ABSTRACT Eleven clinical class 1 integron-containing Pseudomonas aeruginosa isolates from Australia and Uruguay were investigated for the genomic locations of these elements. Several novel class 1 integrons/transposons were found in at least four distinct locations in the chromosome, including genomic islands. These elements seem to be undergoing successful dispersal by lateral gene transfer since integrons were identified across several lineages and more than one clonal line.
ABSTRACT Acinetobacter baumannii , an important pathogen known for its widespread antibiotic resistance, has been the focus of extensive research within its genus, primarily involving clinical isolates. Consequently, data on environmental A. baumannii and other Acinetobacter species remain limited. Here, we utilised Illumina and Nanopore sequencing to analyse the genomes of ten Acinetobacter isolates representing six different species sourced from aquatic environments in South Australia. All ten isolates were phylogenetically distinct compared to clinical and other non-clinical Acinetobacter strains, often tens of thousands of SNPs from their nearest neighbours. Despite the genetic divergence, we identified p dif modules (sections of mobilised DNA) carrying clinically important antimicrobial resistance genes in species other than A. baumannii , including carbapenemase oxa58, tetracycline resistance gene tet ( 39 ) and macrolide resistance genes msr(E)-mph(E). All of these p dif modules were located on plasmids with high sequence homology to those circulating in globally distributed A. baumannii ST1 and ST2 clones. The environmental A. baumannii isolate characterised here (SAAb472; ST350) did not possess any native plasmids; however, it could capture two clinically important plasmids (pRAY and pACICU2) with high transfer frequencies. Furthermore, A. baumannii SAAb472 possessed virulence genes and a capsular polysaccharide type analogous to clinical strains. Our findings highlight the potential for environmental Acinetobacter species to acquire and disseminate clinically important antimicrobial resistance genes, underscoring the need for further research into the ecology and evolution of this important genus. IMPORTANCE Antimicrobial resistance (AMR) is a global threat to human, animal, and environmental health. Studying AMR in environmental bacteria is crucial to understand the emergence and dissemination of resistance genes and pathogens, and to identify potential reservoirs and transmission routes. This study provides novel insights into the genomic diversity and AMR potential of environmental Acinetobacter species. By comparing the genomes of aquatic Acinetobacter isolates with clinical and non-clinical strains, we revealed that they are highly divergent yet carry p dif modules that encode resistance to antibiotics commonly used in clinical settings. We also demonstrated that an environmental A. baumannii isolate can acquire clinically relevant plasmids and carries virulence factors similar to those of hospital-associated strains. These findings suggest that environmental Acinetobacter species may serve as reservoirs and vectors of clinically important genes. Consequently, further research is warranted to comprehensively understand the ecology and evolution of this genus.
Of the 200+ serogroups of Vibrio cholerae , only O1 or O139 strains are reported to cause cholera, and mostly in endemic regions. Cholera outbreaks elsewhere are considered to be via importation of pathogenic strains. Using established animal models, we show that diverse V. cholerae strains indigenous to a non-endemic environment (Sydney, Australia), including non-O1/O139 serogroup strains, are able to both colonize the intestine and result in fluid accumulation despite lacking virulence factors believed to be important. Most strains lacked the type three secretion system considered a mediator of diarrhoea in non-O1/O13 V. cholerae . Multi-locus sequence typing (MLST) showed that the Sydney isolates did not form a single clade and were distinct from O1/O139 toxigenic strains. There was no correlation between genetic relatedness and the profile of virulence-associated factors. Current analyses of diseases mediated by V. cholerae focus on endemic regions, with only those strains that possess particular virulence factors considered pathogenic. Our data suggest that factors other than those previously well described are of potential importance in influencing disease outbreaks.
The spread of antimicrobial resistance (AMR) is a rapidly growing threat to humankind on both regional and global scales. As countries worldwide prepare to embrace a One Health approach to AMR management, which is one that recognizes the interconnectivity between human, animal, and environmental health, increasing attention is being paid to identifying and monitoring key contributing factors and critical control points. Presently, AMR sensing technologies have significantly progressed phenotypic antimicrobial susceptibility testing (AST) and genotypic antimicrobial resistance gene (ARG) detection in human healthcare. For effective AMR management, an evolution of innovative sensing technologies is needed for tackling the unique challenges of interconnected AMR across various and different health domains. This review comprehensively discusses the modern state-of-play for innovative commercial and emerging AMR sensing technologies, including sequencing, microfluidic, and miniaturized point-of-need platforms. With a unique view toward the future of One Health, we also provide our perspectives and outlook on the constantly changing landscape of AMR sensing technologies beyond the human health domain.
We investigated the infectivity, growth ability and the overall biochemical status of Rhizobium purine auxotrophs to determine why these strains show varied nodulation phenotypes on their host plants. Strains ANU2861 (purM::Tn5) and ANU2866 (purY::Tn5), both derivatives of ANU280, are unable to nodulate siratro plants. In contrast, strain L1 (purF::Tn5ssgusA40), a derivative of ANU843, is able to induce fully-developed nodules and normal levels of nodulation on several clover hosts. To our knowledge, strain L1 is the only genetically-defined pur– mutant capable of inducing fully-developed nodules on a legume host. None of the pur– mutants are able to grow at normal rates in vitro even in the presence of purine pathway supplements including AICA-riboside. Further, the nodulation ability of the strain ANU280 pur– mutants is unable to be restored by the addition of AICA-riboside. Full growth and nodulation ability can only be restored to the pur– mutants by genetic complementation. We confirmed that supplementation of pur intermediates failed to fully restore wild-type growth by using proteome analysis to examine the overall biochemical status of the mutants. Proteome analysis demonstrated that the purine mutants possess multiple changes in the protein species present. In addition, strain ANU2866 but not strains ANU2861 or L1 could not tolerate the effects of nutrient step-up or step-down transitions, and failed to produce any colonies on laboratory media. Collectively, these data show that purine auxotrophs suffer pleiotropic effects at the level of protein synthesis and their overall metabolism is compromised. Therefore, care should be exercised in concluding that purine intermediates or by-products of this pathway are required per se for nodule development.
Salmonella genomic island 1 (SGI1) is an integrative mobilisable element that plays an important role in the capture and spread of multiple drug resistance. To date, SGI1 has been found in clinical isolates of Salmonellaenterica serovars, Proteus mirabilis, Morganellamorganii, Acinetobacterbaumannii, Providenciastuartii, Enterobacterspp, and recently in Escherichia coli. SGI1 preferentially targets the 3´-end of trmE, a conserved gene found in the Enterobacteriaceae and among members of the Gammaproteobacteria. It is, therefore, hypothesised that SGI1 and SGI1-related elements (SGI1-REs) may have been acquired by diverse bacterial genera. Here, Bitsliced Genomic Signature Indexes (BIGSI) was used to screen the NCBI Sequence Read Archive (SRA) for putative SGI1-REs in Gammaproteobacteria. Novel SGI-REs were identified in diverse genera including Cronobacter spp, Klebsiella spp, and Vibrio spp and in two additional isolates of Escherichia coli. An extensively drug-resistant human clonal lineage of Klebsiella pneumoniae carrying an SGI1-RE in the United Kingdom and an SGI1-RE that lacks a class 1 integron were also identified. These findings provide insight into the origins of this diverse family of clinically important genomic islands and expand the knowledge of the potential host range of SGI1-REs within the Gammaproteobacteria.
Silver gulls carry phylogenetically diverse Escherichia coli, including globally dominant extraintestinal pathogenic E. coli (ExPEC) sequence types and pandemic ExPEC-ST131 clades; however, our large-scale study (504 samples) on silver gulls nesting off the coast of New South Wales identified E. coli ST457 as the most prevalent. A phylogenetic analysis of whole-genome sequences (WGS) of 138 ST457 samples comprising 42 from gulls, 2 from humans (Australia), and 14 from poultry farmed in Paraguay were compared with 80 WGS deposited in public databases from diverse sources and countries. E. coli ST457 strains are phylogenetic group F, carry fimH145, and partition into five main clades in accordance to predominant flagella H-antigen carriage. Although we identified considerable phylogenetic diversity among the 138 ST457 strains, closely related subclades (<100 SNPs) suggested zoonotic or zooanthroponosis transmission between humans, wild birds, and food-producing animals. Australian human clinical and gull strains in two of the clades were closely related (≤80 SNPs). Regarding plasmid content, country, or country/source, specific connections were observed, including I1/ST23, I1/ST314, and I1/ST315 disseminating blaCMY-2 in Australia, I1/ST113 carrying blaCTX-M-8 and mcr-5 in Paraguayan poultry, and F2:A-:B1 plasmids of Dutch origin being detected across multiple ST457 clades. We identified a high prevalence of nearly identical I1/ST23 plasmids carrying blaCMY-2 among Australian gull and clinical human strains. In summary, ST457 is a broad host range, geographically diverse E. coli lineage that can cause human extraintestinal disease, including urinary tract infection, and displays a remarkable ability to capture mobile elements that carry and transmit genes encoding resistance to critically important antibiotics.
There are currently multiple methods available for the preparation of fresh frozen tissue samples for analysis via matrix-assisted laser desorption/ionisation time-of-flight (MALDI-TOF) imaging mass spectrometry (IMS). Although these methods report excellent results, many are expensive automated approaches. With no published attempt to standardise less expensive manual processes, our work aims to provide a robust and repeatable method of sample preparation for MALDI-TOF-IMS that is applicable to a variety of tissue types, well explained, simple and cost effective.Fresh frozen tissue was sectioned at 12 µm and mounted onto liquid nitrocellulose coated slides, washed in a graded alcohol series and then mounted into a modified sublimation apparatus. Matrix is deposited onto the slide to achieve a desired coating of 0.2 mg/cm(2). Once coated, the slide is mounted into a custom-built vapor chamber and recrystallised with 50% acetonitrile (ACN), 0.1% trifluoroacetic acid (TFA) for 1 h at 37°C. The slide is then analysed using MALDI-IMS.We have successfully implemented this method for a host of tissue samples, including brain, liver, kidney and heart, with no variation in relative spectra or processing method required. When the protocol is followed correctly, sublimations and recrystallisations are highly predictable with limited variation between samples and a very low failure rate. Additional apparatuses can be easily constructed by following the included instructions, that perform as per specifications with no variation.We believe that we have described a complete protocol for MALDI-IMS that is easy to use and highly reproducible. The lack of expensive commercially available equipment makes this process very cheap with a relatively low initial outlay and our hope is that more laboratories will begin IMS-based avenues of research based on the work we have performed.
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