SABRes: in silico detection of drug resistance conferring mutations in subpopulations of SARS-CoV-2 genomes.

The emergence of resistance to antiviral drugs increasingly used to treat SARS-CoV-2 infections has been recognised as a significant threat to COVID-19 control. In addition, some SARS-CoV-2 variants of concern appear to be intrinsically resistant to several classes of these antiviral agents. Therefore, there is a critical need for rapid recognition of clinically relevant polymorphisms in SARS-CoV-2 genomes associated with significant reduction of drug activity in virus neutralisation experiments. Here we present SABRes, a bioinformatic tool, which leverages on expanding public datasets of SARS-CoV-2 genomes and allows detection of drug resistance mutations in consensus genomes as well as in viral subpopulations. We have applied SABRes to detect resistance-conferring mutations in 25,197 genomes generated over the course of the SARS-CoV-2 pandemic in Australia and identified 299 genomes containing resistance conferring mutations to the five antiviral therapeutics that retain effectiveness against currently circulating strains of SARS-CoV-2 - Sotrovimab, Bebtelovimab, Remdesivir, Nirmatrelvir and Molnupiravir. These genomes accounted for a 1.18% prevalence of resistant isolates discovered by SABRes, including 80 genomes with resistance conferring mutations found in viral subpopulations. Timely recognition of these mutations within subpopulations is critical as these mutations can provide an advantage under selective pressure and presents an important step forward in our ability to monitor SARS-CoV-2 drug resistance.

Integrative omics identifies conserved and pathogen-specific responses of sepsis-causing bacteria.

Even in the setting of optimal resuscitation in high-income countries severe sepsis and septic shock have a mortality of 20-40%, with antibiotic resistance dramatically increasing this mortality risk. To develop a reference dataset enabling the identification of common bacterial targets for therapeutic intervention, we applied a standardized genomic, transcriptomic, proteomic and metabolomic technological framework to multiple clinical isolates of four sepsis-causing pathogens: Escherichia coli, Klebsiella pneumoniae species complex, Staphylococcus aureus and Streptococcus pyogenes. Exposure to human serum generated a sepsis molecular signature containing global increases in fatty acid and lipid biosynthesis and metabolism, consistent with cell envelope remodelling and nutrient adaptation for osmoprotection. In addition, acquisition of cholesterol was identified across the bacterial species. This detailed reference dataset has been established as an open resource to support discovery and translational research.

Genomic comparisons of Escherichia coli ST131 from Australia.

Escherichia coli ST131 is a globally dispersed extraintestinal pathogenic E. coli lineage contributing significantly to hospital and community acquired urinary tract and bloodstream infections. Here we describe a detailed phylogenetic analysis of the whole genome sequences of 284 Australian ST131 E. coli isolates from diverse sources, including clinical, food and companion animals, wildlife and the environment. Our phylogeny and the results of single nucleotide polymorphism (SNP) analysis show the typical ST131 clade distribution with clades A, B and C clearly displayed, but no niche associations were observed. Indeed, interspecies relatedness was a feature of this study. Thirty-five isolates (29 of human and six of wild bird origin) from clade A (32 fimH41, 2 fimH89, 1 fimH141) were observed to differ by an average of 76 SNPs. Forty-five isolates from clade C1 from four sources formed a cluster with an average of 46 SNPs. Within this cluster, human sourced isolates differed by approximately 37 SNPs from isolates sourced from canines, approximately 50 SNPs from isolates from wild birds, and approximately 52 SNPs from isolates from wastewater. Many ST131 carried resistance genes to multiple antibiotic classes and while 41 (14 %) contained the complete class one integron-integrase intI1, 128 (45 %) isolates harboured a truncated intI1 (462-1014 bp), highlighting the ongoing evolution of this element. The module intI1-dfrA17-aadA5-qacEΔ1-sul1-ORF-chrA-padR-IS1600-mphR-mrx-mphA, conferring resistance to trimethoprim, aminoglycosides, quaternary ammonium compounds, sulphonamides, chromate and macrolides, was the most common structure. Most (73 %) Australian ST131 isolates carry at least one extended spectrum ß-lactamase gene, typically blaCTX-M-15 and blaCTX-M-27. Notably, dual parC-1aAB and gyrA-1AB fluoroquinolone resistant mutations, a unique feature of clade C ST131 isolates, were identified in some clade A isolates. The results of this study indicate that the the ST131 population in Australia carries diverse antimicrobial resistance genes and plasmid replicons and indicate cross-species movement of ST131 strains across diverse reservoirs.

Molecular Analysis of an IncF ColV-Like Plasmid Lineage That Carries a Complex Resistance Locus with a Trackable Genetic Signature.

IncF ColV plasmids are important plasmid incompatibility group that are currently restricted to the Enterobacteriaceae. These plasmids carry an important repertoire of virulence-associated genes (VAGs) that contribute to the ability of avian pathogenic Escherichia coli to cause disease in poultry. VAGs found on ColV plasmids have also been linked to urosepsis and meningitis in humans but the mechanisms that elicit these disease conditions are not well understood. Recently we described the sequence of a ColV plasmid pSDJ2009-52F that carried the typical repertoire of VAGs and a complex resistance gene locus flanked by IS26, an insertion element that plays an important role in mobilizing antibiotic resistance genes on plasmids and genomic islands. We recovered complete ColV-like plasmid sequences from public databases that shared >80% sequence identity with pSDJ2009-52F in geographically diverse regions of the world over a 20-year timeframe. Previously we noted that pSDJ2009-52F carries a unique genetic signature in the class 1 integron within the complex resistance locus that was presumably created by the action of IS26. Here we show that most ColV-like plasmids that are closely related to pSDJ2009-52F also carry the same signature. Our studies provide insight into how these signature-bearing plasmids and the mobile genetic elements they carry traffic between E. coli sequence types over large geographic distances.

Clonal ST131-H22 Escherichia coli strains from a healthy pig and a human urinary tract infection carry highly similar resistance and virulence plasmids.

The interplay between food production animals, humans and the environment with respect to the transmission of drug-resistant pathogens is widely debated and poorly understood. Pandemic uropathogenic Escherichia coli ST131-H30Rx, with conserved fluoroquinolone and cephalosporin resistance, are not frequently identified in animals. However, the phylogenetic precursor lineage ST131-H22 in animals and associated meat products is being reported with increasing frequency. Here we characterized two highly related ST131-H22 strains, one from a healthy pig and the other from a human infection (in 2007 and 2009, respectively). We used both long and short genome sequencing and compared them to ST131-H22 genome sequences available in public repositories. Even within the context of H22 strains, the two strains in question were highly related, separated by only 20 core SNPs. Furthermore, they were closely related to a faecal strain isolated in 2010 from a geographically distinct, healthy human in New South Wales, Australia. The porcine and hospital strains carried highly similar HI2-ST3 multidrug resistant plasmids with differences in the hospital strain arising due to IS-mediated insertions and rearrangements. Near identical ColV plasmids were also present in both strains, further supporting their shared evolutionary history. This work highlights the importance of adopting a One Health approach to genomic surveillance to gain insights into pathogen evolution and spread.

Genomic analysis of multidrug-resistant Escherichia coli ST58 causing urosepsis.

Sequence type 58 (ST58) phylogroup B1 Escherichia coli have been isolated from a wide variety of mammalian and avian hosts but are not noted for their ability to cause serious disease in humans or animals. Here we determined the genome sequences of two multidrug-resistant E. coli ST58 strains from urine and blood of one patient using a combination of Illumina and Single Molecule, Real-Time (SMRT) sequencing. Both ST58 strains were clonal and were characterised as serotype O8:H25, phylogroup B1 and carried a complex resistance locus/loci (CRL) that featured an atypical class 1 integron with a dfrA5 (trimethoprim resistance) gene cassette followed by only 24 bp of the 3'-CS. CRL that carry this particular integron have been described previously in E. coli from cattle, pigs and humans in Australia. The integron abuts a copy of Tn6029, an IS26-flanked composite transposon encoding blaTEM, sul2 and strAB genes that confer resistance to ampicillin, sulfathiazole and streptomycin, respectively. The CRL resides within a novel Tn2610-like hybrid Tn1721/Tn21 transposon on an IncF, ColV plasmid (pSDJ2009-52F) of 138 553 bp that encodes virulence associated genes implicated in life-threatening extraintestinal pathogenic E. coli (ExPEC) infections. Notably, pSDJ2009-52F shares high sequence identity with pSF-088-1, a plasmid reported in an E. coli ST95 strain from a patient with blood sepsis from a hospital in San Francisco. These data suggest that extraintestinal infections caused by E. coli carrying ColV-like plasmids, irrespective of their phylogroup or ST, may pose a potential threat to human health, particularly to the elderly and immunocompromised.

Multidrug Resistant Uropathogenic Escherichia coli ST405 With a Novel, Composite IS26 Transposon in a Unique Chromosomal Location.

Escherichia coli ST405 is an emerging urosepsis pathogen, noted for carriage of bla CTX-M, bla NDM, and a repertoire of virulence genes comparable with O25b:H4-ST131. Extraintestinal and multidrug resistant E. coli ST405 are poorly studied in Australia. Here we determined the genome sequence of a uropathogenic, multiple drug resistant E. coli ST405 (strain 2009-27) from the mid-stream urine of a hospital patient in Sydney, Australia, using a combination of Illumina and SMRT sequencing. The genome of strain 2009-27 assembled into two unitigs; a chromosome comprising 5,287,472 bp and an IncB/O plasmid, pSDJ2009-27, of 89,176 bp. In silico and phenotypic analyses showed that strain 2009-27 is a serotype O102:H6, phylogroup D ST405 resistant to ampicillin, azithromycin, kanamycin, streptomycin, trimethoprim, and sulphafurazole. The genes encoding resistance to these antibiotics reside within a novel, mobile IS26-flanked transposon, identified here as Tn6242, in the chromosomal gene yjdA. Tn6242 comprises four modules that each carries resistance genes flanked by IS26, including a class 1 integron with dfrA17 and aadA5 gene cassettes, a variant of Tn6029, and mphA. We exploited unique genetic signatures located within Tn6242 to identify strains of ST405 from Danish patients that also carry the transposon in the same chromosomal location. The acquisition of Tn6242 into yjdA in ST405 is significant because it (i) is vertically inheritable; (ii) represents a reservoir of resistance genes that can transpose onto resident/circulating plasmids; and (iii) is a site for the capture of further IS26-associated resistance gene cargo.

A draft genome of Escherichia coli sequence type 127 strain 2009-46.

BACKGROUND: Escherichia coli are a frequent cause of urinary tract infections (UTI) and are thought to have a foodborne origin. E. coli with sequence type 127 (ST127) are emerging pathogens increasingly implicated as a cause of urinary tract infections (UTI) globally. A ST127 isolate (2009-46) resistant to ampicillin and trimethoprim was recovered from the urine of a 56 year old patient with a UTI from a hospital in Sydney, Australia and was characterised here. RESULTS: We sequenced the genome of Escherichia coli 2009-46 using the Illumina Nextera XT and MiSeq technologies. Assembly of the sequence data reconstructed a 5.14 Mbp genome in 89 scaffolds with an N50 of 161 kbp. The genome has extensive similarity to other sequenced uropathogenic E. coli genomes, but also has several genes that are potentially related to virulence and pathogenicity that are not present in the reference E. coli strain. CONCLUSION: E. coli 2009-46 is a multiple antibiotic resistant, phylogroup B2 isolate recovered from a patient with a UTI. This is the first description of a drug resistant E. coli ST127 in Australia.

Genomic interplay in bacterial communities: implications for growth promoting practices in animal husbandry.

The discovery of antibiotics heralded the start of a "Golden Age" in the history of medicine. Over the years, the use of antibiotics extended beyond medical practice into animal husbandry, aquaculture and agriculture. Now, however, we face the worldwide threat of diseases caused by pathogenic bacteria that are resistant to all existing major classes of antibiotic, reflecting the possibility of an end to the antibiotic era. The seriousness of the threat is underscored by the severely limited production of new classes of antibiotics. Evolution of bacteria resistant to multiple antibiotics results from the inherent genetic capability that bacteria have to adapt rapidly to changing environmental conditions. Consequently, under antibiotic selection pressures, bacteria have acquired resistance to all classes of antibiotics, sometimes very shortly after their introduction. Arguably, the evolution and rapid dissemination of multiple drug resistant genes en-masse across microbial pathogens is one of the most serious threats to human health. In this context, effective surveillance strategies to track the development of resistance to multiple antibiotics are vital to managing global infection control. These surveillance strategies are necessary for not only human health but also for animal health, aquaculture and plant production. Shortfalls in the present surveillance strategies need to be identified. Raising awareness of the genetic events that promote co-selection of resistance to multiple antimicrobials is an important prerequisite to the design and implementation of molecular surveillance strategies. In this review we will discuss how lateral gene transfer (LGT), driven by the use of low-dose antibiotics in animal husbandry, has likely played a significant role in the evolution of multiple drug resistance (MDR) in Gram-negative bacteria and has complicated molecular surveillance strategies adopted for predicting imminent resistance threats.

The importance of teaching and learning resilience in the health disciplines: a critical review of the literature.

This paper, based on a review of the literature, discusses resilience and the application of resilience research to nursing education. The paper advances the educational discourse on professional preparation, arguing that resilience theory should be part of the educational content and taught in a way that promotes reflection and application in order to give students strength, focus and endurance in the workplace. In addition, we argue that resilience and similar qualities ought to be emphasised in clinical experience courses, internships, work integrated learning and other work experience courses. Recommendations for building resilience in health professionals through education, training and modifications in workplace culture, as well as suggestions for priorities in research are presented.