This study established the in vitro activity of ceftolozane/tazobactam (C/T) and its genotypic resistance mechanisms by whole-genome sequencing (WGS) in 195 carbapenem-nonsusceptible Pseudomonas aeruginosa (CNSPA) clinical isolates recovered from Singapore between 2009 and 2020. C/T susceptibility rates were low, at 37.9%. Cross-resistance to ceftazidime/avibactam was observed, although susceptibility to the agent was slightly higher, at 41.0%. Whole-genome sequencing revealed that C/T resistance was largely mediated by the presence of horizontally acquired β-lactamases, especially metallo-β-lactamases. These were primarily disseminated in well-recognized high-risk clones belonging to sequence types (ST) 235, 308, and 179. C/T resistance was also observed in several non-carbapenemase-producing isolates, in which resistance was likely mediated by β-lactamases and, to a smaller extent, mutations in AmpC-related genes. There was no obvious mechanism of resistance observed in five isolates. The high C/T resistance highlights the limited utility of the agent as an empirical agent in our setting. Knowledge of local molecular epidemiology is crucial in determining the potential of therapy with novel agents. IMPORTANCE Pseudomonas aeruginosa infection is one of the most difficult health care-associated infections to treat due to the ability of the organism to acquire a multitude of resistance mechanisms and express the multidrug resistance phenotype. Ceftolozane/tazobactam (C/T), a novel β-lactam/β-lactamase inhibitor combination, addresses an unmet medical need in patients with these multidrug-resistant P. aeruginosa infections. Our findings demonstrate geographical variation in C/T susceptibility owing to the distinct local molecular epidemiology. This study adds on to the growing knowledge of C/T resistance, particularly mutational resistance, and will aid in the design of future β-lactams and β-lactamase inhibitors. WGS proved to be a useful tool to understand the P. aeruginosa resistome and its contribution to emerging resistance in novel antimicrobial agents.

In this study, we characterized carbapenem-resistant Klebsiella pneumoniae clinical isolates collected over a 12-year period in the largest public acute-care hospital in Singapore using whole-genome sequencing. The results of this study demonstrate significant genomic diversity with the presence of well-known epidemic, multidrug-resistant clones amid a diverse pool of nonepidemic lineages.

Genomic Characterisation of Carbapenem Non-susceptible Pseudomonas aeruginosa in SingaporePseudomonas aeruginosa is a clinically important pathogen implicated in many hospitalacquired infections. Its propensity to acquire broad-spectrum resistance has earned the organism its status as a severe public health threat requiring urgent control measures.While whole genome sequencing-based genomic surveillance provides a mean to track antimicrobial resistance, its use in molecular epidemiological surveys of Pseudomonas aeruginosa remains limited, especially in the Southeast Asian region. We sequenced the whole genomes of 222 carbapenem non-susceptible P. aeruginosa (CNPA) isolates collected in 2006-2020 at the largest public acute care hospital in Singapore . Antimicrobial susceptibilities were determined using broth microdilution. Clonal relatedness, multi-locus sequence types and antimicrobial resistance determina nts (acquired and chromosomal) were determined. In this study, CNPA exhibited broadspectrum resistance (87.8% multi-drug resistance), retaining susceptibility only to polymyxin B (95.0%) and amikacin (55.0%). Carbapenemases were detected in 51.4% of the isolates, where IMP and NDM metallo-β-lactamases were the most frequent.Carbapenem resistance was also likely associated with OprD alterations or efflux mechanisms (ArmZ/NalD mutations), which occurred in strains with or without carbapenemases. The population of CNPA in the hospital was diverse; the 222 isolates grouped into 68 sequence types (ST), which includes various high-risk clones. We detected an emerging clone, the NDM-1-producing ST308, in addition to the global highrisk ST235 clone which was the predominant clone in our population. Our results thus provide a "snapshot" of the circulating lineages of CNPA locally and the prevailing 4 genetic mechanisms contributing to carbapenem resistance. This database also serves as the baseline for future prospective surveillance studies.

Summary Mycobacterial interspersed repetitive unit-variable number tandem repeat (MIRU-VNTR) typing is widely used to genotype Mycobacterium tuberculosis complex in epidemiological studies for tracking tuberculosis transmission. Recent long-read sequencing technologies from Pacific Biosciences and Oxford Nanopore Technologies can produce reads that are long enough to cover the entire repeat regions in each MIRU-VNTR locus which was previously not possible using the short reads from Illumina high-throughput sequencing technologies. We thus developed MIRUReader for MIRU-VNTR typing directly from long sequence reads. Availability and implementation Source code and documentation for MIRUReader program is freely available at https://github.com/phglab/MIRUReader. Supplementary information Supplementary data are available at Bioinformatics online.

This study determined the susceptibilities of carbapenem-resistant Enterobacterales isolates to various novel antimicrobial agents (ceftazidime-avibactam, imipenem-relebactam, meropenem-vaborbactam, eravacycline, and plazomicin). Whole-genome sequencing was performed for all strains.

No abstract

Objectives To investigate the transmission and origination of MRSA in livestock with limited antimicrobial use. Yak (Bos grunniens) herds in Ganzi Tibetan Autonomous Prefecture, China were chosen for sampling. Methods The yaks from all 18 districts of Ganzi were sampled (anal swabs, n = 657; nasal swabs, n = 634). Based on the WGS data of 83 Staphylococcus aureus isolates, the novel structure of the yak S. aureus population was described. Phylogenetic analyses were utilized for determining the origin of the MRSA lineage in yaks. Results The yak S. aureus population consisted of 11 STs, 6 of which were previously undescribed, with ST6267 being the predominant novel ST. These isolates were generally susceptible to most of the tested antibiotics and lacked the associated antimicrobial resistance genes (ARGs) but showed high penicillin MIC values (MIC90 = 32 mg/L), which were consistent with the high positivity rate for blaZ (61/83). The MRSA identified in yaks were all ST59 and most likely of human origin. The yak ST59 MRSA each carried the human immune evasion cluster (IEC) while lacking the ARGs that are identified in the majority of reported Chinese human ST59 MRSA isolates [erm(B), ant6-Ia and aph(3″)-III]. Conclusions The yak herds living on the Qinghai–Tibet Plateau are important livestock and follow the traditional free-grazing farming model. We surveyed the yak S. aureus population and found that all the yak MRSA isolates belonged to the lineage that might originate from the prevalent community-acquired MRSA ST59 in China. From a ‘One Health’ perspective, the transmission of human MRSA to farming animals with limited antimicrobial exposure highlights the multiple roles of animals in the expansion of MRSA.

Objectives To explore the association of recent hospitalization and asymptomatic carriage of multidrug-resistant Enterobacterales (MDRE) and determine the prevailing strains and antibiotic resistance genes in Siem Reap, Cambodia using WGS. Methods In this cross-sectional study, faecal samples were collected from two arms: a hospital-associated arm consisted of recently hospitalized children (2–14 years), with their family members; and a community-associated arm comprising children in the matching age group and their family members with no recent hospitalization. Forty-two families in each study arm were recruited, with 376 enrolled participants (169 adults and 207 children) and 290 stool specimens collected from participants. The DNA of ESBL- and carbapenemase-producing Enterobacterales cultured from the faecal samples was subject to WGS on the Illumina NovaSeq platform. Results Of the 290 stool specimens, 277 Escherichia coli isolates and 130 Klebsiella spp. were identified on CHROMagar ESBL and KPC plates. The DNA of 276 E. coli (one isolate failed quality control test), 89 Klebsiella pneumoniae, 40 Klebsiella quasipneumoniae and 1 Klebsiella variicola was sequenced. CTX-M-15 was the most common ESBL gene found in E. coli (n = 104, 38%), K. pneumoniae (n = 50, 56%) and K. quasipneumoniae (n = 16, 40%). The prevalence of bacterial lineages and ESBL genes was not associated with any specific arm. Conclusions Our results demonstrate that MDRE is likely to be endemic within the Siem Reap community. ESBL genes, specifically blaCTX-M, can be found in almost all E. coli commensals, indicating that these genes are continuously propagated in the community through various unknown channels at present.