SEARCH · Search bioRxiv

Results for “Microbiology”

Search indexed bioRxiv preprints in genomics, neuroscience, cell biology and bioinformatics. Read source abstracts and check manuscript versions; preprints are not peer reviewed.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 685 records · Page 38Linked to original sources

bioRxiv2026

Enrichments along gradients resolve eco-evolutionary forces on subsurface microbiomes

How a single gram of soil harbors billions of microorganisms, each with distinct genomic variants that collectively maintain coherent ecological function(s), is one of microbiologys grand unsolved problems. A key obstacle is determining which variants contribute to individual- and community-level fitness, in which contexts, and how co-occurring ecotypes interact to divide niche space. Here, using nitrate (NO3-)-contaminated subsurface sediment as inoculum, we have performed high throughput enrichments in laboratory media of defined carbon source compositions across ecologically relevant gradients of pH and NO3-. Long-read metagenomics and link-community decomposition of co-occurrence networks of taxa across these enrichments has revealed context-specific functional interactions among dominant generalist and lower-abundance specialist denitrifier ecotypes that comprise 53 distinct enriched communities (EnComs) across 288 enrichments derived from a single sediment sample. We identified a single enzymatic difference of alternative NO3- reductases (NapAB vs. NarGHI) with differing substrate affinities that provided a mechanistic explanation for competitive niche partitioning between the two dominant taxa, Neorhizobium spp. and Allorhizobium spp., along the NO3- gradient. Genome-wide polymorphism ratios (pN/pS) revealed that selective pressures vary systematically with carbon source availability and gradients of pH and NO3-, which helps explain the natural biodiversity and functional interactions of ecotypes within denitrifying communities in the subsurface sediment. Our findings show that controlled enrichments along ecological gradients can thus uncover eco-evolutionary forces of selection, drift, and diversification that sculpt the biodiversity of microbial populations in the natural environment.

microbiology↗

bioRxiv2026

Statistical learning of bacterial growth in combinatorially constructed environments

Optimizing growth conditions and culture media is a major goal in microbiology. A challenge is that nutrients can have complex, non-additive effects on growth. The fact that resources interact with one another has long been known in ecology, but systematic maps of resource interactions at all orders have been lacking and it is not known whether interactions are primarily low order or fundamentally complex. To tackle this problem, we have followed a full factorial design approach and measured the growth of seven different bacterial species in all possible combinations of 8 carbon sources under carbon-limiting conditions. Our approach allows us to directly estimate interactions at all orders. Even though all C-sources stimulate growth on their own as well as in combination with other nutrients, most of them can also have negative effects on growth when they are added to at least some nutrient mixtures. We show that the switch from positive to negative fitness effects is governed by global epistasis among resources. An analysis of variance shows that additive effects and pairwise interactions explain most of the variation in fitness, allowing us to train simple regression models that accurately predict bacterial growth in novel environments. The generality of these findings across seven different bacterial strains belonging to two different families indicates that interactions between carbon sources under carbon-limiting conditions may be generally learnable from a relatively sparse set of constructed environments, enabling the rational optimization of growth conditions.

microbiology↗

bioRxiv2026

Epistasis limits but does not prevent the transfer of mutation-drug resistance mapping across 600 million years of fungal evolution

Whether different pathogens acquire resistance to antimicrobials through the same mutations is a major question in evolution and microbiology. Most antifungal drugs are used to treat infections caused by multiple fungal species, many of which have diverged for millions of years. If the evolution of resistance was to converge onto the same set of mutations across species, knowing the mechanism of resistance in one would allow us to predict and track it in others. The extent of this convergence remains unknown due to the lack of systematic data on resistance mutations. Here, we quantify the conservation of resistance mutations in the cytosine deaminase (CD), a protein responsible for resistance to flucytosine, one of the oldest antifungal drugs. By comparing the crystal structures of this enzyme through 600 My of evolution, we show that the CD structure is highly conserved. We compared the full CD mutational resistance spectrum of resistance from an ascomycete and a basidiomycete. We found that resistance mutations in one ortholog can be used to predict resistance in the other at a high level of accuracy. However, because of epistasis, around 10% of mutations have distinct effects in the two orthologs, which imposes an upper limit to the transferability of the knowledge of resistance mutations from one species to another. Using biochemical assays and by structural characterization of several mutants, we identify distinct mechanisms of epistasis, one important being that the local physiochemical environment of some position has evolved in a way that makes the same substitutions destabilizing or entirely inactivating in an ortholog-specific manner. Our results show that resistance mutations can be conserved in fungi across hundreds of millions of years of evolution but that epistasis eventually limits the accuracy of these predictions.

microbiology↗

bioRxiv2026

Lytic bacteriophages against Salmonella Typhi as a potential alternative to antibiotics

IntroductionEnteric fever, being endemic with seasonal peaks in low- and middle-income countries, is a major health concern. Moreover, the rise in antibiotic resistance has exacerbated the situation. This study was undertaken to investigate the lytic bacteriophages against Salmonella Typhi with a potential for phage therapy. Materials and MethodsA hospital-based cross-sectional study was conducted from October 2023 to March 2024. Blood cultures were processed by the BACTEC automated culture system following standard microbiological techniques to isolate typhoidal Salmonella. Antibiotic susceptibility was tested by the modified Kirby-Bauer disc diffusion method. Lytic bacteriophages isolated by the double-layer agar method were assessed for their host range and lytic ability with spot and turbidimetric assays. ResultsOf the total 1054 blood specimens, 35 (3.2%) were positive for S. Typhi. All the isolates were susceptible to first-line antibiotics--ampicillin, chloramphenicol, and cotrimoxazole. The isolates were also sensitive to nalidixic acid (80%) as well as fluoroquinolones; ciprofloxacin (62.86%), levofloxacin (77.14%), and ofloxacin (80%). Fifteen lytic phages were isolated against S. Typhi Ty2 and CT18 strains. Four phages--vB_SaTy_ST2, vB_SaTy_ST7, vB_SaTy_ST17, and vB_SaTy_ST18--lysed all 35 clinical S. Typhi isolates. While vB_SaTy_ST17 and vB_SaTy_ST18 also lysed 7 out of 20 S. Paratyphi A isolates. Three phages (vB_SaTy_ST2, vB_SaTy_ST7, vB_SaTy_ST17) were tested against S. Typhi isolate S30. Individually, vB_SaTy_ST17 suppressed the growth for 13 hours, vB_SaTy_ST2 and vB_SaTy_ST7 for 10 hours. The phage cocktail vB_SaTy_ST2 + vB_SaTy_ST17 was the most effective, which extended the inhibition time to 15 hours. ConclusionThis study highlights the ongoing burden of enteric fever in Nepal and the increase in susceptibility of S. Typhi to nalidixic acid and fluoroquinolones. It also demonstrates the promising lytic potential of bacteriophages, particularly vB_SaTy_ST17 and the phage cocktail vB_SaTy_ST2 + vB_SaTy_ST17, against clinical S. Typhi, highlighting their potential as alternatives to antibiotics.

microbiology↗

bioRxiv2026

MTB-LysB1: A Novel Endolysin Against Multidrug-resistant Mycobacterium tuberculosis

The phenotypic plasticity, slow replication, and complex, hydrophobic cell envelope of Mycobacterium tuberculosis contribute to its successful survival as a pathogen and its drug tolerance. Consequently, the global threat of multidrug-resistant Tuberculosis (MDR-TB), coupled with lengthy and highly toxic treatment regimens, necessitates the development of innovative treatment solutions. Mycobacteriophages are natural viruses of mycobacteria that typically encode two endolysins, which cooperatively facilitate host cell lysis at the end of the lytic life cycle: LysA, a peptidoglycan hydrolase, and LysB, a lipolytic enzyme, targeting the mycolylarabinogalactan-peptidoglycan complex. Their precise and efficient lytic activity, along with their low propensity to induce resistance, make them, particularly LysBs, promising candidates for new treatment solutions. In this study, we report MTB-LysB1, a novel LysB enzyme from an F1 sub-cluster mycobacteriophage isolated from our laboratory collection. While studying its structural features by comparing the modelled structure with representative mycobacteriophage LysB homologues, we found that the /{beta}-hydrolase fold and key motifs are conserved. Also, we identified putative membrane-interaction motifs that may play a role in LysB1s cell permeation. Significantly, we found MTB-LysB1 to be active against both drug-susceptible and multidrug-resistant (MDR) M. tuberculosis strains at nanomolar concentrations, comparable to the well-characterised D29 LysB reference enzyme. Beyond its standalone activity, MTB-LysB1 exhibits an additive effect when combined with the TB drugs rifampicin and moxifloxacin, and co-administration reduces the drugs minimum inhibitory concentrations (MICs), which holds clinical significance. By structurally damaging the mycobacterial cell wall, the enzyme appears to act as a permeability enhancer for the chemotherapeutic drugs, thereby improving antibiotic efficacy. Collectively, our findings position the enzyme not only as a novel antimycobacterial agent but also provide a structural framework for its rational engineering as a promising next-generation adjunct to TB drug regimens. HighlightsO_LIA novel F1 sub-cluster phage-derived LysB is discovered and characterised using integrated computational, biochemical and microbiological methods. C_LIO_LIAlphaFold2 modelling, molecular dynamics simulations and comparative structural analyses revealed an /{beta}-hydrolase fold with conserved catalytic and membrane-interaction features. C_LIO_LIThe enzyme exhibited high esterase activity, thermal stability and potent lytic activity against Mycobacterium tuberculosis. C_LIO_LIAn additive effect with TB drugs rifampicin and moxifloxacin highlights MTB-LysB1s potential as an adjunct therapeutic. C_LI

microbiology↗

bioRxiv2026

No-take marine reserves promote oligotrophic reef bacterioplankton communities across the Great Barrier Reef

Australias Great Barrier Reef is a biodiversity hotspot critical to ocean health, yet it faces increasing threats from climate change and localised impacts requiring effective conservation and management action. Rezoning of the Great Barrier Reef Marine Park in 2004 expanded No-Take Marine Reserves (NTMRs) to restrict extractive activities like fishing and collecting, creating one of the largest networks of marine reserves globally. Benefits like increased biomass of fisheries-targeted species and improved coral community health metrics have been reported, though the effects of zoning on water chemistry and seawater microbiology remain unexplored. Using data from the Great Barrier Reef Microbial Genomics Database, we investigated the structure of seawater microbiomes on 48 offshore reefs within NTMRs and fished reefs. A supervised classification method (MINT sPLS-DA) identified 350 indicator species that predict zoning with [~]71% accuracy (range 58-85%). Microbial communities broadly reflected reef states, with NTMR zones enriched in streamlined microbial oligotrophs (Pelagibacter and SAR86) correlating with higher cover of hard coral, crustose coralline algae, and herbivore fish abundance under lower nutrient conditions. By contrast, fished reefs harbored opportunists (Flavobacteriales, especially UA16, and Pseudomonadales) associating with elevated nutrients and turf algae cover. Co-occurrence networks revealed stronger competitive interactions in fished reefs, where nutrient-responsive taxa may outcompete other microbes, underscoring the need to investigate how these shifts influence reef nutrient cycling and function. Our findings reveal ecosystem-wide effects of marine zoning beyond fish protection, with distinct seawater microbiomes between fished reefs and NTMRs, which will help build decision tools for more targeted reef health monitoring assessments.

microbiology↗

bioRxiv2026

Comparative Evaluation of Commercial Gradient Diffusion and Disk Diffusion Tests for Aztreonam-Avibactam Susceptibility Testing

BackgroundAztreonam-avibactam (AZA) is a recently approved agent active against metallo-{beta}-lactamase-producing Enterobacterales. However, independent evaluations of commercially available antimicrobial susceptibility testing (AST) products remain limited. MethodsWe evaluated 56 carbapenem-resistant Enterobacterales isolates using four newly available AZA AST products, including two gradient diffusion (bioMerieux Etest; Liofilchem MIC Test Strip) and two disk diffusion (Hardy, Liofilchem) tests across three Mueller-Hinton agar manufacturers at two independent clinical laboratories. A diverse isolate collection enriched for nonsusceptible and breakpoint-adjacent MICs was used to rigorously assess categorical agreement. Broth microdilution (BMD) was used as the reference method. Essential agreement (EA), categorical agreement (CA), error rates, and reproducibility were analyzed. ResultsAll four commercial AST products demonstrated acceptable analytical performance compared with reference BMD. Gradient diffusion demonstrated high EA (bioMerieux 94.3%, Liofilchem 90.5%), whereas CA ranged from 83.3% to 89.0% across all four products. Minor errors accounted for all categorical discrepancies, and occurred almost exclusively among breakpoint-adjacent isolates. Media-related variability was modest, and quality control performance was generally high, with strong within-site precision and consistent performance across laboratories. ConclusionsThis study provides the first comprehensive head-to-head comparison of commercial AZA susceptibility testing products and supports their implementation in clinical microbiology laboratories.

microbiology↗

bioRxiv2026

Challenges of Dry Sanitization to Control Salmonella Dry Surface Biofilms

Salmonella is a pathogen linked to foodborne outbreaks, including low-moisture foods. Its ability to resist desiccation can contribute to the formation of dry surface biofilms (DSB). This study evaluated the impact of 3 DSB formation protocols (P1=48-h hydrated phase/48-h dry phase, P2=24-h/120-h and P3=8-h/48-h) on the resistance of Salmonella DSB to 70% alcohol, a commercial product (based on 0.015% quaternary ammonium and 25% isopropyl alcohol), gaseous ozone (45 ppm), hot air (90 {degrees}C) and UV-C light (254 nm). The type of DSB protocol impacted the efficacy of the sanitizers (p < 0.05). The biofilm with the shortest hydration phase showed the greatest susceptibility; three out of the five sanitizers evaluated (70% alcohol, commercial product, and UV-C) promoted significant reductions in P3, with counts below the detection limit (0.8 log CFU/cm{superscript 2}) after 5 to 15 min exposure. Regarding protocols P1 and P2, in general, the best performance was from UV-C, especially against DSB on polypropylene, where it achieved reductions of 1.3 log CFU/cm{superscript 2} for P1 and 2.9 log CFU/cm{superscript 2} for P2 after 15 to 30 min of exposure. In contrast, hot air and ozone showed less effectiveness, with reductions [≤]1.2 log CFU/cm{superscript 2}. In most scenarios, confocal microscopy images corroborated the plate count results (log CFU/cm{superscript 2}). In summary, our data indicates limited action of dry sanitizers on Salmonella DSB, requiring validation and optimization of sanitization processes to ensure the microbiological safety of low-moisture products.

microbiology↗

bioRxiv2026

BACTERIAL AND FUNGAL CONTAMINATION OF STAIRCASE BANISTERS AT THE COLLEGE OF SCIENCE, KWAME NKRUMAH UNIVERSITY OF SCIENCE AND TECHNOLOGY, GHANA

Staircase banisters are frequently touched surfaces that may receive microorganisms from hands, dust, air and other environmental sources, but their microbial status in Ghanaian university buildings has received limited attention. This cross-sectional environmental microbiology study assessed bacterial and fungal contamination of staircase banisters at the Kwame Nkrumah University of Science and Technology, Kumasi. Six banisters from the Aboagye Menyah Building Complex, Chemistry Block and Biology Block were purposively selected to include high-traffic locations and both wooden and metal surfaces. Upper and lower sections were sampled over three consecutive Monday afternoons after classes, giving 12 surface samples. Approximately 150 cm{superscript 2} of each section was swabbed with sterile buffered peptone water, cultured on standard bacteriological and mycological media, and analysed using phenotypic and morphological methods. Bacterial loads were compared by independent samples t-test. Thirty-one bacterial isolates were recovered. The study found Gram-positive bacteria which accounted for 74% of isolates and Gram-negative bacteria for 26%. Staphylococcus spp., Streptococcus spp., Enterobacteriaceae, Bacillus spp. and Corynebacterium-+ spp. were the main presumptive bacterial groups. Metal banisters had higher mean bacterial loads than wooden banisters (4.38 {+/-} 0.86 versus 1.24 {+/-} 1.44 log10 CFU/mL; p = 0.014), whereas upper and lower sections did not differ significantly (p = 0.539). Fungal growth was detected in all samples, with Aspergillus fumigatus, Colletotrichum spp. and Aspergillus niger being frequent presumptive fungi. The findings support the routine inclusion of staircase banisters in cleaning and disinfection programmes for academic buildings.

microbiology↗

bioRxiv2026

Isolation, Identification and Antibiogram Assay of Escherichia coli from the Environment of Live Bird Markets in Bangladesh

BackgroundLive bird markets (LBMs) may facilitate the persistence and dissemination of Escherichia coli and antimicrobial-resistant bacteria because of intensive bird handling, environmental contamination and inadequate sanitation. However, information on E. coli contamination and antimicrobial susceptibility in LBM environments in Rajshahi District, Bangladesh, remains limited. ObjectiveThis study aimed to determine the prevalence, identify the cultural and biochemical characteristics, and assess the antimicrobial susceptibility pattern of E. coli isolated from water, soil and bird-dropping samples collected from LBMs in Rajshahi District. MethodsA total of 60 environmental samples, comprising 20 water, 20 soil and 20 bird-dropping samples, were collected from LBMs across all ten upazillas of Rajshahi District between January and June 2023. E. coli was isolated and identified using cultural characteristics, Gram staining and biochemical tests. Antimicrobial susceptibility was determined by the Kirby- Bauer disc diffusion method against seven antimicrobial agents using CLSI interpretive criteria. ResultsE. coli was detected in 33 of 60 samples, giving an overall prevalence of 55.00%. Prevalence was highest in bird-dropping samples (75.00%), followed by water (55.00%) and soil (35.00%). Among the 33 isolates, resistance was highest to oxytetracycline (78.79%) and amoxicillin (63.64%), followed by ciprofloxacin (48.48%), doxycycline (33.33%), levofloxacin (9.09%), erythromycin (9.09%) and neomycin (6.06%). Sensitivity was highest to neomycin (60.61%), followed by levofloxacin and erythromycin (51.51% each). ConclusionThe high prevalence of E. coli and substantial resistance to several commonly used antimicrobials indicate considerable microbiological and antimicrobial-resistance concerns in LBM environments. Improved sanitation, biosecurity, hygienic poultry handling and prudent antimicrobial use are warranted to reduce environmental contamination and potential transmission of resistant bacteria.

microbiology↗

bioRxiv2026

Environmental and spatiotemporal drivers of marine microbial communities from Antarctic and Subantarctic water masses

The Southern Ocean harbors diverse marine microbial communities shaped by both local oceanographic conditions and dispersal limitations. However, this knowledge is mainly based on coastal Antarctic sites, whereas circumpolar Antarctic open sea and subantarctic ecosystems remain poorly explored. Here, we characterize marine microbial communities (using 16S rDNA high-throughput sequencing) and marine oceanographic data across two regions: the Subantarctic, involving two localities (the Magellan Strait and the Beagle Channel), and Antarctic open sea, involving two localities (Eastern Indian and Central South Pacific). We found extensive differences across regions and localities, characterized by distinct taxonomic patterns, alpha diversity, microbial composition, and enriched taxa profiles. Despite these differences, Clade Ia, Amylibacter, NS5 marine group, and NS2b marine group exhibited high prevalence across regions. Oceanographic parameters had variable relationships with microbial alpha diversity across regions: Sea surface temperature and salinity had a negative and positive correlation, respectively, in the Magellan Strait during 2024. In the Antarctic region, dissolved oxygen displayed a negative correlation in the Indian Ocean during 2024, whereas salinity displayed a more variable relationship in the Indian Ocean: positively correlated during 2024, while negatively correlated during 2025. Collectively, our results highlight a strong microbiological biogeographic structure in the Southern Ocean, both across broad scales (between Subantarctic and Antarctic regions) and within regions. Furthermore, our results show dynamic relationships between oceanographic variables and marine microbial diversity across Antarctic and Subantarctic regions.

microbiology↗

bioRxiv2026

Isolation and Characterization of Bacteriocin-Producing Lactic Acid Bacteria from Cheese and Functional Evaluation of Their Synthesized Bioactive Peptides

Biopreservatives including nisin and its derivatives are becoming more desirable in the food processing industry because of the growing demand for naturally preserved and minimally processed foods free from artificial preservatives. However, ensuring microbiological safety while meeting these consumer preferences remains a major challenge. This has necessitated the continuous investigation of potential new antimicrobial agents produced by naturally occurring microorganisms. Hence, this study explored the synthesis, characterisation, and optimisation of a bacteriocinogenic lactic acid bacterium and its antimicrobial product, possibly novel bacteriocin (Nisin 2A) from Lactococcus lactis isolated from commercial brined cheese. The isolation was achieved by screening for wild-type bacteriocin-producing lactic acid bacteria from dairy products using MRS media. Screening was performed using antagonism assays, yielding five producer organisms. Of these, the isolate whose metabolites exhibited the most potent antimicrobial activity was identified as Lactococcus lactis, which synthesised an active antimicrobial peptide designated as Nisin 2A, with a molecular mass of approximately 3.3 kDa as determined by UHPLC-MS and SDS-PAGE. Production of Nisin 2A was scaled up through fed-batch fermentation of Lactococcus lactis in modified MRS broth following process optimisation using a Plackett-Burman experimental design and purified by ammonium sulphate precipitation and solid-phase extraction (SPE). Furthermore, the antimicrobial potential of the bacteriocin was evaluated by the agar well diffusion assay and quantified using the tube dilution method. The purified peptide demonstrated broad-spectrum antimicrobial activity, particularly against the test Gram-positive bacteria Bacillus cereus and retained its bioactivity across a wide pH range (3-9) and high thermal conditions (up to 100 {degrees}C). Furthermore, it had high sensitivity to proteolytic enzymes (Proteinase K and Trypsin). Notably, the peptide was thermostable and retained up to 90% of its initial activity after thermal treatment and maintained consistent inhibitory performance after extended storage. These findings highlight the potential application of Nisin 2A as a natural biopreservative in food systems.

microbiology↗

bioRxiv2026

Heterobenzamides exhibit bacteriostatic activity against intracellular Mycobacterium tuberculosis by targeting aerobic respiration

We previously identified a series of heterobenzamides (HBAs) with potent growth inhibitory activity against Mycobacterium tuberculosis in axenic culture. We also provided evidence that these target QcrB, a component of the terminal cytochrome oxidase in the electron transport chain. We expanded our studies to look at the full microbiological profile: key molecules from the series were tested for activity under different conditions and against additional strains. HBA analogs were active against intracellular bacteria where they exhibited bacteriostatic activity. A strain of M. tuberculosis with a mutation in QcrB (T313I) was resistant to HBAs in both axenic culture and inside macrophages. HBAs retained potency against lineages and mono-resistant strains of M. tuberculosis. HBAs had a narrow spectrum of activity, since they were not active against the ESKAPEE pathogens. Combination of the key HBA with bedaquiline was synergistic, as expected for a QcrB inhibitor, but there was no strong synergy with other drugs. Exposure of M. tuberculosis to the key HBA led to ATP depletion and boosted the oxygen consumption rate. This effect was specific to M. tuberculosis, since human THP-1 macrophage-like cells were unaffected by exposure to the HBA. HBA did not induce the production of reactive oxygen species or affect membrane potential but did affect pH homeostasis. Taken together, these data provide further evidence to support the identification of QcrB as the target and indicate that they are suitable for further drug development.

microbiology↗

bioRxiv2026

Multi-Species, Genome-Wide Metabolic Network Reconstructions Reveal the Basis for Metabolic Versatility in Mycobacteria

The genus Mycobacterium comprises over 200 species, many of which now have complete genome sequences. Some are major pathogens causing diseases like tuberculosis and leprosy, while others are harmless environmental organisms with useful abilities such as degrading pollutants. Environmental mycobacteria are often seen as metabolic generalists, able to utilise a wider range of carbon sources than host-associated species, which are typically more specialised due to their restricted habitats. This metabolic versatility has been proposed to stem from differences in nutrient uptake capabilities rather than catabolic pathways. In order to test this explanation, we developed and validated genome-scale metabolic models for five Mycobacterium species with varying lifestyles and growth rates, creating a computational approach enabled by CarveMe that allows rapid construction of models from genome information. By combining these models with microbiology experiments the study showed that the capacity of the bacteria to transport nutrients into the cell is indeed key to metabolic versatility. We notably found through load-partition experiments that, if a transporter is present but cannot take up its substrate at a rate sufficient for growth, the supply of multiple substrates can mitigate this rate-limiting step. This suggests that mycobacterial species have evolved high-affinity, low-rate systems for nutrient uptake in their ecological niches. More generally, our results demonstrate that a combination of automated annotation methods and straightforward bacterial physiology experiments allow the reconstruction of metabolic models of good predictive quality for hitherto little studied mycobacterial species.

microbiology↗

bioRxiv2026

Phylogenetic Analysis of cpn60 UT region reveal oligonucleotide regions for the detection of prominent Enterobacteriaceae pathogens

The Enterobacteriaceae pathogens like E. coli, Shigella, Klebsiella and Salmonella cause a wide range of gastrointestinal and other mucosal infections when the contaminated food and water are consumed. Conventional microbiological methods are time-consuming and laborious process to detect these pathogens. Molecular assays are fast and sensitive to detect the pathogens. But a potential target for the detection is very essential to develop a molecular assay for the rapid detection. In this study, we explored the cpn60 UT region as a potential target for the detection of significant Enterobacteriaceae members like E. coli, Shigella sp, Salmonella, Klebsiella etc. We did the phylogenetic analysis of cpn60 UT region among the Enterobacteriaceae pathogenic groups. This analysis has revealed the presence of 13 clusters and two oligonucleotide regions with highest nucleotide conservation. Targeting these regions we have designed a set of oligonucleotide primer pairs and developed a SYBR green real time PCR assay to detect the Enterobacteriaceae pathogenic members. The degenerate oligonucleotide primers designed amplified a 132 bp UT region of cpn60 gene present in Enterobacteriaceae pathogens. The sensitivity of the assay was initially investigated by 10-fold dilution series of overnight broth culture of E. coli ATCC 10536 and had a detection limit of 101 CFU/mL. The specificity was assessed by comparing the amplification plot of a set of Enterobacteriaceae pathogens and non-Enterobacteriaceae members like Staphylococcus aureus (S. aureus) and Listeria monocytogenes (L. monocytogenes) and found to detect all the Enterobacteriaceae cultures included in the study whereas amplification was not seen in S. aureus and L. monocytogenes.

microbiology↗

bioRxiv2026

Giant bacteria in gut microbiomes

The advent of sequencing-based omics has transformed our understanding of the gut microbiome, enabling microbial communities to be characterised at unprecedented scale and resolution. However, these approaches provide limited information about cellular traits such as size and morphology. Giant bacteria remain largely overlooked in gut microbiome research, despite their extraordinary cell size, extreme polyploidy and unusual reproductive strategies. Epulopiscium, one of the largest known heterotrophic bacteria, has been studied primarily as an intestinal symbiont of surgeonfish, leaving its broader host range unresolved. Here we show, by a large-scale analysis of publicly available 16S ribosomal RNA amplicon samples, alongside locally collected human samples, that Epulopiscium spp. occur across diverse hosts, including humans and a broad diversity of animals. Full-length 16S rRNA gene phylogenies resolved multiple Epulopiscium-related lineages with distinct host compositions. Among human samples, Epulopiscium sequences were most abundant during the first year of life. In adults, abundance varied across disease states, increasing in inflammatory bowel disease and colorectal cancer while decreasing in neurological conditions compared with a healthy adult baseline. Similar patterns were also observed in mouse models. Our study broadens the known host range of Epulopiscium and establishes giant bacteria as an overlooked component of gut microbiome diversity. These findings highlight the need to combine sequence-based studies with classical microbiological techniques to identify organisms with unique traits, otherwise hidden by taxonomy or phylogeny.

microbiology↗

bioRxiv2026

Redundancy relocated: the distributed metaparalog pool of an Arctic Pelagibacterales population

The presence of metaparalogs (related genes diverging in function) has been proven to be extensive in the pangenomes of streamlined Pelagibacterales genomes. Here by leveraging a large number of complete genomes from an Arctic population we have been able to establish their universality and mechanistic origins. The diversity of some metaparalog families was as high within a single population than comparing with another (Mediterranean) population/species, which points to an ancient diversification of metaparalog families predating genomospecies separation. Metaparalog families provided diversity for transport and envelope diversity which would provide vast capacities to degrade substrates within DOM and interact with other cells or viruses or other subcellular entities (like EVs). The pool does not saturate at fifteen genomes. Our results highlight the immense importance of the pangenome in understanding the biology of bacteria in natural environments and how critical is the study of population genomics for the future of microbiology.

microbiology↗

bioRxiv2026

AMMPER-2: A spatially explicit agent-based model of microbial radiobiology with redox dye simulation

To reduce health risks for human space exploration, it is important to model the effects of deep-space radiation on biological systems. The budding yeast Saccharomyces cerevisiae is a common model organism in space radiobiology, but little is known about how radiation damage to individual cells translates into the population-level effects that experiments measure. The Agent-Based Model for Microbial Populations Exposed to Radiation (AMMPER) is a spatially explicit agent-based model that simulates the effects of ionizing radiation on S. cerevisiae cells and produces population-level results. Its goal is to improve experiment design and data interpretation. AMMPER-1 demonstrated that agent-based simulations can qualitatively reproduce the effects of proton radiation on the growth rate of wild-type and DNA-repair mutant yeast. Here, we present AMMPER-2, which addresses limitations of the first model and expands its capabilities. AMMPER-2 incorporates the dynamics of the redox dye alamarBlue, a data type commonly generated in spaceflight microbiology experiments, and models the fate of reactive oxygen species in more detail. It also adds an improved graphical interface and runs faster. AMMPER-2 remains easy to execute in Python on a standard personal laptop, providing space biologists a tool to explore experimental conditions, generate hypotheses, and interpret data from microbial space radiation experiments.

microbiology↗