₂O in soil) and Aeromicrobium ponti (a beneficial species for the soil environment, essential for the proper functioning of the crop agroecosystem), increased in reduced cultivation technologies. These species can determine soil fertility and crop yields, and therefore, they are very important for sustainable and even regenerative agriculture. Further studies of soil samples collected from other crop plantations under different cropping systems may indicate beneficial microbial species that are important for soil fertility. Full article

Agricultural practices significantly impact soil properties and ecological functions, highlighting the importance of comprehensive soil health assessments. Traditionally, these assessments have focused on physical and chemical indicators, often neglecting microbiological properties. This review explores the potential of microbiological indicators in evaluating the effects of agricultural practices on soil ecological functions, emphasizing their significance and addressing challenges associated with their application. A key advantage of microbiological indicators is their high sensitivity and rapid response to environmental changes. These indicators can be grouped into three categories: microbial biomass and abundance, microbial taxonomic composition and diversity, and microbial activity. Among these, microbial biomass carbon, basal respiration, and decomposition rates are considered the most reliable and interpretable indicators. Microbial taxonomic composition and diversity remain limited in their diagnostic and predictive capabilities due to challenges in interpretation. Integrating microbiological indicators offers a more holistic understanding of the interactions between agricultural practices and soil health, enhancing our ability to monitor, manage, and preserve soil ecosystems. To facilitate their adoption in agricultural production and land management, further efforts are needed to improve the interpretability of these indicators and to establish standardized criteria for soil health assessment. Full article

Introduction: Nosocomial lower respiratory tract infections (nLRTIs), including hospital-acquired pneumonia (HAP) and ventilator-associated pneumonia (VAP), remain significant challenges due to high mortality, morbidity, and healthcare costs. Implementing accurate and timely diagnostic strategies is pivotal for guiding optimized antimicrobial therapy and addressing the growing threat of antimicrobial resistance. Areas Covered: This review examines emerging microbiological diagnostic methods for nLRTIs. Although widely utilized, traditional culture-based techniques are hindered by prolonged processing times, limiting their clinical utility in timely decision-making. Advanced molecular tools, such as real-time PCR and multiplex PCR, allow rapid pathogen identification but are constrained by predefined panels. Metagenomic next-generation sequencing (mNGS) provides comprehensive pathogen detection and resistance profiling yet faces cost, complexity, and interpretation challenges. Non-invasive methods, including exhaled breath analysis using electronic nose (e-nose) technology, gene expression profiling, and biomarker detection, hold promise for rapid and bedside diagnostics but require further validation to establish clinical applicability. Expert Opinion: Integrating molecular, metagenomic, biomarker-associated, and traditional diagnostics is essential for overcoming limitations. Continued technological refinements and cost reductions will enable broader clinical implementation. These innovations promise to enhance diagnostic accuracy, facilitate targeted therapy, and improve patient outcomes while contributing to global efforts to mitigate antimicrobial resistance. Full article

Fish is an essential lean protein source worldwide. Unfortunately, fresh fish food products deteriorate rapidly due to microbial spoilage. With consumers’ growing concerns about using chemical preservatives, we propose using natural preservatives as safer alternatives to prevent microbial spoilage. In this study, we used Next-Generation Sequencing (NGS) metagenomics to study microbiomes on catfish fillets at early (day one for all samples), middle (day seven for control store-bought and aquaculture-raised samples, day nine for other treatment store-bought samples, and day eleven for other treatment aquaculture-raised samples), and late (day fifteen for all store-bought, day eleven for control aquaculture-raised samples, and day twenty-seven for other treatment aquaculture-raised samples) points. Store-bought and aquaculture-raised catfish were treated individually with natural preservatives (vinegar, lemon, and grapefruit seed [GSE]). We observed bacterial populations and sequenced 16S NGS libraries of catfish microbes. Vinegar treatment showed the greatest suppression of bacterial growth in both groups, and GSE and lemon treatment had similar levels of suppression in the mid and late points (−4 to −5 Log CFU/g vinegar and −0.1 to −4 Log CFU/g other treatments in aquaculture and −1 to −2 Log CFU/g vinegar and −0.2 to −0.5 Log CFU/g other treatments in store-bought). Aquaculture-raised vinegar treatment samples had similar proportional taxonomy abundance values through storage duration. Pseudomonas, Janthinobacterium, and Camobacteriaceae were the dominant bacteria species in the early point for store-bought fish. Still, Pseudomonas was suppressed by vinegar treatment in the middle point, which allowed for less biased relative abundance compared to other treatments. Chryseobacterium, CK-1C4-19, and Cetobacterium were the dominant bacteria species for early point treatments in aquaculture-raised fish. Still, they remained the predominant bacteria for only aquaculture-raised vinegar samples in the middle and late points, which allowed for a similar relative abundance to fresh catfish. Meanwhile, Pseudomonas in most lemon and GSE samples became the dominant species at a later point. This study provides a better understanding of bacterial spoilage of catfish during storage. Additionally, we showed that natural preservative treatments can effectively extend the shelf-life of fishery products. Full article

Among the various causes of rhomboencephalitis, Listeria monocytogenes infection is the most common. However, conventional microbiological methods often yield negative results, making diagnosis challenging and leading to extensive, often inconclusive, diagnostics. Advanced molecular techniques like metagenomic next-generation sequencing (mNGS) offer a powerful and efficient approach to pathogen identification. We present a case of life-threatening rhomboencephalitis in a 32-year-old immunocompetent patient where extensive microbiological, immunological, and biochemical tests were inconclusive. Given the patient’s consumption of unpasteurized homemade cheese, neurolisteriosis was suspected, and mNGS was employed on clinical samples (CSF, serum, urine) and the food source to identify the pathogen. mNGS detected L. monocytogenes in both patient samples and the cheese. Mapping reads were distributed across the genome, with 18.9% coverage in clinical samples and 11.8% in the cheese sample. Additionally, the Listeriolysin (hlyA) gene was detected with 22.3% coverage in clinical samples and 12.3% in the food source, confirming neurolisteriosis. The patient fully recovered following antibiotic treatment. This case underscores the importance of mNGS in diagnosing CNS infections when conventional methods yield negative results, and supports its inclusion in diagnostic protocols for suspected neurolisteriosis, particularly when traditional methods prove inadequate. Full article

Background: For investigating the host response in Acinetobacter baumannii associated pneumonia, we analyzed the host genetic sequences obtained from metagenomic next-generation sequencing (mNGS). Methods: The samples for mNGS were bronchoalveolar lavage fluid (BALF) collected from the lungs of patients infected with A. baumannii and from patients without bacterial infections. BALF samples from patients with pneumonia were collected from the lungs of patients infected with A. baumannii with New Delhi metallo-β-lactamase (NDM, before treatment), A. baumannii with NDM (post-treatment), A. baumannii without resistant genes, and those without bacterial infection. Partek was used for investigating enriched functions and pathways related to the pulmonary host response to pneumonia caused by A. baumannii with NDM infection and A. baumannii without antimicrobial-resistant genes. The STRING was employed for identifying protein interaction pathways related to the pulmonary host response to pneumonia caused by A. baumannii without antimicrobial-resistant genes. Results: In pulmonary host response to pneumonia caused by A. baumannii with NDM, five immune system-related pathways and five pathways related to signal transduction were identified. No significant differences were observed in the immune system and signal transduction pathways in the pulmonary host response to pneumonia caused by A. baumannii without antimicrobial-resistant genes. However, significant differences were noted in the phagosome, ferroptosis, and regulation of the actin cytoskeleton in cellular processes. Conclusions: mNGS provides information not only on pathogen gene expression but also on host gene expression. In this study, we found that pneumonia with A. baumannii carrying the NDM resistance gene triggers stronger immune responses in the lung, while pneumonia with A. baumannii lacking antimicrobial resistance genes is more linked to iron-related pathways. Full article