Genomic epidemiology and antimicrobial resistance of Neisseria gonorrhoeae in New Zealand.

Background: Antimicrobial-resistant Neisseria gonorrhoeae is a major threat to public health. No studies to date have examined the genomic epidemiology of gonorrhoea in the Western Pacific Region, where the incidence of gonorrhoea is particularly high. Methods: A population-level study of N. gonorrhoeae in New Zealand (October 2014 to May 2015). Comprehensive susceptibility testing and WGS data were obtained for 398 isolates. Relatedness was inferred using phylogenetic trees, and pairwise core SNPs. Mutations and genes known to be associated with resistance were identified, and correlated with phenotype. Results: Eleven clusters were identified. In six of these clusters, >25% of isolates were from females, while in eight of them, >15% of isolates were from females. Drug resistance was common; 98%, 32% and 68% of isolates were non-susceptible to penicillin, ciprofloxacin and tetracycline, respectively. Elevated MICs to extended-spectrum cephalosporins (ESCs) were observed in 3.5% of isolates (cefixime MICs ≥ 0.12 mg/L, ceftriaxone MICs ≥ 0.06 mg/L). Only nine isolates had penA XXXIV genotypes, three of which had decreased susceptibility to ESCs (MIC = 0.12 mg/L). Azithromycin non-susceptibility was identified in 43 isolates (10.8%); two of these isolates had 23S mutations (C2611T, 4/4 alleles), while all had mutations in mtrR or its promoter. Conclusions: The high proportion of females in clusters suggests transmission is not exclusively among MSM in New Zealand; re-assessment of risk factors for transmission may be warranted in this context. As elevated MICs of ESCs and/or azithromycin were found in closely related strains, targeted public health interventions to halt transmission are urgently needed.

Antimicrobial Resistance in Selected Bacteria from Food Animals in New Zealand 2018-2022.

Antimicrobial resistance (AMR) presents a significant threat to human health worldwide. One important source of antimicrobial-resistant infections in humans is exposure to animals or animal products. In a phased survey, we investigated AMR in 300 Escherichia coli isolates and 300 enterococci (Enterococcus faecalis and E. faecium) isolates each from the carcasses of poultry, pigs, very young calves, and dairy cattle (food animals); all Salmonella isolates from poultry, very young calves, and dairy cattle; and 300 Campylobacter (Campylobacter jejuni and C. coli) isolates from poultry. The highest resistance levels in E. coli were found for sulfamethoxazole, tetracycline, and streptomycin, for all food animals. Cefotaxime-resistant E. coli were not found and low resistance to ciprofloxacin, colistin, and gentamicin was observed. The majority of enterococci isolates from all food animals were bacitracin-resistant. Erythromycin- and/or tetracycline-resistant enterococci isolates were found in varying proportions from all food animals. Ampicillin- or vancomycin-resistant enterococci isolates were not identified, and ciprofloxacin-resistant E. faecalis were not found. Salmonella isolates were only recovered from very young calves and all eight isolates were susceptible to all tested antimicrobials. Most Campylobacter isolates were susceptible to all tested antimicrobials, although 16.6% of C. jejuni were resistant to quinolones and tetracycline. Results suggest that AMR in E. coli, enterococci, Salmonella, and Campylobacter isolates from food animals in New Zealand is low, and currently, AMR in food animals poses a limited public health risk. Despite the low prevalence of AMR in this survey, ongoing monitoring of antimicrobial susceptibility in bacteria from food animals is recommended, to ensure timely detection of AMR with potential impacts on animal and human health.

Daptomycin Resistance Occurs Predominantly in vanA-Type Vancomycin-Resistant Enterococcus faecium in Australasia and Is Associated With Heterogeneous and Novel Mutations.

Healthcare associated infections caused by vancomycin-resistant Enterococcus faecium (VREfm) have a major impact on health outcomes. VREfm is difficult to treat because of intrinsic and acquired resistance to many clinically used antimicrobials, with daptomycin being one of the few last line therapeutic options for treating multidrug-resistant VREfm. The emergence of daptomycin-resistant VREfm is therefore of serious clinical concern. Despite this, the impact that daptomycin-resistant VREfm have on patient health outcomes is not clearly defined and knowledge on the mechanisms and genetic signatures linked with daptomycin resistance in VREfm remains incomplete. To address these knowledge gaps, phenotypic daptomycin susceptibility testing was undertaken on 324 E. faecium isolates from Australia and New Zealand. Approximately 15% of study isolates were phenotypically resistant to daptomycin. Whole genome sequencing revealed a strong association between vanA-VREfm and daptomycin resistance, with 95% of daptomycin-resistant study isolates harbouring vanA. Genomic analyses showed that daptomycin-resistant VREfm isolates were polyclonal and carried several previously characterised mutations in the liaR and liaS genes as well as several novel mutations within the rpoB, rpoC, and dltC genes. Overall, 70% of daptomycin-resistant study isolates were found to carry mutations within the liaR, rpoB, rpoC, or dltC genes. Finally, in a mouse model of VREfm bacteraemia, infection with the locally dominant daptomycin-resistant clone led to reduced daptomycin treatment efficacy in comparison to daptomycin-susceptible E. faecium. These findings have important implications for ongoing VREfm surveillance activities and the treatment of VREfm infections.

Salivaricin G32, a Homolog of the Prototype Streptococcus pyogenes Nisin-Like Lantibiotic SA-FF22, Produced by the Commensal Species Streptococcus salivarius.

Salivaricin G32, a 2667 Da novel member of the SA-FF22 cluster of lantibiotics, has been purified and characterized from Streptococcus salivarius strain G32. The inhibitory peptide differs from the Streptococcus pyogenes-produced SA-FF22 in the absence of lysine in position 2. The salivaricin G32 locus was widely distributed in BLIS-producing S. salivarius, with 6 (23%) of 26 strains PCR-positive for the structural gene, slnA. As for most other lantibiotics produced by S. salivarius, the salivaricin G32 locus can be megaplasmid encoded. Another member of the SA-FF22 family was detected in two Streptococcus dysgalactiae of bovine origin, an observation supportive of widespread distribution of this lantibiotic within the genus Streptococcus. Since the inhibitory spectrum of salivaricin G32 includes Streptococcus pyogenes, its production by S. salivarius, either as a member of the normal oral microflora or as a commercial probiotic, could serve to enhance protection of the human host against S. pyogenes infection.

Sequence variation in the porB gene from B:P1.4 meningococci causing New Zealand's epidemic.

Since mid-1991, New Zealand has experienced an epidemic of meningococcal disease. The epidemic has been caused by serogroup B meningococci expressing PorA type P1.7-2,4, belonging to the ST-41/ST-44 complex, lineage III. Most B:P1.7-2,4 meningococci express type 4 PorB (87.0%), although case isolates with porB other than type 4 have been identified throughout the duration of the epidemic. To assess the genetic relatedness of case isolates with an alternative porB gene, multilocus restriction typing validated against multilocus sequence typing was used. This determined that B:P1.7-2,4 meningococci with a porB gene that was other than type 4 had the same clonal origin. It was concluded that strains with alternative porB genes had diverged from the original type 4 porB. Variation in porB was also shown to be associated with the uptake of DNA encoding one or two of the PorB variable regions leading to mosaic porB. Point mutation rather than horizontal transfer and recombination was implicated as the mechanism of sequence variation in some strains. This work will serve as a reference point to determine if the administration of a strain-specific vaccine increases the level of porB divergence and variation already observed in New Zealand case isolates. It also complements the study undertaken of PorA stability which showed that variation in P1.7-2,4 PorA was almost exclusively due to deletions in the P1.4 epitope of the epidemic strain.

Evaluation of porB PCR-amplicon restriction endonuclease analysis as a method to determine porB variable-region sequences in nonserotypeable meningococci.

porB PCR-amplicon restriction endonuclease analysis is a rapid, simple method developed to assess porB variation in nonserotypeable meningococci isolated during New Zealand's epidemic of meningococcal disease. Most nonserotypeable meningococci isolated between 1990 and 1999 inclusively either were type 4 (40.5%) or contained the porB variable region 1 (VR1)-19, VR2-D, VR3-7, and VR4-14a sequences (45.1%).

Multilocus restriction typing method to predict the sequence type of meningococci.

A multilocus restriction typing (MLRT) method was developed to reduce the number of sequencing reactions required to determine the clonal relationships among serogroup B meningococci causing an epidemic in New Zealand. MLRT was a rapid, simple, and inexpensive method, and the results had an excellent correlation with multilocus sequence typing results.