Access Microbiology

The COVID-19 pandemic has imposed a reevaluation of safety protocols across various sectors, including the arts. This study addresses a critical gap in understanding SARS-CoV-2 persistence on materials commonly associated with musical instruments and scores, such as alloys, varnishes, reeds, and paper. While previous research has explored viral survival on various surfaces, limited data exists for materials specific to musical contexts. In this work, we investigate the efficacy of quarantine as a non-destructive method for inactivating SARS-CoV-2 on 16 materials, including brass, silver plating, ABS plastic, ebonite, and various varnishes and paper types. Results revealed significant variability in viral persistence across materials. Non-porous surfaces like metals and ABS plastic cleared infectivity within 3 days, while porous materials such as reeds and music scores required up to 7 days. Gold-plated brass and certain varnishes showed intermediate persistence, with infectivity clearing after 4 days. These findings are in agreement with prior studies indicating that SARS-CoV-2 survival is highly dependent on surface composition, with porous and organic-coated materials retaining viable virus longer due to reduced environmental stress. Our results highlight the feasibility of stratified quarantine protocols based on material type, offering practical guidelines for musicians and institutions and provides critical insights for mitigating SARS-CoV-2 transmission risks in musical settings.

Antimicrobial resistance (AMR) is a major global public health threat. Wild birds, including seagulls, are increasingly recognised as potential reservoirs and disseminators of resistant bacteria linked to human activity. The objective of the study was to assess the association between human population density and the occurrence of Escherichia coli resistant to critically important antimicrobials in Australian seagulls. Faecal samples were collected from seagull populations in coastal regions across Australia representing differing human population densities. Resistant E. coli isolates were identified and characterised using multilocus sequence typing and plasmid incompatibility group analysis to determine relatedness to human associated lineages. The frequency of resistant E. coli isolation increased with human population density. The predominant sequence types ST10, ST131 and ST354 comprised 24.5% of isolates and belong to globally distributed human associated lineages linked to extraintestinal pathogenic E. coli. Many isolates carried IncF and IncI plasmids, which are key vectors of blaCTX-M extended spectrum beta lactamase genes and plasmid mediated quinolone resistance determinants commonly reported in human clinical strains. IMPORTANCEThese findings support the contention that seagulls primarily acquire resistant bacteria through contact with anthropogenic activities. Once acquired, these bacteria may be disseminated to other seagulls, birds and animals, including being transmitted to humans.

Melanins are biological macromolecule with immense functionality synthesised by a wide spectrum of living organism. It is mainly synthesised by the oxidative polymerization of indolic and phenolic compounds through several enzymatic process. It has wide spread application in agriculture, cosmetic and therapeutic industry due to its various properties including antioxidation ability, UV protection efficiency and anticancer activity. Because of this wide range of application in different sectors, large scale production and commercialization attains enormous consideration. The present study deals with the effect of 12 different process parameters on melanin production viz., production media, incubation time, inoculum concentration, pH, temperature, agitation, carbon source, phosphate and magnesium source, CuSO4.5H2O, sodium chloride and L-tyrosine on melanin production by Pseudomonas stutzeri strain BTCZ 109 obtained from Arabian sea sediments was evaluated. After optimizing the important process parameters, the bacteria showed about ~4.65 fold increase in melanin production compared to unoptimized cultural conditions. The melanin optimized through this method was found to be nano sized. The Nano sized DOPA melanin in treating Skin cancer cell line SK ML28 which showed a dose-dependent activity with an IC50 value of 164 g/mL. All these results highlight the therapeutic efficiency of DOPA melanin Nano particle as promising bioactive molecule.

Climate-driven increases in sea surface temperature have been associated with the expansion of Vibrio species and a corresponding rise in vibriosis cases in both human populations and aquaculture systems. Coastal waters across the south of England are increasingly becoming suitable for the growth and establishment of both human- and aquaculture-associated Vibrio species, potentially increasing vulnerability to the types of infections and disease outbreaks already reported elsewhere in the world. In this study, we report the presence of a diverse and well-established Vibrio community within the Teign Estuary, (Southwest, UK), including the human-pathogenic species V. parahaemolyticus, V. cholerae (non-O1/non-O139), V. alginolyticus, and V. diabolicus, as well as the important aquaculture pathogens V. jasicida, V. aestuarianus, and V. anguillarum. We identified V. diabolicus, a species that was indistinguishable from V. alginolyticus using conventional biochemical identification methods and could only be accurately resolved by whole-genome sequencing and developed novel PCR targets to differentiate these species in the lab. Using the insect infection model Galleria mellonella, we demonstrate that environmental isolates of V. cholerae (non-O1/non-O139), V. parahaemolyticus, and V. alginolyticus possess virulence potential. We also investigated the effects of sewage effluent on the growth of Vibrio isolates from the Teign Estuary and found that sewage can preferentially promote the growth of Vibrio species. Furthermore, several Vibrio isolates were multidrug resistant and carried antimicrobial resistance genes, highlighting the potential role of environmental Vibrio populations in the Teign Estuary as reservoirs of antimicrobial resistance. Together, these findings demonstrate how rising sea surface temperatures and sewage pollution may influence the emergence, persistence, and public health and aquaculture significance of Vibrio species in UK coastal waters.

Macrolides are key agents for treating infections caused by non-tuberculous mycobacteria (NTM). Nevertheless, chromosomal erm genes conferring inducible macrolide resistance are described in some NTM species, such as Mycobacterium abscessus and M. fortuitum, whereas M. chelonae had long been considered as lacking a functional erm. Recent descriptions from the USA and Japan of a new plasmid-borne erm(55) (erm(55)P) in M. chelonae and other rapidly growing mycobacteria (RGM) have challenged this assumption. We investigated erm(55)P occurrence in clinical RGM referred to the French National Reference Centre for Mycobacteria between 2012 and 2026 by genome screening and erm(55)P specific real-time PCR. Positive isolates underwent long-read whole genome sequencing (GridIon, Oxford Nanopore Technologies). Clarithromycin (CLR) minimum inhibitory concentration (MIC) was determined by broth microdilution (RAPMYCO and FRATMYC, Thermo Fisher) and read up to 14 days. Five clinical isolates showing inducible CLR resistance (MIC range <0.25-64 mg/L on day 3-4 and 128 - >128 mg/L on day 14) were positive for erm(55)P: one M. chelonae, three M. neoaurum, and one M. parafortuitum. erm(55)P-positive M. chelonae genomes from this and previous descriptions did not cluster together in the phylogenetic analysis of 263 genomes. The assembled plasmids showed high similarity to previously reported erm(55)-carrying plasmids, especially within the erm(55)P region. The upstream sequence of erm(55)P showed a secondary structure compatible with a possible translation attenuation mechanism. These findings document the first report of a plasmid-borne erm(55) in Europe in M. chelonae and other RGM and raise concern about the emergence of plasmid macrolide resistance in NTM.

Plasmid-encoded mobile colistin resistance (mcr) genes have raised concern due to dissemination potential. While mcr variants are reported across One Health compartments, they remain unreported in grass. This study characterises a novel mcr variant (mcr-13.1), detected in Escherichia coli isolated from the grass phylosphere within an agricultural grassland. The two mcr-positive isolates were clonal copies isolated at timepoints eight weeks apart. They belonged to the serotype O17:H18 and were of the sequence type ST394. The E. coli were phenotypically susceptible to {beta}-lactams, aminoglycosides, quinolones, sulphonamides, phenicols, tetracyclines, diaminopyrimidine and colistin (Minimum Inhibitory Concentration (MIC) = 0.5 {micro}g/mL). The mcr-13.1 gene was encoded on an IncFIB plasmid. This plasmid was transferable by conjugation but the colistin MIC of the E. coli J53 transconjugant did not change (0.5 {micro}g/mL). Further, cloned pUC19::mcr-13.1 did not alter the colistin MIC for E. coli DH5 (0.25 {micro}g/mL). The translated amino acid sequence showed highest homology (82 %) to MCR-10.2 and MCR-10.4. Our findings identify grass as a previously unrecognised reservoir for E. coli carrying mobile mcr genes, reports the identification of the novel mcr-13.1 variant from this niche and demonstrates the importance of genomic screening in identifying mcr genes that would otherwise remain undetected.

Oligella otitidis MSHR-50489EDL strain (ATCC: TSD462; DSMZ: DSM118617) is a new species of the genus Oligella that was isolated from a middle ear discharge swab from a child with chronic suppurative otitis media (CSOM). This Gram-negative coccobacillus produces small, circular, smooth, whitish-opaque and occasionally mucoid colonies. It grows in aerobic conditions at a temperature range from 25-42oC. Phylogenetic analysis demonstrates a relationship to other species of the genera Oligella and average nucleotide identity and digital DNA/DNA hybridization values indicate a distinct species in comparison to other Oligella species. Thus far, the majority of isolates exhibit resistance to ciprofloxacin, the first line treatment for CSOM.

The vancomycin-resistant Enterococcus faecium (VREfm) has been declared as a high priority pathogen by World Health Organisation (WHO). It presents a major therapeutic challenge to healthcare, remaining with limited antibiotic options. Bacteriophage therapy has emerged as a promising alternative for combating antimicrobial-resistant pathogens. This study reports the isolation and characterization of bacteriophages active against VREfm and MDR E. faecium clinical isolates from India. Two bacteriophages, NIMS_EF375_N69_P9 ({Phi}1) and NIMS_EF375_N74_P12 ({Phi}2), were isolated from sewage by enrichment using MDR E. faecium strain EF375 as the propagation host. Lytic activity was confirmed by spot assay and double-layer soft agar plaque assay; both phages produced clear plaques of 1.5-2.0 mm. Transmission electron microscopy showed icosahedral heads with long non-contractile tails, presenting siphovirus-like morphotype within the class Caudoviricetes. Host range was assessed against 10 MDR E. faecium isolates (including the propagation host), 2 of which were VREfm. {Phi}1 lysed 5 of 10 isolates, including the VanA-type VREfm, whereas {Phi}2 lysed 7 of 10, including both VanA- and VanB-type VREfm. In time-kill assays against EF375 at an MOI of 1, {Phi}1 produced effective decline in optical density sustained through 24 h (~80% reduction relative to the untreated control), whereas {Phi}2 suppressed bacterial growth with only ~50% reduction. To the best of our knowledge, this study represents the first report from India on the isolation and characterization of bacteriophages active against clinical VREfm isolates. The broader host range of {Phi}2 and the stronger killing kinetics of {Phi}1 suggest complementary roles in a phage cocktail, warranting genomic characterization and in vivo evaluation.

This study investigated the genomic and biochemical characteristics of two amylolytic microbial strains, NATIVEDY160T (= JE7B6T = NRRL B-68523T) and NATIVEDY161T (= JL13D9T, = NRRL B-68524T) isolated from the rumen of healthy Holstein dairy cattle. Both strains are obligately anaerobic, non-motile, Gram positive, catalase-negative, and oxidase-negative. Morphologically, NATIVEDY160T grows in long coccoid chains while NATIVEDY161T grows in short chains or pairs. NATIVEDY160T can catabolize amygdalin, esculin/ferric citrate, and starch, compared to NATIVEDY161T which utilizes amygdalin, arbutin, esculin/ferric citrate, glycogen, and D-maltose as determined by API 50 CH carbon panels. Starch degradation ability was verified for both strains, but neither showed cellulolytic activity as confirmed by starch agar and Congo red agar assays, respectively. HPLC analysis revealed that lactate was the primary end product of both strains carbohydrate fermentation, while strain NATIVEDY161T also produced small amounts of acetate. 16S rRNA sequences from both strains cluster with the Oscillospiraceae (formerly Ruminococcaceae) lineage Ruminococcus species, but average nucleotide identity of either strain compared to closely related Ruminococcus members was under the species threshold (95%). Genomic, phylogenetic, and phenotypic interrogation support NATIVEDY160T and NATIVEDY161T as novel species. Each strain was isolated from the rumen of dairy cows located within the central valley of southern California, which has a rich history of Dutch and Basque dairy farm ownership and is still the case today in the region. In recognition of the contributions and heritage of the central and southern California dairy industry, the names Ruminococcus hollandia and Ruminococcus vasco are proposed with NATIVEDY160T and NATIVEDY161T as their respective type strains.

AO_SCPLOWBSTRACTC_SCPLOWUnderstudied bacterial genera present a dynamic phylogenetic landscape and opportunities for discovering new taxa as more strains are isolated and genomic data is added. Here, through phylogenomic analysis, we describe three novel species of the globally distributed cosmopolitan marine bacterial genus Microbulbifer. This genus is ubiquitous in saltwater microbiomes and is a validated source of biodegradation enzymes as well as high value small molecule natural products. Average nucleotide identity (ANI) to the closest known species, Microbulbifer variabilis ATCC 700307T, was less than 88.4% for all three novel species. Isolates of the three novel species, designated as PAAF003T (T = type strain), ZKSA006T, and SSSA003T were imaged to reveal their phormological characteristics. Based on phylogenetic data, strains PAAF003T, ZKSA006T, and SSSA003T represent three new species of the genus Microbulbifer, for which the names Microbulbifer maximicatervae sp. nov., Microbulbifer regidiadema sp. nov., and Microbulbifer mixtoriginis sp. nov. are proposed, respectively, under the SeqCode. We also reconstructed a robust phylogeny of available Microbulbifer genomes, which should faciliatate future isolation and strain description studies.

The use of antimicrobial compounds in humans, animals and in agriculture leads to environmental antimicrobial contamination through domestic, industrial and agricultural wastewater. Efforts have been made to perform environmental risk assessments based on the potential of these compounds to select for antimicrobial resistance (AMR) at certain concentrations in bacteria. This has resulted in predicted no effect concentrations (PNEC) which determine the minimum thresholds required to select for resistance. However, the effects of these compounds on intracellular transposition within bacterial isolates, a major driver of AMR, have not been previously assessed. Here, we assess the effect of differing sub-inhibitory concentrations of the third-generation cephalosporin, ceftriaxone, on the rate of intracellular transposition in combination with colistin and kanamycin. Two triple replicons systems (RS1 and RS2) were developed to assess this, each containing a chromosome, plasmid and entrapment vector. We show that sub-inhibitory concentrations of ceftriaxone exert hormetic effects on the intracellular transposition rate in RS1 and a steady linear increase in RS2. This defines a predicted no effect concentration for transposition (PNECT) for ceftriaxone as 320 ng/L in RS1 and 3200 ng/L in RS2. This provides a minimum threshold for the environmental impact of ceftriaxone on biological systems at the sub-cellular scale, which is applicable to industrial standards of waste management, where consideration of ecological impact is central.

Different plasmids exist at different copy numbers per cell, and there is an inverse relationship between a plasmids copy number and its size. Two recent studies quantified this relationship into a scaling law, but both the form and the interpretation of this law are contested. Here, I explore the issues with fitting a single law across plasmid diversity and suggest a consistent synthesis. First, I explore some potential problems with using sequencing-based estimates of copy number. Then, I discuss plasmid copy number through a series of case studies. I argue in favour of interpreting plasmid copy numbers not through a single law, but through the lens of two dominant evolutionary strategies. I suggest that small plasmids which lack active segregation mechanisms have a resulting tradeoff between plasmid inheritance and fitness cost to the host, which is responsible for an inverse relationship between copy number and size. In contrast, larger plasmids with active segregation mechanisms show a much weaker relationship, in line with evidence that their metabolic costs are dominated by the expression of specific genes rather than their size. Where plasmids in the 20-100kb range have higher copy numbers, I argue these probably arise more from selection at the level of the host cell for plasmid-associated phenotypes (e.g. antibiotic resistance) rather than from plasmid-level selection for inheritance.

Multilocus sequence typing (MLST) remains the predominant method for typing bacterial strains. A common method for investigating particularly successful epidemic lineages within a species is to cluster isolates with similar MLST profiles into clonal complexes (CCs). Some CCs, such as international clones (ICs) in A. baumannii, are identified with specific sequence types (STs) and are of particular importance to human health. Although theoretically simple, there is a lack of convenient, user-friendly tools perform this analysis. Here we present PhyloMLST, a tool to cluster bacterial isolates into CCs and map them to ICs using the output from existing MLST tools and user-provided STs. As there is potential for aberrant IC assignment arising from excessively large CCs constructed with spurious links, PhyloMLST provides additional functionality to correct IC assignment with a user-provided phylogenetic tree. Although designed with A. baumannii in mind, PhyloMLST can be applied to any bacteria where construction and investigation of CCs based on MLST is performed. Impact statementMany bacterial pathogens are characterised by successful epidemic lineages that are responsible for a substantial number of infections, may be more virulent, and may carry an abundance of antimicrobial resistance genes. These epidemic lineages are comprised of a number of multilocus sequence typing (MLST) sequence types (STs), clustered into clonal complexes (CCs). To date, identifying which STs belong to which epidemic lineage has been challenging, with no simple analytical tools available. Here, we present PhyloMLST - a phylogenetically-aware method for assigning STs to epidemic lineages. The customisable nature of the tool will enable researchers to straightforwardly characterise any population of bacteria that they are working on using MLST data and user-defined definitions of epidemic lineages. Data summaryThe PhyloMLST source code and example data shown here is available at https://github.com/Mattn286/PhyloMLST.

Recombinase polymerase amplification (RPA) enables rapid nucleic acid testing in low-resource environments, but poorly characterized byproducts can compromise assay specificity and cause false-positive results. Here, we amplified the thirteen original CODIS core loci and Amelogenin to characterize recurrent RPA artefacts and establish conditions that reduce their formation. First, RPA products were analyzed for two reference samples by Oxford Nanopore Technologies sequencing. This revealed two distinct classes of multimeric products: primer multimers and amplicon multimers, consisting of repeated primer or amplicon sequences, respectively. Individual artefacts contained up to 281 primer copies or 22 amplicon copies, demonstrating the extensive range of these products. Next, we performed an optimization study to evaluate the effects of reaction temperature and reagent concentrations at two representative loci, D3S1358 and D5S818. Among the conditions tested, temperature had the most pronounced effect. Reducing the temperature from 42{degrees}C to 34{degrees}C increased the relative target amplicon fraction from 15% to 83% for D3S1358 and from 84% to 98% for D5S818, while maintaining or increasing absolute target concentration. Lower primer concentrations and higher T4 UvsX concentrations also reduced multimer formation, although lower primer concentrations reduced target yield and caused allelic dropout. Finally, amplification at 34{degrees}C was evaluated across all fourteen loci by sequencing. Relative to 42{degrees}C, the target read fraction increased by more than 5 percentage points for 7/14 loci in one reference sample and 9/14 loci in the other, with the largest improvements at multimer-prone loci. These findings identify multimers as an important class of RPA artefacts and establish reaction temperature and T4 UvsX concentration as promising conditions to improve RPA specificity.

Thermostable DNA polymerases are indispensable tools in molecular biology, yet enzymes from the most extreme hyperthermophiles remain largely uncharacterized. Here, we report the biochemical and structural characterization of a family B DNA polymerase from Pyrolobus fumarii A1 (Pyrfu pol), one of the most thermoresistant archaea described to date. The enzyme was efficiently overproduced in E. coli Rosetta 2(DE3)[pLysS] and purified to homogeneity using a two-step protocol that combined heat treatment with immobilized metal affinity chromatography (IMAC). Bioinformatic analysis confirmed the canonical family B architecture, while AlphaFold-based structural modeling and comparative analysis with mesophilic RB69 DNA polymerase revealed a well-conserved structural core alongside thermoadaptive features. Radiolabel incorporation assays demonstrated enzymatic activity over a broad ionic strength range and an absolute requirement for Mg ions. PCR-based optimization confirmed these findings and revealed broad pH tolerance (6.5-11.0). Notably, Tris inhibited radiolabel-based assays (pH 7.0) yet proved essential for efficient PCR amplification (pH 8.5), suggesting a context-dependent role of buffer composition in polymerase activity. Processivity assays confirmed amplification of DNA fragments up to approximately 8,000 bp. Replication fidelity, assessed by the lacZ-based assay, showed a 2.9-fold improvement over Taq polymerase. Urea-nanoDSF yielded an exceptional melting temperature of 105.9 {+/-} 0.08 {degrees}C. Pyrfu pol also demonstrated tolerance to common PCR inhibitors, highlighting its potential utility in molecular biology applications.

Probiotics produce antimicrobial peptides and small molecules that are secreted into the medium. Antimicrobial activity of cell-free supernatants can be tested using various qualitative and quantitative methods. Many of these techniques employ methods which introduce uncontrolled variables, impacting reproducibility and making comparison of reported results difficult. Here, we present a simple procedure for quantitative estimation of antimicrobial activity of cell-free supernatants which overcomes drawbacks of commonly used methods.

H5N1 clade 2.3.4.4b avian influenza A viruses pose a significant threat to wild animal populations, domesticated animals, and potentially, the human population. For H5N1s to infect and transmit among mammalian species, mutations for improved utilization of mammalian receptors and enhanced replication at the lower temperatures of the upper respiratory tract need to be acquired. A human H1N1pdm09-like virus was compared to H5N1 genotypes B3.13 and D1.1 for replication at 33{o}C, 37{o}C, and 39{o}C - temperatures consistent with the upper and lower respiratory tract in humans, and dairy cow udder tissue. All H5N1 viruses had increased plaque sizes on MDCK cells at 37{o}C and 39{o}C compared to H1N1pdm09. In primary, differentiated human nasal and bronchial epithelial cultures, all H5N1 viruses show restricted infectious virus production compared to H1N1 at 33{o}C. While H5N1 D1.1 also showed restricted replication at 37{o}C and 39{o}C, the H5N1 B3.13 replicated to nearly equivalent titers as H1N1pdm09. All H5N1 viruses demonstrated similar cell tropism in cells from the upper and lower respiratory tract, infecting more ciliated than non-ciliated cells relative to H1N1pdm09. H1N1, H5N1 B3.13 D1.1 infection induced similar innate immune factors, with nasal epithelial cells producing higher levels compared to bronchial epithelial cells. These data suggest that genotype B3.13 and D1.1 H5N1 viruses show different temperature dependent replication patterns compared to H1N1pdm09.

Streptococcus pyogenes is a globally important pathogen responsible for at least 500,000 deaths a year, causing significant burden on healthcare systems. It is the causative agent for ailments such as impetigo and strep throat to septicaemia and necrotizing fasciitis. Assessment of genetic relatedness for the detection of outbreaks within communities or healthcare facilities is vital in decreasing the propagation of S. pyogenes within these settings, alongside epidemiological data. As the volume of isolates being sequenced increases year on year, more scalable and sharable methodologies of assessing genetic relatedness are required by reference laboratories and for international collaboration. LIN codes, applied to core genome MLST (cgMLST) represent a method which is extensible to large scale whole genome sequencing (WGS) while still being sufficiently sensitive to detect outbreak clusters. Here we present a novel cgMLST and LIN code scheme, hosted by PubMLST, enabling international collaboration and global tracking of variants, that is highly scalable and usable for all. The schemes are available at https://pubmlst.org/organisms/streptococcus-pyogenes. Data SummaryGenome sequences and metadata are available at https://pubmlst.org/organisms/streptococcus-pyogenes. PubMLST and ENA accessions and metadata can additionally be found in the supplementary data. Raw reads for UKHSA sequences are available in ENA study PRJEB115996. Impact StatementStreptococcus pyogenes is a globally relevant pathogen capable of causing invasive and non-invasive disease across a multitude of settings. Assessment of genetic relatedness is an increasingly important aspect of managing outbreaks, requiring solutions that are scalable, high resolution and comparable across laboratories. Here we present a high resolution core genome multi locus sequence typing (MLST) and associated life identification number (LIN) code scheme, The schemes were developed using a combination of 4,916 UKHSA and 2,391 publicly available S. pyogenes isolates in order to cover a wide range of EMM types both within the UK and globally. These new schemes enable high resolution typing of S. pyogenes isolates, suitable for analysis of lineages to genomic epidemiology in outbreak detection and management. Both cgMLST and LIN code schemes are available on PubMLST as an open access resource for the public health and academic communities and can enable both intra laboratory and global coordination.

A rapid expansion of influenza A virus (IAV) genome sequencing has transformed global surveillance but has also created major challenges for interpreting the biological significance of viral mutations, particularly amino acid replacements associated with host adaptation. Resources have been created to support mutation annotation and phylogenetic analysis, but there is a need for a tool that integrates experimentally derived phenotypic evidence with evolutionary context in a framework suitable for users without prior training in bioinformatics. Here, we present the Flu Mutation Explorer, an interactive web application that combines large-scale influenza phylogenies with a manually curated database of reported mammalian adaptation mutations, to enable the exploration and interpretation of IAV genetic variation. The underlying database comprises over 1.5 million publicly available IAV sequences and over 1000 mutations associated with mammalian adaptation. The Flu Mutation Explorer enables users to query protein sequences, visualise amino acid distributions across viral lineages, examine host-specific conservation patterns, and identify adaptation mutation with links to supporting literature. We include case studies which demonstrate the platforms use in assessing amino acid conservation at sites of interest and in rapidly identifying candidate mammalian adaptation mutations during the ongoing H5N1 panzootic. By integrating genomic, phylogenetic, and functional information into an intuitive interface, the Flu Mutation Explorer lowers the barriers to interpreting influenza sequences for specialists and non-specialists alike.

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.