Biology subjects

Negus, D.

Publications and source records attributed to Negus, D..

7 recordsLinked to original sources

bioRxiv2026

Hippurate hydrolases encoded by Klebsiella pneumoniae and Klebsiella oxytoca are functional

Human urine is non-sterile, harbouring its own microbiota (urobiota). Klebsiella species are commensals of the urobiota but can also cause urinary tract infections (UTIs). Gut-microbiota-host co-metabolites (MHCMs) such as hippurate are major components of urine. We previously showed UTI-associated strains of Klebsiella pneumoniae break down hippurate in human urine, though the enzymes responsible for this bioconversion are unknown. Here we sought to confirm that K. pneumoniae hydrolyses hippurate, and to determine whether the enzyme(s) responsible for this bioconversion are present and/or active in other Klebsiella species. Ninhydrin assays confirmed that strains of K. pneumoniae (n=3) and, albeit to a lesser extent, Klebsiella oxytoca (n=4) isolated from human urine could produce glycine from hippurate. Comparative protein (KEGG, UniProt, phylogenetic) and structural (AlphaFold, ChimeraX) analyses were used to predict putative hippurate hydrolases (HHs) encoded by Klebsiella species. Their gene products (n=2 K. pneumoniae; n=3 K. oxytoca) were cloned, expressed and purified. The proteins belonged to three distinct groups: only group 1 and group 2 HHs hydrolysed hippurate under conditions used in this study. Both HHs co-occurred with high prevalence in 15/20 (75 %) Klebsiella species (2885/3012 genomes, 95.8 %). Most Klebsiella species encode two distinct HHs that release glycine and benzoate from hippurate. The functional group 2 HH is predicted to facilitate delivery of amino acids such as glycine to K. pneumoniae cells in the nutrient-limited environment of urine, and its activity demonstrates that members of the urobiota and opportunistic pathogens can contribute to catabolism of MHCMs in human urine.

microbiology↗

bioRxiv2025

Isolation and characterization of broad-range Staphylococcus epidermidis sepunaviruses

Staphylococcus epidermidis is a major contributor to biofilm-associated infections, bacteraemia, and sepsis in humans. Additionally, it is an important veterinary pathogen. The spread of multi-drug resistant S. epidermidis (MRSE) poses an even greater challenge, requiring the development of antibiotic-alternative approaches. Here, we isolated four new broad-range phages capable of infecting a large panel of 65 clinical and veterinary isolates of S. epidermidis. Phages with the broadest host range produced plaques on 37% of the hosts tested and halos on an additional 40%. These phages belonged to the Sepunavirus genus, as supported by their morphology and genome analysis. A genome-wide association study identified a significant correlation between resistance to phage lysis and the presence of the sbcCD complex and the ica operon, highlighting the protective role of biofilms against the phages isolated in this study. Furthermore, distinct phage-encoded methyltransferases identified in the phage genomes may contribute to differences in host ranges. This study advances our knowledge on the diversity and characteristics of S. epidermidis phages, which could be considered as interesting tools for phage therapeutics. RepositoriesPhage genomes are available under Genbank accession numbers PV426907-PV426910. Bacterial genome sequences determined as part of this study, and all Bakta annotations, are available via Figshare (doi:10.6084/m9.figshare.29066417 and doi:10.6084/m9.figshare.29079968 respectively).

microbiology↗

bioRxiv2025

Phenotype-genotype discordance in antimicrobial resistance profiles of Gram-negative uropathogens recovered from catheter-associated urinary tract infections in Egypt

Catheter-associated urinary tract infections (CAUTIs) are among the most common healthcare-associated infections in low- and middle-income countries (LMICs). In this study, phenotypic (EUCAST AMR profiles) and genomic data were generated for 132 isolates (67 Escherichia coli, 14 Pseudomonas aeruginosa, 11 Klebsiella pneumoniae, 9 Proteus mirabilis, 8 Providencia spp., 5 Enterobacter hormaechei, and 18 rare uropathogens) recovered from CAUTIs in an Egyptian hospital. Among the Escherichia coli isolates, phylogroup B2 was most represented (53.7 %), followed by B1 (19.4 %), A (11.9 %), D (7.4 %), F (5.9 %) and C (1.4 %). Several (22/132, 16.6 %) isolates were multidrug-resistant, while almost half (62/132, 46.9 %) were extensively drug-resistant. Comparison of phenotypic data with genotypic data from three different AMR-profiling tools (ResFinder, CARD, AMRFinder) highlighted phenotype-genotype discordance as an important consideration in resistome studies in Egypt, with a total concordance of 91.1 %, 85.7 %, and 80.3 % for ResFinder, CARD and AMRFinder, respectively. Pseudomonas, at the species level, exhibited the greatest discordance. At the antimicrobial level, meropenem was subject to greatest discordance. In addition to the findings from our comparative analyses, new AMR variants are reported for Egypt for Pseudomonas (OXA-486, OXA-488, OXA-905, IMP-43, PDC-35, PDC-45, PDC-201) and Escherichia coli (TEM-176, TEM-190). In summary, we show that there is phenotype-genotype discordance in AMR profiling among CAUTI isolates, highlighting the need for comprehensive approaches in resistome studies. We also show the genomic diversity of Gram-negative uropathogens contributing to disease burden in a little-studied LMIC setting.

microbiology↗

bioRxiv2024

Weberviruses are gut-associated phages that infect Klebsiella spp.

Weberviruses are bacteriophages (phages) that can infect and lyse clinically relevant, multidrug-resistant (MDR) strains of Klebsiella. They are an attractive therapeutic option to tackle Klebsiella infections due to their high burst sizes, long shelf life and associated depolymerases. In this study we isolated and characterized seven new lytic phages and compared their genomes with those of their closest relatives. Gene-sharing network, ViPTree proteome and terL gene-sequence-based analyses incorporating all publicly available webervirus genomes [n=258 from isolates, n=65 from metagenome-assembled genome (MAG) datasets] confirmed the seven phages as members of the genus Webervirus and identified a novel genus (Defiantjazzvirus) within the family Drexlerviridae. Using our curated database of 265 isolated phage genomes and 65 MAGs (n=330 total), we found that weberviruses are distributed globally and primarily associated with samples originating from the gut: sewage (154/330, 47 %), wastewater (83/330, 25 %) and human faeces (66/330, 20 %). We identified three distinct clusters of potential depolymerases encoded within the 330 genomes. Due to their global distribution, frequency of isolation and lytic activity against the MDR clinical Klebsiella strains used in this study, we conclude that weberviruses and their depolymerases show promise for development as therapeutic agents against Klebsiella spp.

microbiology↗

bioRxiv2023

Lelliottia amnigena recovered from the lung of a harbour porpoise, and comparative analyses with Lelliottia spp.

Strain M1325/93/1 (= GFKo1) of Lelliottia amnigena was isolated from the lung of a harbour porpoise in 1993. The genome sequence and antimicrobial resistance profile (genomic, phenotypic) of the strain were generated, with the genomic data compared with those from closely related bacteria. We demonstrate the recently described chromosomally-encoded AmpC {beta}-lactamase blaLAQ is a core gene of L. amnigena, and suggest new variants of this class of lactamase are encoded by other members of the genus Lelliottia. Although presence of blaLAQ is ubiquitous across the currently sequenced members of L. amnigena, we highlight that strain GFKo1 is sensitive to ampicillin and cephalosporins. These data suggest blaLAQ may act as a useful genetic marker for identification of L. amnigena strains, but its presence may not correlate with expected phenotypic resistances. Further studies are required to determine the regulatory mechanisms of blaLAQ in L. amnigena.

microbiology↗

bioRxiv2023

Phage vB_KmiS-Kmi2C infects members of the Klebsiella oxytoca complex and represents a novel genus of lytic bacteriophage

AIMSThis study aimed to characterise the lytic phage vB_KmiS-Kmi2C, isolated from sewage water on a GES-positive strain of Klebsiella michiganensis. METHODS AND RESULTSComparative phylogenetic and network-based analyses were used to characterise the genome of phage vB_KmiS-Kmi2C (circular genome of 42,234 bp predicted to encode 55 genes), demonstrating it shared little similarity with other known phages. The phage was lytic on clinical strains of K. oxytoca (n=2) and K. michiganensis (n=4), and was found to both prevent biofilm formation and disrupt established biofilms produced by these strains. CONCLUSIONSWe have identified a phage capable of killing clinically relevant members of the Klebsiella oxytoca complex (KoC). The phage represents a novel virus family (proposed name Dilsviridae) and genus (proposed name Dilsvirus). SIGNIFICANCE AND IMPACT OF THE STUDYIdentification a novel lytic phage active against clinically relevant strains of the KoC provides an alternative to antibiotics to treat these increasingly antimicrobial-resistant opportunistic pathogens. The unusual way in which the phage can disrupt established biofilms may allow us to identify novel phage-based approaches for biofilm remediation in the future.

microbiology↗

bioRxiv2022

Extended genomic analyses of the broad-host-range phages vB_KmiM-2Di and vB_KmiM-4Dii reveal slopekviruses have highly conserved genomes

High levels of antimicrobial resistance among members of the Klebsiella oxytoca complex (KoC) have led to renewed interest in the use of bacteriophage (phage) therapy to tackle infections caused by these bacteria. In this study we characterized two lytic phages, vB_KmiM-2Di and vB_KmiM-4Dii, that were isolated from sewage water against two GES-5-positive Klebsiella michiganensis strains (PS_Koxy2 and PS_Koxy4, respectively). ViPTree analysis showed both phages belonged to the genus Slopekvirus. rpoB gene-based sequence analysis of 108 presumptive K. oxytoca isolates (n=59 clinical, n=49 veterinary) found K. michiganensis to be more prevalent (46 % clinical and 43 % veterinary, respectively) than K. oxytoca (40 % clinical and 6 % veterinary, respectively). Host range analysis against these 108 isolates found both vB_KmiM-2Di and vB_KmiM-4Dii showed broad lytic activity against KoC species. Several putative homing endonuclease genes were encoded within the genomes of both phages, which may contribute to their broad host range. Pangenome analysis of 24 slopekviruses found that genomes within this genus are highly conserved, with more than 50 % of all predicted coding sequences representing core genes at [≥]95 % identity and [≥]70 % coverage. Given their broad host ranges, our results suggest vB_KmiM-2Di and vB_KmiM-4Dii represent attractive potential therapeutics. In addition, current recommendations for phage-based pangenome analyses may require revision.

microbiology↗