UK Isolates Research Articles

Multiple routes to fungicide resistance: Interaction of Cyp51 gene sequences, copy number and expression.

We examined the molecular basis of triazole resistance in Blumeria graminis f. sp. tritici (wheat mildew, Bgt), a model organism among powdery mildews. Four genetic models for responses to triazole fungicides were identified among US and UK isolates, involving multiple genetic mechanisms. Firstly, only two amino acid substitutions in CYP51B lanosterol demethylase, the target of triazoles, were associated with resistance, Y136F and S509T (homologous to Y137F and S524T in the reference fungus Zymoseptoria tritici). As sequence variation did not explain the wide range of resistance, we also investigated Cyp51B copy number and expression, the latter using both reverse transcription-quantitative PCR and RNA-seq. The second model for resistance involved higher copy number and expression in isolates with a resistance allele; thirdly, however, moderate resistance was associated with higher copy number of wild-type Cyp51B in some US isolates. A fourth mechanism was heteroallelism with multiple alleles of Cyp51B. UK isolates, with significantly higher mean resistance than their US counterparts, had higher mean copy number, a high frequency of the S509T substitution, which was absent from the United States, and in the most resistant isolates, heteroallelism involving both sensitivity residues Y136+S509 and resistance residues F136+T509. Some US isolates were heteroallelic for Y136+S509 and F136+S509, but this was not associated with higher resistance. The obligate biotrophy of Bgt may constrain the tertiary structure and thus the sequence of CYP51B, so other variation that increases resistance may have a selective advantage. We describe a process by which heteroallelism may be adaptive when Bgt is intermittently exposed to triazoles.

A core genome multi-locus sequence typing scheme for Streptococcus uberis: an evolution in typing a genetically diverse pathogen.

Streptococcus uberis is a globally endemic and poorly controlled cause of bovine mastitis impacting the sustainability of the modern dairy industry. A core genome was derived from 579 newly sequenced S. uberis isolates, along with 305 publicly available genome sequences of S. uberis isolated from 11 countries around the world and used to develop a core genome multi-locus sequence typing (cgMLST) scheme. The S. uberis core genome comprised 1475 genes, and these were used to identify 1447 curated loci that were indexed into the cgMLST scheme. This was able to type 1012 of 1037 (>97 %) isolates used and differentiated the associated sequences into 932 discrete core genome sequence types (cgSTs). Analysis of the phylogenetic relationships of cgSTs revealed no clear clustering of isolates based on metadata such as disease status or year of isolation. Geographical clustering of cgSTs was limited to identification of a UK-centric clade, but cgSTs from UK isolates were also dispersed with those originating from other geographical regions across the entire phylogenetic topology. The cgMLST scheme offers a new tool for the detailed analysis of this globally important pathogen of dairy cattle. Initial analysis has re-emphasized and exemplified the genetically diverse nature of the global population of this opportunistic pathogen.

Genomic diversity and epidemiological significance of non-typhoidal Salmonella found in retail food collected in Norfolk, UK.

Non-typhoidal Salmonella (NTS) is a major cause of bacterial gastroenteritis. Although many countries have implemented whole genome sequencing (WGS) of NTS, there is limited knowledge on NTS diversity on food and its contribution to human disease. In this study, the aim was to characterise the NTS genomes from retail foods in a particular region of the UK and assess the contribution to human NTS infections. Raw food samples were collected at retail in a repeated cross-sectional design in Norfolk, UK, including chicken (n=311), leafy green (n=311), pork (n=311), prawn (n=279) and salmon (n=157) samples. Up to eight presumptive NTS isolates per positive sample underwent WGS and were compared to publicly available NTS genomes from UK human cases. NTS was isolated from chicken (9.6 %), prawn (2.9 %) and pork (1.3 %) samples and included 14 serovars, of which Salmonella Infantis and Salmonella Enteritidis were the most common. The S. Enteritidis isolates were only isolated from imported chicken. No antimicrobial resistance determinants were found in prawn isolates, whilst 5.1 % of chicken and 0.64 % of pork samples contained multi-drug resistant NTS. The maximum number of pairwise core non-recombinant single nucleotide polymorphisms (SNPs) amongst isolates from the same sample was used to measure diversity and most samples had a median of two SNPs (range: 0-251). NTS isolates that were within five SNPs to clinical UK isolates belonged to specific serovars: S. Enteritidis and S. Infantis (chicken), and S. I 4,[5],12:i- (pork and chicken). Most NTS isolates that were closely related to human-derived isolates were obtained from imported chicken, but further epidemiological data are required to assess definitively the probable source of the human cases. Continued WGS surveillance of Salmonella on retail food involving multiple isolates from each sample is necessary to capture the diversity of Salmonella and determine the relative importance of different sources of human disease.

Prevalence and Molecular Characterization of Deformed Wing, Acute Bee Paralysis and Black Queen Cell Viruses Infecting Honey Bees and Varroa Mites

Abstract Surveys were conducted in Malatya and Elazığ provinces of Eastern Türkiye between 2018–2019 to detect the presence and distribution of Deformed wing virus (DWV), Acute bee paralysis virus (ABPV) and Black queen cell virus (BQCV) infecting honey bees and Varroa mites. Thirty Varroa mites feeding on honey bees and 147 honey bees were collected from twenty-five apiaries of Malatya and fifteen apiaries of Elazığ. The collected samples were subjected to reverse transcriptase-polymerase chain reaction. All tested viruses were found to be present in the samples singly or as mixed infections. The prevalence of the viruses were 23.81%, 12.93%, and 10.20% for DWV, ABPV and BQCV, respectively. Five out of thirty Varroa mites tested for the viruses were found to be infected only with DWV. Randomly chosen sequences of each detected virus’s partial polyprotein gene region were registered in GenBank under the accession numbers OP805878, OP805879, OP805880, OP805887, OP805888, OP805889, OP805890, OP805891 (DWV), OP805881, OP805882, OP805883, OP805884 (ABPV) and OP805885, OP805886 (BQCV). The phylogenetic tree of the viral isolates were compared with the world isolates. DWV isolates were found to be closely related to the UK, Lebanon, Türkiye, France, Germany and Israel isolates, whereas ABPV isolates were related to Türkiye, South Africa, Slovenia, Serbia, France, Hungary, Syria and USA isolates and BQCV isolates were related to China, South Korea, Czech Republic, France, Hungary, Poland, South Africa, Australia and Sweden isolates. The presence of DWV, ABPV and BQCV infections in Malatya and Elazığ provinces of Türkiye is revealed for the first time in this study.

Emerging patterns of fluoroquinolone resistance in Campylobacter jejuni in the UK [1998-2018

Campylobacter jejuni (C.jejuni) is the most common causative agent of bacterial food poisoning worldwide and is known to be genetically highly diverse. C. jejuni is increasingly resistant to fluoroquinolone antibiotics, but very few studies have investigated variant-specific patterns of resistance across time. Here we use statistical modelling and clustering techniques to investigate patterns of fluoroquinolone resistance amongst 10,359 UK isolates from human disease sampled over 20 years. We observed six distinct patterns of fluoroquinolone sensitivity/resistance in C. jejuni across time, grouping by clonal complex (CC). Some CCs were fully resistant, some shifted from susceptible to resistant following a sigmoidal shape, and some remained susceptible over time. Our findings indicate that the fluoroquinolone resistance patterns of C. jejuni are complicated and cannot be analysed as a single species but divided into variant dynamics so that the factors driving resistance can be thoroughly investigated.

Genomic Diversity of Bradyrhizobium from the Tree Legumes Inga and Lysiloma (Caesalpinioideae-Mimosoid Clade)

We identified diverse bradyrhizobia having distinct ERIC-PCR genomic fingerprints from native American trees Inga vera and Lysiloma spp. In addition, two nodule isolates recovered from Lysiloma divaricatum seedlings inoculated with soil from the tropical house of a UK botanical garden were also identified as Bradyrhizobium. Genomes were obtained (with sizes around 9 Mb each) from nine Inga and Lysiloma bradyrhizobial isolates; plasmids were detected in two of the Inga strains analyzed. Average Nucleotide Identity of whole genomes revealed five novel Bradyrhizobium genomospecies from Mexican trees, while the UK isolates were identified as Bradyrhizobium cajani and Bradyrhizobium brasilense. Inga vera and Lysiloma isolates, despite their genetic distances and different hosts, shared a common set of nod genes that suggested that I. vera and Lysiloma bradyrhizobia produce fucosylated, methylated and carbamylated lipochitooligosaccharides. Uptake hydrogenase hup, hyp and secretion system genes were found in some of the isolates. Lysiloma strains were found to be ineffective on I. vera. Some of the isolates may be used as plant inoculants.

The ecology and antimicrobial resistance of Staphylococci colonising neonates

Coagulase Negative Staphylococci (CoNS) are common commensals of human skin, account for nearly 20% of the microbiota in infants and are thought to promote early immune responses in healthy babies. However, CoNS are opportunistic pathogens and in the UK between 2005 and 2014 were responsible for 57% of episodes of Late Onset Sepsis (LOS). In neonatal intensive care units (NICUs) this is a major concern and antiseptics are used to prevent vascular catheter infections. Chlorhexidine (CHX) and Octenidine (OCT) are the most common agents used for skin antisepsis, but evidence is emerging of antiseptic tolerance amongst CoNS. We undertook a longitudinal survey of CoNS from skin and rectal swabs isolated from babies in two NICUs from countries with different antiseptic regimens (UK and Germany). Over 1000 isolates were characterised for antimicrobial susceptibility and sequenced. The most frequent species isolated were S. epidermidis and S. haemolyticus with similar strain types present in both units. Reduced susceptibility to CHX and OCT was observed in UK isolates (where CHX is used), compared to German isolates (where OCT is used). Analysis of genome data using GWAS and clustering techniques has identified loci associated with antimicrobial susceptibility. Comparison of isolates taken on admission and thereafter, demonstrated that babies acquired isolates with decreased antiseptic tolerance after admission. This data provides new information about the phylogeny of CoNS in NICUs and suggest different potentials for selection of resistance between antiseptics commonly used in neonatal care.

Genotypic and Phenotypic Structure of the Population of Phytophthora infestans in Egypt Revealed the Presence of European Genotypes.

Late blight disease of potato and tomato, caused by Phytophthora infestans, results in serious losses to Egyptian and global potato and tomato production. To understand the structure and dynamics of the Egyptian population of P. infestans, 205 isolates were collected from potato and tomato plants during three growing seasons in 2010–2012. The characterization was achieved by mating-type assay, metalaxyl sensitivity assay, and virulence pattern. Additionally, genotyping of 85 Egyptian isolates and 15 reference UK isolates was performed using 12 highly informative microsatellite (SSR) markers David E. L. Cooke and five effector (RxLR) genes. Mating-type testing showed that 58% (118 of 205) of the isolates belonged to mating type A1, 35% (71 isolates) to mating type A2, and the rest 8% (16 isolates) were self-fertile. The phenotype of metalaxyl response was represented as 45% resistant, 43% sensitive, and 12% as intermediate. Structure analysis grouped the 85 identified genotypes into two main clonal lineages. The first clonal lineage comprised 21 isolates belonging to A2 mating type and 8 self-fertile isolates. This clonal lineage was identified as Blue_13 or EU_13_A2. The second main clonal lineage comprised 55 isolates and was identified as EU_23_A1. A single isolate with a novel SSR genotype that formed a distinct genetic grouping was also identified. The effector sequencing showed good correspondence with the virulence data and highlighted differences in the presence and absence of loci as well as nucleotide polymorphism that affect gene function. This study indicated a changing population of P. infestans in Egypt and discusses the findings in the context of late blight management.

Wheat stem rust recorded for the first time in decades in Ireland.

Wheat stem rust, caused by the fungus Puccinia graminis f. sp. tritici (Pgt), occurs in most wheat‐growing areas worldwide, and, in western Europe since 2013, has started to re‐emerge after many decades of absence. Following this trend across western Europe, in 2020, we also detected and recorded wheat stem rust for the first time in five decades in experimental plots across five locations in Ireland. To examine the potential origin of the Irish Pgt infection in 2020, we carried out transcriptome sequencing on 12 Pgt‐infected wheat samples collected across Ireland and compared these to 76 global P. graminis isolates. This analysis identified a close genetic relationship between the Irish Pgt isolates and those from Ethiopia collected in 2015 after a severe stem rust epidemic caused by the TKTTF Pgt race, and with the UK‐01 Pgt isolate that was previously assigned to the TKTTF race. Subsequent pathology‐based race profiling designated two Irish isolates and recent UK and French Pgt isolates to the TKTTF Pgt race group. This suggests that the Irish Pgt occurrence most probably originated from recent long‐distance windborne dispersal of Pgt urediniospores from neighbouring countries in Europe where we confirmed the Pgt TKTTF race continues to be prevalent. The identification of wheat stem rust in Ireland at multiple locations in 2020 illustrates that the disease can occur in Ireland and emphasizes the need to re‐initiate local monitoring for this re‐emergent threat to wheat production across western Europe.

Chlorhexidine gluconate usage is associated with antiseptic tolerance in staphylococci from the neonatal intensive care unit.

BackgroundIntravascular catheters are essential for care in Neonatal Intensive Care Units (NICUs) but predispose infants to catheter-associated infections including late-onset sepsis, commonly caused by CoNS. Antiseptics are applied to prevent infection with chlorhexidine (CHG) and octenidine (OCT) the most common agents used.ObjectivesTo investigate the association between antiseptic use and bacterial susceptibility.MethodsCoNS isolates were collected from two NICUs with differing antiseptic regimens: Norwich, UK (using CHG) and Lubeck, Germany (using OCT). CoNS were isolated from different body sites of babies upon admission, and weekly thereafter. Antiseptic susceptibility testing was performed, and a selection underwent genome sequencing.ResultsA total of 1274 isolates were collected. UK isolates (n = 863) were significantly less susceptible than German isolates (n = 411) to both CHG (mean MIC: 20.1 mg/L versus 8.9 mg/L) and OCT (mean MIC: 2.3 mg/L versus 1.6 mg/L). UK isolates taken on admission were more susceptible to CHG than subsequent isolates. No cross-resistance between the agents was seen. Genome sequencing of 122 CoNS showed the most common species to be Staphylococcus epidermidis and Staphylococcus haemolyticus and phylogenetic analysis suggested antiseptic tolerance evolved multiple times in independent lineages. There was no evidence of dominant antiseptic tolerant clones and carriage of genes previously implicated in antimicrobial susceptibility (qac, smr, norA/B), did not correlate with CHG or OCT susceptibility.ConclusionsLong-term CHG use may select for CHG and OCT tolerance in CoNS. This highlights the different potential for separate antiseptic regimens to select for resistance development. This could be an important factor in developing future infection control policies.

Genome-wide analysis provides a deeper understanding of the population structure of the Salmonella enterica serotype Paratyphi B complex in Bangladesh.

The Salmonella enterica serotype Paratyphi B complex causes a wide range of diseases, from gastroenteritis to paratyphoid fever, depending on the biotypes Java and sensu stricto. The burden of Paratyphi B biotypes in Bangladesh is still unknown, as these are indistinguishable by Salmonella serotyping. Here, we conducted the first whole-genome sequencing (WGS) study on 79 Salmonella isolates serotyped as Paratyphi B that were collected from 10 nationwide enteric disease surveillance sites in Bangladesh. Placing these in a global genetic context revealed that these are biotype Java, and the addition of these genomes expanded the previously described PG4 clade containing Bangladeshi and UK isolates. Importantly, antimicrobial resistance (AMR) genes were scarce amongst Bangladeshi S. Java isolates, somewhat surprisingly given the widespread availability of antibiotics without prescription. This genomic information provides important insights into the significance of S. Paratyphi B biotypes in enteric disease and their implications for public health.

Molecular characterization of Deformed wing viruses identified in honeybee (Apis mellifera L.) colonies in Erzincan province of Turkey

Deformed wing virus (DWV), which belongs to the Iflaviridae family, is a devastating virus causing severe financial losses in honey bee colonies all over the world. In 2020 (November), virus-suspected honey bee specimens collected from Erzincan province were screened by molecular assays against DWV infection. In 6 of 12 samples, an expected approximately 700 bp DNA bands were achieved, indicating probably virus-positive. Two randomly selected amplified DNA band suggesting the presence of the virus was further cloned into a pGEM-T Easy vector and electrically transformed into E. coli competent bacteria. Recombinant plasmids obtained from cloning were purified from E. coli and sequenced by next-generation sequencing (NGS). The partial nucleotide sequences of DWV were ascertained to contain 711 nucleotides and these sequences were recorded in the NCBI GenBank with MW962981 and MW962982 accession number. The highest sequence similarity of both isolates ranged from 98.03% to 98.17%, at the nucleotide level. Furthermore, phylogenetic relationships of Erzincan DWV sequences were revealed using 15 unique sequences from different ecological regions registered in the GenBank. Phylogenetic tree displayed that DWV Erzincan isolates are clustered with UK (HM067438), Spain (MK262743), and Sweden (MN746311) isolates. According to our best knowledge, DWV has been reported for the first time in the honey bees from Erzincan province of Turkey by this study.

Changes in Interaction Between Accessory Protein 8 and IL-17RA in UK Isolates Caused by Mutations in the SARS-CoV-2 Open Reading Frame 8

SARS-CoV-2 is the infectous agent of Covid-19, one of the most important health problems of the twenty-first century. IL-17RA is an crucial receptor in the generation of the host immune response. ORF8 is the viral accessory protein of SARS-CoV-2 that suppresses the host immune response. Mutations can alter the viral properties and clinical course of SARS-CoV-2. In this study, we investigated the changes that SARS-CoV-2 ORF8 mutations may cause in the interaction of IL-17RA with ORF8. The study was carried out using 825 complete genome sequences from UK isolates. Mutation analyzes were performed using RDP4 and MEGAX. The protein model was created using the Swiss Model. Protein protein interaction was analyzed by Haddock ver 2.4. Analysis of changes in protein stability was performed using SDM2, mCSM stability and DUET tools. The change in ORF8 - IL-17RA binding affinity before and after the mutation was evaluated using mCSM-PPI2. We detected P30S, R52I, Y73C and L118V mutations in SARS-CoV-2 ORF8. Mutations have been shown to reduce protein stability and affinity. After the mutation, the binding dynamics of ORF8 to IL-17RA were changed. Molecular attachment scores were -78.0±3.4 kcal.mol-1 and -76.3±11.9 kcal.mol-1, for wild type and mutant, respectively. After the mutations, the hydrogen bond number and position between ORF8 and IL-17RA changed. While establishing ten hydrogen bonds between the wild type and IL-17RA, four hydrogen bonds were established between the mutant ORF8 and IL-17RA. The decreased affinity between ORF8 and IL-17RA can be seen as a stronger immune response and a milder clinical course. Although our results contain important data for understanding ORF8, which is an important drug target, it needs to be repeated with in-vivo and crystallgraphy studies.

Antimicrobial resistance progression in the United Kingdom: A temporal comparison of Clostridioides difficile antimicrobial susceptibilities

ObjectivesClostridioides difficile (CD) is widely reported as one of the most prevalent multi-drug resistant (MDR) organisms. Assessment of temporally disparate isolate collections can give valuable epidemiological data to further the understanding of antimicrobial resistance progression. MethodsA collection of 75 CD isolates (1980–86) was characterised by PCR ribotyping, cell cytotoxicity assay and susceptibility testing with a panel of 16 antimicrobials and compared to a modern surveillance collection consisting of 416 UK isolates (2012–2016). Agar-incorporation was performed to ascertain susceptibility data for vancomycin, metronidazole, rifampicin, fidaxomicin, moxifloxacin, clindamycin, imipenem, chloramphenicol, tigecycline, linezolid, ciprofloxacin, piperacillin/tazobactam, ceftriaxone, amoxicillin, tetracycline and erythromycin. Genomes were obtained using Illumina HiSeq3000 sequencing and assembled using CLC Genomics Workbench. Resistance genes were identified using the Comprehensive Antibiotic Research Database's Resistance Gene Identifier and ResFinder3.0. ResultsTwenty-six known and one previously unobserved ribotype (RT) were detected. RT015 and RT020 dominated; 21.3% and 17.3%, respectively. Three moxifloxacin resistant (16–32 mg/L) RT027 isolates were recovered, pre-dating the earliest reports of this phenotype/genotype. Phenotypic resistance was observed to moxifloxacin (9.3% of isolates), ciprofloxacin (100%), erythromycin (17.3%), tetracycline (9.3%), linezolid and chloramphenicol (4.0%). Phenotypic comparisons with modern strains revealed increasing minimum inhibitory concentrations (MIC), with MIC50 elevations of one doubling-dilution for the majority of compounds, excluding clindamycin and imipenem. Moxifloxacin MIC90 comparisons revealed a two doubling-dilution increase between temporal isolate collections. Historical genomes revealed twenty different resistance determinants, including ermB (8.0% of isolates), tetM (9.3%), cfr (5.3%) and gyrA substitution Thr-82→Ile (9.3%). Seventeen isolates (22.7%) were resistant to ≥3 compounds (MDR), demonstrating ten different combinations. Intra-RT diversity was observed. ConclusionsAntibiotic resistance in CD has increased since the early 1980s, across the majority of classes. Moxifloxacin resistance determinants may pre-date its introduction.

Transmission, adaptation and geographical spread of the Pseudomonas aeruginosa Liverpool epidemic strain.

The Liverpool epidemic strain (LES) is an important transmissible clonal lineage of Pseudomonas aeruginosa that chronically infects the lungs of people with cystic fibrosis (CF). Previous studies have focused on the genomics of the LES in a limited number of isolates, mostly from one CF centre in the UK, and from studies highlighting identification of the LES in Canada. Here we significantly extend the current LES genome database by genome sequencing 91 isolates from multiple CF centres across the UK, and we describe the comparative genomics of this large collection of LES isolates from the UK and Canada. Phylogenetic analysis revealed that the 145 LES genomes analysed formed a distinct clonal lineage when compared with the wider P. aeruginosa population. Notably, the isolates formed two clades: one associated with isolates from Canada, and the other associated with UK isolates. Further analysis of the UK LES isolates revealed clustering by clinic geography. Where isolates clustered closely together, the association was often supported by clinical data linking isolates or patients. When compared with the earliest known isolate, LESB58 (from 1988), many UK LES isolates shared common loss-of-function mutations, such as in genes gltR and fleR. Other loss-of-function mutations identified in previous studies as common adaptations during CF chronic lung infections were also identified in multiple LES isolates. Analysis of the LES accessory genome (including genomic islands and prophages) revealed variations in the carriage of large genomic regions, with some evidence for shared genomic island/prophage complement according to clinic location. Our study reveals divergence and adaptation during the spread of the LES, within the UK and between continents.

Legionella antimicrobial sensitivity testing: comparison of microbroth dilution with BCYE and LASARUS solid media.

ObjectivesThere is a lack of international unification for AST methodology for Legionella pneumophila. Current literature contains multiple possible methods and this study compares each of them to determine methodological concordance.MethodsAntibiotic susceptibility of 50 L. pneumophila strains was determined using broth microdilution (BMD), serial antimicrobial dilution in traditional buffered charcoal yeast extract (BCYE) agar (as well as comparison with gradient strip overlay on BCYE) and in a novel charcoal-free agar (LASARUS) for rifampicin, azithromycin, levofloxacin and doxycycline.ResultsThe deviation of tested media relative to BMD highlighted the overall similarity of BMD and LASARUS across all antimicrobials tested (within one serial dilution). BCYE agar dilution showed an increased MIC of up to five serial dilutions relative to BMD, while MICs by gradient strip overlay on BCYE were elevated by two to three serial dilutions, with the exception of doxycycline, which was decreased by three serial dilutions relative to MIC values determined by BMD. The MIC range for azithromycin was wider than for other antimicrobials tested and found to be caused by the presence or absence of the lpeAB gene.ConclusionsBMD-based antimicrobial susceptibility testing (AST) methodology should be the internationally agreed gold standard for Legionella spp. AST, as is common for other bacterial species. Traditional BCYE gave significantly elevated MIC results and its use should be discontinued for Legionella spp., while MIC determination using LASARUS solid medium gave results concordant (within one serial dilution) with BMD for all antimicrobials tested. To the best of our knowledge, this study is the first to identify the lpeAB gene in UK isolates.

Sequence analysis of 43‐year old samples of Plantago lanceolata show that Plantain virus X is synonymous with Actinidia virus X and is widely distributed

AbstractPlantain virus X was first recognized by the ICTV as a species in the genus Potexvirus in 1982. However, because no sequence was available for plantain virus X (PlVX), abolishing the species was proposed to the Flexiviridae working group of the ICTV in 2015. This initiated efforts to sequence the original isolates from Plantago lanceolata samples. Here we report the full‐genome sequencing of two original isolates of PlVX, which demonstrate that the virus is synonymous to Actinidia virus X, a species previously reported from kiwifruit (Actinidia sp.) and blackcurrant (Ribes nigrum). PlVX was previously noted to be widespread in the UK in P. lanceolata. This report additionally presents novel data on the distribution and diversity of PlVX, collected at the same site as the original UK isolates, and from three independent surveys, two in the Netherlands and one in Belgium. This study also includes two new host records for PlVX, Browallia americana and Capsicum annuum (sweet pepper), indicating the virus is more widespread and infects a broader range of hosts than previously reported. This stresses the importance of surveys of noncultivated species to gain insight into viral distribution and host range. This study also demonstrates the value of generating sequence data for isolates retained in virus collections. Additionally, it demonstrates the potential value in prepublication data sharing for giving context to virus detections such as the four independent studies here which, when combined, give greater clarity to the identity, diversity, distribution, and host range of plantain virus X.

Activity of β-lactam/taniborbactam (VNRX-5133) combinations against carbapenem-resistant Gram-negative bacteria.

Boronates are of growing interest as β-lactamase inhibitors. The only marketed analogue, vaborbactam, principally targets KPC carbapenemases, but taniborbactam (VNRX-5133, Venatorx) has a broader spectrum. MICs of cefepime and meropenem were determined combined with taniborbactam or avibactam for carbapenem-resistant UK isolates. β-Lactamase genes and porin alterations were sought by PCR or sequencing. Taniborbactam potentiated partner β-lactams against: (i) Enterobacterales with KPC, other class A, OXA-48-like, VIM and NDM (not IMP) carbapenemases; and (ii) Enterobacterales inferred to have combinations of ESBL or AmpC activity and impermeability. Potentiation of cefepime (the partner for clinical development) by taniborbactam was slightly weaker than by avibactam for Enterobacterales with KPC or OXA-48-like carbapenemases, but MICs of cefepime/taniborbactam were similar to those of ceftazidime/avibactam, and the spectrum was wider. MICs of cefepime/taniborbactam nonetheless remained >8 + 4 mg/L for 22%-32% of NDM-producing Enterobacterales. Correlates of raised cefepime/taniborbactam MICs among these NDM Enterobacterales were a cefepime MIC >128 mg/L, particular STs and, for Escherichia coli only: (i) the particular blaNDM variant (even though published data suggest all variants are inhibited similarly); (ii) inserts in PBP3; and (iii) raised aztreonam/avibactam MICs. Little or no potentiation of cefepime or meropenem was seen for Pseudomonas aeruginosa and Acinetobacter baumannii with MBLs, probably reflecting slower uptake or stronger efflux. Potentiation of cefepime was seen for Stenotrophomonas maltophilia and Elizabethkingia meningoseptica, which have both chromosomal ESBLs and MBLs. Taniborbactam broadly reversed cefepime or meropenem non-susceptibility in Enterobacterales and, less reliably, in non-fermenters.

Geographical Variations in Virulence Factors and Antimicrobial Resistance Amongst Staphylococci Isolated From Dogs From the United Kingdom and Romania.

The objective of this study was to compare virulence and resistance factors of mucosal and cutaneous staphylococci from dogs with pyoderma in the UK and Romania, two countries with different approaches to antimicrobial use in companion animals. Staphylococcal isolates (n = 166) identified to the species level as being Staphylococcus pseudintermedius or coagulase negative (CoNS) were analyzed for their antimicrobial resistance (AMR) profile and presence of resistance and virulence genes. Of the investigated isolates, 26 were methicillin-resistant S. pseudintermedius (MRSP), 89 were methicillin-susceptible S. pseudintermedius (MSSP) and 51 were coagulase negative staphylococci (CoNS). A significantly larger number of isolates originating from Romania were resistant to clindamycin, tetracycline, and chloramphenicol compared to the UK isolates (P < 0.05). Resistance to amoxicillin-clavulanic acid, gentamicin, and trimethoprim-sulphamethoxazole was more evident in UK isolates. Fusidic acid resistance was common in Staphylococcus spp. isolates from both countries. Most isolates carried virulence factors associated with siet (exfoliative toxin) and luk (leucocidin) genes. All MRSP UK isolates exhibited fusidic acid resistance genes whilst this was very rare in the MRSP isolates from Romania. The chlorhexidine resistance gene qacA/B was frequently identified in CoNS isolates from the UK (P < 0.001). The current study documented differences in antimicrobial resistance profiles of Staphylococcus spp. isolates from dogs in two geographical locations in Europe, which could reflect differences in antimicrobial prescribing patterns. The study also highlights the need for further studies and interventions on antimicrobial use, prescribing patterns and AMR surveillance in companion animals in Romania.

Genetic Diversity of Nine Non-Recombinant Potato virus Y Isolates From Three Biological Strain Groups: Historical and Geographical Insights.

Potato virus Y (PVY) isolates from potato currently exist as a complex of six biologically defined strain groups all containing nonrecombinant isolates and at least 14 recombinant minor phylogroups. Recent studies on eight historical UK potato PVY isolates preserved since 1984 found only nonrecombinants. Here, four of five PVY isolates from cultivated potato or wild Solanum spp. collected recently in Australia, Mexico, and the U.S.A. were typed by inoculation to tobacco plants and/or serological testing using monoclonal antibodies. Next, these five modern isolates and four additional historical UK isolates belonging to biological strain groups PVYC, PVYZ, or PVYN obtained from cultivated potato in 1943 to 1984 were sequenced. None of the nine complete PVY genomes obtained were recombinants. Phylogenetic analysis revealed that the four historical UK isolates were in minor phylogroups PVYC1 (YC-R), PVYO-O (YZ-CM1), PVYNA-N (YN-M), or PVYEu-N (YN-RM), Australian isolate YO-BL2 was in minor phylogroup PVYO-O5, and both Mexican isolate YN-Mex43 and U.S.A. isolates YN-MT12_Oth288, YN-MT12_Oth295, and YN-WWAA150131G42 were in minor phylogroup PVYEu-N. When combined, these new findings and those from the eight historical UK isolates sequenced earlier provide important historical insights concerning the diversity of early PVY populations in Europe and the appearance of recombinants in that part of the world. They and four recent Australian isolates sequenced earlier also provide geographical insights about the geographical distribution and diversity of PVY populations in Australia and North America.