Ceftriaxone-resistant Neisseria gonorrhoeae detected in England, 2015-24: an observational analysis.

OBJECTIVES: Since June 2022, there has been a rise in the number of ceftriaxone-resistant Neisseria gonorrhoeae cases detected in England (n = 15), of which a third were XDR. We describe the demographic and clinical details of the recent cases and investigate the phenotypic and molecular characteristics of the isolates. For a comprehensive overview, we also reviewed 16 ceftriaxone-resistant cases previously identified in England since December 2015 and performed a global genomic comparison of all publicly available ceftriaxone-resistant N. gonorrhoeae strains with mosaic penA alleles. METHODS: All N. gonorrhoeae isolates resistant to ceftriaxone (MIC > 0.125 mg/L) were whole-genome sequenced and compared with 142 global sequences of ceftriaxone-resistant N. gonorrhoeae. Demographic, behavioural and clinical data were collected. RESULTS: All cases were heterosexual, and most infections were associated with travel from the Asia-Pacific region. However, some had not travelled outside England within the previous few months. There were no ceftriaxone genital treatment failures, but three of five pharyngeal infections and the only rectal infection failed treatment. The isolates represented 13 different MLST STs, and most had the mosaic penA-60.001 allele. The global genomes clustered into eight major phylogroups, with regional associations. All XDR isolates belonged to the same phylogroup, represented by MLST ST16406. CONCLUSIONS: Most cases of ceftriaxone-resistant N. gonorrhoeae detected in England were associated with travel from the Asia-Pacific region. All genital infections were successfully treated with ceftriaxone, but there were extragenital treatment failures. Ceftriaxone resistance continues to be associated with the penA-60.001 allele within multiple genetic backgrounds and with widespread dissemination in the Asia-Pacific region.

Molecular Basis of Antimicrobial Resistance in Group B Streptococcus Clinical Isolates from Saudi Arabia.

Published data on the molecular mechanisms underlying antimicrobial resistance in Group B Streptococcus (GBS) isolates from Saudi Arabia are lacking. Here, we aimed to determine the genetic basis of resistance to relevant antibiotics in a collection of GBS clinical isolates (n = 204) recovered from colonized adults or infected patients and expressing serotypes Ia, Ib, II, III, V, and VI. Initial susceptibility testing revealed resistance to tetracycline (76.47%, n = 156/204), erythromycin (36.76%, n = 75/204), clindamycin (25.49%, n = 52/204), levofloxacin (6.37%, n = 13/204), and gentamicin (2.45%, n = 5/204). Primers designed for the detection of known resistance determinants in GBS identified the presence of erm(A), erm(B), mef(A), and/or lsa(C) genes at the origin of resistance to macrolides and/or clindamycin. Of these, erm(B) and erm(A) were associated with the cMLSB (n = 46) and iMLSB (n = 28) phenotypes, respectively, while mef(A) was linked to the M phenotype (n = 1) and lsa(C) was present in isolates with the L phenotype (n = 8). Resistance to tetracycline was mainly mediated by tet(M) alone (n = 112) or in combination with tet(O) (n = 10); the remaining isolates carried tet(O) (n = 29), tet(L) (n = 2), or both (n = 3). Isolates resistant to gentamicin (n = 5) carried aac(6')-Ie-aph(2')-Ia, and those exhibiting resistance to levofloxacin (n = 13) had alterations in GyrA and/or ParC. Most isolates with the erm gene (93.24%, n = 69/74) also had the tet gene and were therefore resistant to erythromycin, clindamycin, and tetracycline. Overall, there were no clear associations between serotypes and resistance genotypes except for the presence of erm(B) in serotype Ib isolates. Dissemination of antibiotic resistance genes across different serotypes represents a public health concern that requires further surveillance and appropriate antibiotic use in clinical practice.

Genomic insights into the diversity, virulence, and antimicrobial resistance of group B Streptococcus clinical isolates from Saudi Arabia.

Introduction: Detailed assessment of the population structure of group B Streptococcus (GBS) among adults is still lacking in Saudi Arabia. Here we characterized a representative collection of isolates from colonized and infected adults. Methods: GBS isolates (n=89) were sequenced by Illumina and screened for virulence and antimicrobial resistance determinants. Genetic diversity was assessed by single nucleotide polymorphisms and core-genome MLST analyses. Results: Genome sequences revealed 28 sequence types (STs) and nine distinct serotypes, including uncommon serotypes VII and VIII. Majority of these STs (n=76) belonged to the human-associated clonal complexes (CCs) CC1 (33.71%), CC19 (25.84%), CC17 (11.24%), CC10/CC12 (7.87%), and CC452 (6.74%). Major CCs exhibited intra-lineage serotype diversity, except for the hypervirulent CC17, which exclusively expressed serotype III. Virulence profiling revealed that nearly all isolates (94.38%) carried at least one of the four alpha family protein genes (i.e., alphaC, alp1, alp2/3, and rib), and 92.13% expressed one of the two serine-rich repeat surface proteins Srr1 or Srr2. In addition, most isolates harbored the pilus island (PI)-2a alone (15.73%) or in combination with PI-1 (62.92%), and those carrying PI-2b alone (10.11%) belonged to CC17. Phylogenetic analysis grouped the sequenced isolates according to CCs and further subdivided them along with their serotypes. Overall, isolates across all CC1 phylogenetic clusters expressed Srr1 and carried the PI-1 and PI-2a loci, but differed in genes encoding the alpha-like proteins. CC19 clusters were dominated by the III/rib/srr1/PI-1+PI-2a (43.48%, 10/23) and V/alp1/srr1/PI-1+PI-2a (34.78%, 8/23) lineages, whereas most CC17 isolates (90%, 9/10) had the same III/rib/srr2/P1-2b genetic background. Interestingly, genes encoding the CC17-specific adhesins HvgA and Srr2 were detected in phylogenetically distant isolates belonging to ST1212, suggesting that other highly virulent strains might be circulating within the species. Resistance to macrolides and/or lincosamides across all major CCs (n=48) was associated with the acquisition of erm(B) (62.5%, 30/48), erm(A) (27.1%, 13/48), lsa(C) (8.3%, 4/48), and mef(A) (2.1%, 1/48) genes, whereas resistance to tetracycline was mainly mediated by presence of tet(M) (64.18%, 43/67) and tet(O) (20.9%, 14/67) alone or in combination (13.43%, 9/67). Discussion: These findings underscore the necessity for more rigorous characterization of GBS isolates causing infections.

Molecular typing and antimicrobial resistance of group B Streptococcus clinical isolates in Saudi Arabia.

OBJECTIVES: Group B Streptococcus (GBS) has emerged as an important cause of severe infections in adults. However, limited data are available regarding the epidemiology of GBS in Saudi Arabia. METHODS: Isolates were collected over a period of eight months from colonized (n = 104) and infected adults (n = 95). Serotypes and virulence determinants were detected by polymerase chain reactions (PCRs). Genetic relatedness was assessed using Multiple Locus Variable Number Tandem Repeat Analysis (MLVA). Antimicrobial susceptibilities were determined by disk diffusion. RESULTS: Serotypes III and V (25% each) were the most prevalent, followed by serotypes II (16.18%), Ia (13.24%), VI (9.31%), and Ib (8.82%), while five isolates remained non-typeable (2.45%). Hypervirulent serotype III/CC17 clone (n = 21) accounted for 41.18% of the serotype III isolates. Most isolates (53.92%) harboured pilus island (PI) 1 and 2a types, while PI-2b was predominantly detected in the hypervirulent clone. Isolates were variably resistant to tetracycline (76.47%), erythromycin (36.76%), clindamycin (25.49%), and levofloxacin (6.37%), but remained susceptible to penicillin. Macrolide resistant isolates exhibited constitutive (55.42%) and inducible macrolide-lincosamide-streptogramin B resistance phenotypes (33.74%), while a few had L (9.64%) or M (1.2%) phenotypes. MLVA patterns of dominant serotypes III and V revealed 40 different types divided into 12 clusters and 28 singletons. Interestingly, macrolide resistance was significantly associated with two major MLVA types. CONCLUSIONS: GBS isolates belonged predominantly to serotypes III and V, but there were no clear associations between serotypes and patient groups. The studied isolates exhibited high levels of resistance to erythromycin and clindamycin that need further surveillance.

Molecular characterization and differential effects of levofloxacin and ciprofloxacin on the potential for developing quinolone resistance among clinical Pseudomonas aeruginosa isolates.

Background: Fluoroquinolones are some of the most used antimicrobial agents for the treatment of Pseudomonas aeruginosa. This study aimed at exploring the differential activity of ciprofloxacin and levofloxacin on the selection of resistance among P. aeruginosa isolates at our medical center. Methods: 233 P. aeruginosa clinical isolates were included in this study. Antimicrobial susceptibility testing (AST) was done using disk diffusion and broth microdilution assays. Random Amplification of Polymorphic DNA (RAPD) was done to determine the genetic relatedness between the isolates. Induction of resistance against ciprofloxacin and levofloxacin was done on 19 isolates. Fitness cost assay was done on the 38 induced mutants and their parental isolates. Finally, whole genome sequencing was done on 16 induced mutants and their 8 parental isolates. Results: AST results showed that aztreonam had the highest non-susceptibility. RAPD results identified 18 clusters. The 19 P. aeruginosa isolates that were induced against ciprofloxacin and levofloxacin yielded MICs ranging between 16 and 256 µg/mL. Levofloxacin required fewer passages in 10 isolates and the same number of passages in 9 isolates as compared to ciprofloxacin to reach their breakpoints. Fitness cost results showed that 12 and 10 induced mutants against ciprofloxacin and levofloxacin, respectively, had higher fitness cost when compared to their parental isolates. Whole genome sequencing results showed that resistance to ciprofloxacin and levofloxacin in sequenced mutants were mainly associated with alterations in gyrA, gyrB and parC genes. Conclusion: Understanding resistance patterns and risk factors associated with infections is crucial to decrease the emerging threat of antimicrobial resistance.

In vitro activity of cefiderocol against clinically important carbapenem non-susceptible Gram-negative bacteria from Saudi Arabia.

OBJECTIVES: Cefiderocol is a novel catechol-substituted siderophore cephalosporin with broad-spectrum activity against Gram-negative pathogens. However, variation of its activity among carbapenemase producers from various regions and countries has been reported. Here, we checked the in vitro activity against Gram-negative carbapenem non-susceptible bacteria collected in Saudi Arabia. METHODS: Cefiderocol MICs were determined using the iron-depleted cation-adjusted Mueller-Hinton broth and interpreted according to the Clinical and Laboratory Standards Institute guidelines. Isolates (n = 288) included carbapenemase-producing Escherichia coli (n = 46), Klebsiella pneumoniae (n = 98), Acinetobacter baumannii (n = 65), and Pseudomonas aeruginosa (n = 79) clinical isolates. RESULTS: Cefiderocol inhibited 73.26% (211/288) of the isolates studied at concentrations of ≤ 4 mg/L. Cefiderocol inhibited all carbapenem-resistant A. baumannii isolates (65/65, 100%) producing OXA-23-like, OXA-24-like, and NDM, and nearly all P. aeruginosa isolates (75/79, 94.94%), including those producing VIM and NDM. In contrast, the carbapenemase-producing isolates from the Enterobacterales group demonstrated significantly higher MICs with only 53.06% (52/98) of K. pneumoniae and 41.3% (19/46) of E. coli isolates exhibiting MICs of ≤4 mg/L. Isolates showing elevated MICs (73/144, 50.69%) included NDM (20/29, 68.97%), NDM/OXA-48-like (34/59, 57.63%), OXA-48-like (18/52, 34.62%), and KPC (1/4, 25%) producers, thus showing no clear association with the production of serine-type or metallo-type carbapenemases. However, high cefiderocol MICs (≥ 32mg/L) were associated with isolates producing NDM, and in particular, among those coproducing the OXA-232-type enzyme. CONCLUSIONS: Cefiderocol had excellent activity against multi-drug resistant non-fermenting Gram-negative pathogens. Reasons behind the high cefiderocol MICs in certain Enterobacterales isolates need further investigation.

Epididymo-Orchitis Caused by POM-1 Metallo-ß-Lactamase-Producing Pseudomonas otitidis in an Immunocompetent Patient: Case Report and Molecular Characterization.

Pseudomonas otitidis is a rare and unique species among the Pseudomonas genus that has not been previously reported as a cause of male genitourinary tract infection. In this report, we describe a case of a 20-year-old immunocompetent male who presented with recurrent epididymo-orchitis, which was initially misidentified as Vibrio vulnificus and treated successfully. The causative agent could not be identified appropriately using the available routine methods, but a final identification was established using 16S rRNA targeted sequencing followed by whole-genome sequencing.

Epidemiological and molecular description of nosocomial outbreak of COVID-19 Alpha (B.1.1.7) variant in Saudi Arabia.

BACKGROUND: Nosocomial outbreaks frequently occurred during the Coronavirus disease 2019 (COVID-19) pandemic; however, sharing experiences on outbreak containment is vital to reduce the related burden in different locations. OBJECTIVES: This article aims at sharing a practical experience on COVID-19 outbreak containment, including contact tracing, screening of target population, testing including molecular analysis, and preventive modalities. It also provides an epidemiological and molecular analysis of the Severe Acute Respiratory Syndrome Coronavirus 2 (SARS­CoV­2) infection outbreak in a tertiary care hospital in Saudi Arabia. METHODS: The outbreak occurred in a non-COVID medical ward at a tertiary care hospital in Jeddah, Saudi Arabia, from 22nd March and 15th April 2021. The multidisciplinary outbreak response team performed clinical and epidemiological investigations. Whole-Genome Sequencing (WGS) was implemented on selected isolates for further molecular characterization. RESULTS: A total of eight nurses (20 % of the assigned ward nurses) and six patients (16.2 % of the ward admitted patients at the time of the outbreak) tested positive for the SARS-CoV-2 virus based on PCR testing. The outbreak investigation identified strong evidence of an epidemiologic link between the affected cases. WGS revealed a set of spike mutations and deletions specific to the Alpha variant (B.1.1.7 lineage). All the nurses had mild symptoms, and the fatality among the patients was 50 % (three out of the six patients). CONCLUSIONS: The current nosocomial COVID-19 outbreak, caused by the Alpha variant, revealed multiple breaches in the adherence to the hospital infection control recommended measures. Containment strategies were successful in controlling the outbreak and limiting infection spread. Molecular analysis and genome sequencing are essential tools besides epidemiological investigation to inform appropriate actions, especially with emerging pathogens.

Diversity of carbapenemase-producing Enterobacterales in England as revealed by whole-genome sequencing of isolates referred to a national reference laboratory over a 30-month period.

Introduction. Increasing numbers of carbapenemase-producing Enterobacterales (CPE), which can be challenging to treat, have been referred to the national reference laboratory in England since the early 2000s.Gap Statement/Aim. Previous studies on CPE in the UK have focussed on localized outbreaks. We applied whole-genome sequencing (WGS) to isolates referred to the national reference laboratory over 30 months to inform our understanding of CPE epidemiology in England.Methodology. The first confirmed CPE from each new patient referred by an English diagnostic laboratory between 1 January 2014 and 30 June 2016 was sequenced on an Illumina HiSeq 2500. Multiple isolates from the same patient were included from either different species or the same species with different carbapenemase genes. The data were analysed using an in-house bioinformatics pipeline that determines species identification, multi-locus sequence typing (MLST) profile and antimicrobial resistance gene content.Results. A total of 2658 non-duplicate CPE were sequenced amongst which three host organisms belonging to diverse sequence types (STs) predominated: Klebsiella pneumoniae (1380/2658, 51.9 %; 177 STs), Escherichia coli (723/2658, 27.2 %; 133 STs) and Enterobacter cloacae (294/2658, 11.1 %; 88 STs). Thirty different carbapenemase gene variants were identified, although bla OXA-48-like (1122/2658, 42.2%), bla NDM (692/2658, 26.0 %), bla KPC (571/2658, 21.5 %), bla VIM (100/2658, 3.8 %) and bla IMP (33/2658, 1.2 %) predominated. ST/carbapenemase gene pairings represented widely distributed high-risk clones or clusters at a regional or hospital level.Conclusion. CPE referred to the national reference laboratory are diverse, suggesting multiple introductions to England and a role for horizontal transfer of carbapenemase genes in English CPE epidemiology.

Acquired resistome and plasmid sequencing of mcr-1 carrying MDR Enterobacteriaceae from poultry and their relationship to STs associated with humans.

OBJECTIVES: To investigate the acquired resistome in 18 colistin-resistant Escherichia coli isolated from different poultry farms in Lebanon, analyse Inc plasmids associated with mcr and assess potential transmission to humans. METHODS: A total of 18 E. coli were recovered from poultry faeces collected from different poultry farms in Lebanon. Broth microdilution (BMD) assay was performed to determine the antimicrobial resistance profiles. WGS was used to identify the genetic determinants behind the resistance in these isolates. RESULTS: BMD results showed that all of the 18 isolates were colistin resistant. Furthermore, resistance to trimethoprim/sulfamethoxazole was the most recorded among the isolates and only one isolate was resistant to cefepime. Sequencing results showed that the isolates were distributed into seven different STs and that the most abundant was ST1140. The number of antimicrobial resistant determinants ranged from 4 to 21 among the 18 isolates, with tet(A) and floR being the most frequent. Moreover, a total of 15 different plasmid replicon types were identified. The mcr-1 gene was shown to be predominantly located on IncX4 plasmids. Additionally, two isolates harboured the IncI2-type self-conjugative plasmid. CONCLUSIONS: The findings show that mcr and other important resistance determinants occur in MDR E. coli isolated poultry farms in Lebanon. The occurrence of mcr on mobile plasmids and the zoonotic potential and clinical relevance of some strains highlight a risk of transmission to humans.

Invasive fungal infection of the brain caused by Neoscytalidium dimidiatum in a post-renal transplant patient: A case report.

Neoscytalidium is a phytopathogen that is often found in plants and soil. It mostly leads to skin and nail infections, and invasive diseases of the sinuses, lung, and brain have been described mostly in immunocompromised patients. We report a case of a post-renal transplant patient who received anti-thymocyte globulin for induction immunosuppression. A month after her transplant, she presented with fever and new-onset seizures, and computed tomography revealed a brain abscess with mass effects and herniation. The patient underwent abscess drainage and craniectomy. The pathological findings showed filamentous septate hyphae. The surgical culture rapidly grew wool-like colonies with a black reverse on Sabouraud agar. Lactophenol cotton blue staining showing septate branched hyphae with one to two arthroconidia cells with flattened ends. The patient was given a combination of amphotericin B and voriconazole but unfortunately died ten days after the diagnosis. This case highlights Neoscytalidium as a cause of invasive fungal disease in immunocompromised patients that is difficult to treat and is often fatal, even when combined surgical and medical therapies are used as treatment modalities.

Successful treatment of infective endocarditis due to pandrug-resistant Klebsiella pneumoniae with ceftazidime-avibactam and aztreonam.

Pandrug-resistant (PDR) K. pneumoniae refractory to conventional treatment has been reported worldwide, causing a huge burden on the healthcare system, patient safety and the economy. K. pneumoniae is a prominent opportunistic pathogen causing hospital-acquired and community-acquired infections, but is rarely associated with infective endocarditis. Currently, there are sparse data guiding the optimal regimen when commonly used antibiotics fail, notably for the treatment of endocarditis infections. Here we report our experience in treating a 40-year-old female with PDR K. pneumoniae infection of cardiovascular implantable electronic device (CIED) and right-sided infective endocarditis. Initial susceptibility testing of the incriminated pathogen showed an apparent susceptibility to colistin but the prolonged course of colistin, gentamicin and meropenem did not resolve the infection. However, the synergistic combinations of aztreonam with ceftazidime-avibactam was able to overcome resistance and clear the infection rapidly. Genome sequencing showed that the PDR K. pneumoniae isolate belongs to the international high-risk clone ST14. The isolate harbored genes encoding NDM-1, OXA-48, CTX-M-14b, SHV-28 and OXA-1, explaining resistance to all ß-lactams, including carbapenems. It carried the armA gene conferring resistance to all clinically important aminoglycosides and had alterations in GyrA, ParC and MgrB, explaining resistance to ciprofloxacin and colistin.

Genomic Characterization of Carbapenem-Non-susceptible Pseudomonas aeruginosa Clinical Isolates From Saudi Arabia Revealed a Global Dissemination of GES-5-Producing ST235 and VIM-2-Producing ST233 Sub-Lineages.

Carbapenem-resistant P. aeruginosa has become a major clinical problem due to limited treatment options. However, studies assessing the trends in the molecular epidemiology and mechanisms of antibiotic resistance in this pathogen are lacking in Saudi Arabia. Here, we reported the genome characterization in a global context of carbapenem non-susceptible clinical isolates from a nationally representative survey. The antibiotic resistance profiles of the isolates (n = 635) collected over 14 months between March 2018 and April 2019 from different geographical regions of Saudi Arabia showed resistance rates to relevant ß-lactams, aminoglycosides and quinolones ranging between 6.93 and 27.56%. Overall, 22.52% (143/635) of the isolates exhibited resistance to both imipenem and meropenem that were mainly explained by porin loss and efflux overexpression. However, 18.18% of resistant isolates harbored genes encoding GES (69.23%), VIM (23.07%), NDM (3.85%) or OXA-48-like (3.85%) carbapenemases. Most common GES-positive isolates produced GESs -5, -15 or -1 and all belonged to ST235 whereas the VIM-positive isolates produced mainly VIM-2 and belonged to ST233 or ST257. GES and VIM producers were detected at different sampling periods and in different surveyed regions. Interestingly, a genome-wide comparison revealed that the GES-positive ST235 and VIM-2-positive ST233 genomes sequenced in this study and those available through public databases from various locations worldwide, constituted each a phylogenetically closely related sub-lineage. Profiles of virulence determinants, antimicrobial resistance genes and associated mobile elements confirmed relatedness within each of these two different sub-lineages. Sequence analysis located the bla GES gene in nearly all studied genomes (95.4%) in the same integrative conjugative element that also harbored the acc(6')-Ib, aph(3')-XV, aadA6, sul1, tet(G), and catB resistance genes while bla VIM-2 in most (98.89%) ST233-positive genomes was co-located with aac(6')-I1, dfrB-5, and aac(3')-Id in the same class I integron. The study findings revealed the global spread of GES-5 ST235 and VIM-2 ST233 sub-lineages and highlighted the importance of routine detection of rare ß-lactamases.

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

BACKGROUND: Boronates are of growing interest as ß-lactamase inhibitors. The only marketed analogue, vaborbactam, principally targets KPC carbapenemases, but taniborbactam (VNRX-5133, Venatorx) has a broader spectrum. METHODS: MICs of cefepime and meropenem were determined combined with taniborbactam or avibactam for carbapenem-resistant UK isolates. ß-Lactamase genes and porin alterations were sought by PCR or sequencing. RESULTS: Taniborbactam potentiated partner ß-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. CONCLUSIONS: Taniborbactam broadly reversed cefepime or meropenem non-susceptibility in Enterobacterales and, less reliably, in non-fermenters.

The characterization of mobile colistin resistance (mcr) genes among 33 000 Salmonella enterica genomes from routine public health surveillance in England.

To establish the prevalence of mobile colistin resistance (mcr) genes amongst Salmonella enterica isolates obtained through public health surveillance in England (April 2014 to September 2017), 33 205 S. enterica genome sequences obtained from human, food, animal and environmental isolates were screened for the presence of mcr variants 1 to 8. The mcr-positive genomes were assembled, annotated and characterized according to plasmid type. Nanopore sequencing was performed on six selected isolates with putative novel plasmids, and phylogenetic analysis was used to provide an evolutionary context for the most commonly isolated clones. Fifty-two mcr-positive isolates were identified, of which 32 were positive for mcr-1, 19 for mcr-3 and 1 for mcr-5. The combination of Illumina and Nanopore sequencing identified three novel mcr-3 plasmids and one novel mcr-5 plasmid, as well as the presence of chromosomally integrated mcr-1 and mcr-3. Monophasic S. enterica serovar Typhimurium accounted for 27/52 (52 %) of the mcr-positive isolates, with the majority clustering in clades associated with travel to Southeast Asia. Isolates in these clades were associated with a specific plasmid range and an additional extended-spectrum beta-lactamase genotype. Routine whole-genome sequencing for public health surveillance provides an effective screen for novel and emerging antimicrobial determinants, including mcr. Complementary long-read technologies elucidated the genomic context of resistance determinants, offering insights into plasmid dissemination and linkage to other resistance genes.

Genomic analysis of the first KPC-producing Klebsiella pneumoniae isolated from a patient in Riyadh: A new public health concern in Saudi Arabia.

Klebsiella pneumoniae carbapenemase (KPC)-producing Enterobacteriaceae have been isolated from human patients in many countries across the globe but rarely in Saudi Arabia. Here we provide the genomic characterization of the first KPC-producing K. pneumoniae isolated from the urine of a patient in Riyadh, Saudi Arabia, who had a recent travel history to Egypt involving a medical procedure. Presence of KPC-encoding gene initially detected with the Xpert Carba-R assay was confirmed by traditional PCR. Susceptibility testing using the VITEK 2 system, E-test and microbroth dilution methods showed that the K. pneumoniae isolate, namely SA01_KPC-2, was resistant to all antibiotics except colistin and ceftazidime/avibactam. Whole-genome sequencing (WGS), performed on the Illumina Miseq instrument, identified the isolate as sequence type (ST) 383 and serotype KL30-D1 O1v2. Genome assembly of SA01_KPC-2 indicated the presence of two plasmids. Plasmid pSA01_KPC-2, of approximately 45.9 kb in size, harbored the blaKPC-2 flanked by ISKpn27 and ISKpn6 and had a backbone similar to published KPC-carrying IncX6 plasmids. The second plasmid pSA01_incHIB1, a derivative of published 372-kb plasmid pKpvST383, carried genes encoding virulence factors and resistance to five classes of antibiotics. The isolation of the first KPC producer in Saudi Arabia requires high attention and rapid interventions to prevent further spread.

Comparison of phenotypic and WGS-derived antimicrobial resistance profiles of Campylobacter jejuni and Campylobacter coli isolated from cases of diarrhoeal disease in England and Wales, 2015-16.

OBJECTIVES: To compare and evaluate phenotypic and genotypic methods for the detection of antimicrobial resistance (AMR) in Campylobacter jejuni and Campylobacter coli in England and Wales. METHODS: WGS data from 528 isolates of Campylobacter spp. (452 C. jejuni and 76 C. coli) from human (494), food (21) and environmental (2) sources, collected between January 2015 and December 2016, and from the PHE culture collection (11) were mapped to genes known to be associated with phenotypic resistance to antimicrobials in the genus. Phenotypic antibiotic susceptibility (erythromycin, ciprofloxacin, tetracycline, gentamicin and streptomycin) testing using an in-agar dilution method was performed on all isolates. RESULTS: Concordance between phenotypic resistance and the presence of corresponding AMR determinants was 97.5% (515/528 isolates). Only 13 out of 528 isolates (10 C. jejuni and 3 C. coli) had discordant interpretations for at least one of the five antibiotics tested, equating to a total of 15 (0.6%) discrepancies out of 2640 isolate/antimicrobial combinations. Seven discrepant results were genotypically resistant but phenotypically susceptible (major errors) and eight discrepant results were genotypically susceptible but phenotypically resistant (very major errors). CONCLUSIONS: The use of this bioinformatics approach for predicting AMR from WGS data for routine public health surveillance is a reliable method for real-time monitoring of changing AMR patterns in isolates of C. jejuni and C. coli.

Extended-spectrum ß-lactamase-producing Escherichia coli in human-derived and foodchain-derived samples from England, Wales, and Scotland: an epidemiological surveillance and typing study.

BACKGROUND: Extended-spectrum ß-lactamase-producing Escherichia coli isolates (ESBL-E coli) cause more than 5000 cases of bacteraemias annually in the UK. The contribution of the food chain to these infections is debated. We aimed to identify the most important reservoirs of ESBL-E coli that colonise and infect humans to identify strategic intervention points. METHODS: Sampling for ESBL-E coli was done between Aug 1, 2013, and Dec 15, 2014. We used selective media to seek ESBL-E coli in routinely submitted samples from human faeces, and prospectively collected samples from sewage, farm slurry, and retail foodstuffs in London, East Anglia, northwest England, Scotland, and Wales. We sequenced recovered isolates and compared these isolates with 293 bloodstream and 83 veterinary surveillance ESBL-E coli isolates from the same regions. FINDINGS: 2157 (11%) of 20 243 human faeces samples contained ESBL-E coli, including 678 (17%) of 3995 in London. ESBL-E coli also were frequent in sewage and retail chicken (104 [65%] of 159 meat samples), but were rare in other meats and absent from plant-based foods (0 of 400 fruit and vegetable samples). Sequence type (ST) 131 dominated among ESBL-E coli from human blood (188 [64%] of 293 isolates), faeces (128 [36%] of 360), and sewage (14 [22%] of 65) with STs 38 and 648 also widespread; CTX-M-15 was the predominant ESBL in these lineages (319 [77%] of 416). By contrast, STs 602, 23, and 117-mostly with CTX-M-1 ESBL-dominated among food and veterinary isolates (68 [31%] of 218), with only two ST131 organisms recovered. ST10 occurred in both animals and humans, being frequent in surveillance bovines (11 [22%] of 51 cattle) and representing 15 (4%) of 360 human faecal isolates (but only three [1%] of 293 from bacteraemias); however, both human and animal ST10 isolates were diverse in serotype. INTERPRETATION: Most human bacteraemias with ESBL-E coli in the UK involve internationally prevalent human-associated STs, particularly ST131; non-human reservoirs made little contribution to invasive human disease. Any interventions that seek to target food or livestock can affect the numbers of human infections caused by ESBL-E coli; prevention of the spread of resistant lineages among humans is more vital. FUNDING: NIHR Policy Research.