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2.
Antimicrob Agents Chemother ; : AAC0120021, 2021 Sep 20.
Artigo em Inglês | MEDLINE | ID: mdl-34543095

RESUMO

Typhoid fever is an invasive bacterial disease of humans that disproportionately affects low- and middle-income countries. Antimicrobial resistance (AMR) has been increasingly prevalent in recent decades in Salmonella enterica serovar Typhi (S. Typhi), the causative agent of typhoid fever, limiting treatment options. In Australia most cases of typhoid fever are imported due to travel to regions where typhoid fever is endemic. Here, all 116 isolates of S. Typhi isolated in Victoria, Australia between 1st July 2018 and 30th June 2020, underwent whole genome sequencing and antimicrobial susceptibility testing. Genomic data were linked to international travel data collected from routine case interviews. Travel to South Asia accounted for most cases, with 92.2% imported from seven primary countries (the top two were India n = 87 and Pakistan n = 12). A total of 17 S. Typhi genotypes were detected in the two-year cohort, with 47.4% genotyped as part of global AMR lineages. Ciprofloxacin resistance was detected in two lineages, 3.3 and 4.3.1.2, all from cases with reported travel to India. Nearly all multidrug and extensively drug resistant isolates (90%) were from cases with reported travel to Pakistan in genotypes 4.3.1.1 and 4.3.1.1.P1. Extended spectrum beta-lactamases, blaCTX-M-15 and blaSHV-12, were detected in cases with travel to Pakistan and India, respectively. Linking epidemiological data with genomic studies of S. Typhi provides an opportunity to improve understanding of the emergence, spread and risk of drug-resistant S. Typhi infections and to better inform empiric treatment guidelines in returned travellers.

3.
ACS Biomater Sci Eng ; 7(10): 4982-4990, 2021 10 11.
Artigo em Inglês | MEDLINE | ID: mdl-34521204

RESUMO

The ability to detect SARS-CoV-2 is critical to implementing evidence-based strategies to address the COVID-19 global pandemic. Expanding SARS-CoV-2 diagnostic ability beyond well-equipped laboratories widens the opportunity for surveillance and control efforts. However, such advances are predicated on the availability of rapid, scalable, accessible, yet high-performance diagnostic platforms. Methods to detect viral RNA using reverse transcription loop-mediated isothermal amplification (RT-LAMP) show promise as rapid and field-deployable tests; however, the per-unit costs of the required diagnostic hardware can be a barrier for scaled deployment. Here, we describe a diagnostic hardware configuration for LAMP technology, named the FABL-8, that can be built for approximately US$380 per machine and provide results in under 30 min. Benchmarking showed that FABL-8 has a similar performance to a high-end commercial instrument for detecting fluorescence-based LAMP reactions. Performance testing of the instrument with RNA extracted from a SARS-CoV-2 virus dilution series revealed an analytical detection sensitivity of 50 virus copies per microliter-a detection threshold suitable to detect patient viral load in the first few days following symptom onset. In addition to the detection of SARS-CoV-2, we show that the system can be used to detect the presence of two bacterial pathogens, demonstrating the versatility of the platform for the detection of other pathogens. This cost-effective and scalable hardware alternative allows democratization of the instrumentation required for high-performance molecular diagnostics, such that it could be available to laboratories anywhere-supporting infectious diseases surveillance and research activities in resource-limited settings.

4.
Pathology ; 53(6): 689-699, 2021 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-34425991

RESUMO

Emerging testing technologies for detection of SARS-CoV-2 include those that are rapid and can be used at point-of-care (POC), and those facilitating high throughput laboratory-based testing. Tests designed to be performed at POC (such as antigen tests and molecular assays) have the potential to expedite isolation of infectious patients and their contacts, but most are less sensitive than standard-of-care reverse transcription polymerase chain reaction (RT-PCR). Data on clinical performance of the majority of emerging assays are limited with most evaluations performed on contrived or stored laboratory samples. Further evaluations of these assays are required, particularly when performed at POC on symptomatic and asymptomatic patients and at various time-points after symptom onset. A few studies have so far shown several of these assays have high specificity. However, large prospective evaluations are needed to confirm specificity, particularly before the assays are implemented in low prevalence settings or asymptomatic populations. High throughput laboratory-based testing includes the use of new sample types (e.g., saliva to increase acceptability) or innovative uses of existing technology (e.g., sample pooling). Information detailing population-wide testing strategies for SARS-COV-2 is largely missing from peer-reviewed literature. Logistics and supply chains are key considerations in any plan to 'scale up' testing in the Australian context. The strategic use of novel assays will help strike the balance between achieving adequate test numbers without overwhelming laboratory capacity. To protect testing of high-risk populations, the aims of testing with respect to the phase of the pandemic must be considered.


Assuntos
Teste para COVID-19/métodos , COVID-19/diagnóstico , Austrália , Humanos , SARS-CoV-2
5.
ACS Biomater Sci Eng ; 7(9): 4669-4676, 2021 09 13.
Artigo em Inglês | MEDLINE | ID: mdl-34437802

RESUMO

The COVID-19 pandemic has exposed the dependence of diagnostic laboratories on a handful of large corporations with market monopolies on the worldwide supply of reagents, consumables, and hardware for molecular diagnostics. Global shortages of key consumables for RT-qPCR detection of SARS-CoV-2 RNA have impaired the ability to run essential, routine diagnostic services. Here, we describe a workflow for rapid detection of SARS-CoV-2 RNA in upper respiratory samples including nasal swabs and saliva, utilizing low-cost equipment and readily accessible reagents. Using repurposed Creality3D Ender-3 three-dimensional (3D) printers, we built a semiautomated paramagnetic bead RNA extraction platform. The hardware for the system was built for $300 USD, and the material cost per reaction was $1 USD. Named the Ender VX500, instrument performance when paired with RT-qPCR for SARS-CoV-2 detection in nasal and saliva specimens was two virus copies per microliter. There was a high-performance agreement (assessed using 458 COVID-19 nasal swab specimens) with the Aptima SARS-CoV-2 assay run on the Hologic Panther, a commercial automated RNA extraction and detection platform. Inter- and intrainstrument precision was excellent (coefficients of variation (CoV) of 1.10 and 0.66-1.32%, respectively) across four instruments. The platform is scalable with throughput ranging from 23 specimens on a single instrument run by one user in 50 min to 364 specimens on four instruments run by four users in 190 min. Step-by-step instructions and protocols for building and running the Ender VX500 have been made available without restriction.


Assuntos
COVID-19 , Humanos , Pandemias , Patologia Molecular , RNA Viral/genética , SARS-CoV-2
6.
Nat Commun ; 12(1): 4786, 2021 08 09.
Artigo em Inglês | MEDLINE | ID: mdl-34373455

RESUMO

Salmonella enterica serovar 4,[5],12:i:- (Salmonella 4,[5],12:i:-) is a monophasic variant of Salmonella Typhimurium that has emerged as a global cause of multidrug resistant salmonellosis. We used Bayesian phylodynamics, genomic epidemiology, and phenotypic characterization to describe the emergence and evolution of Salmonella 4,[5],12:i:- in Australia. We show that the interruption of the genetic region surrounding the phase II flagellin, FljB, causing a monophasic phenotype, represents a stepwise evolutionary event through the accumulation of mobile resistance elements with minimal impairment to bacterial fitness. We identify three lineages with different population dynamics and discrete antimicrobial resistance profiles emerged, likely reflecting differential antimicrobial selection pressures. Two lineages are associated with travel to South-East Asia and the third lineage is endemic to Australia. Moreover antimicrobial-resistant Salmonella 4,[5],12:i- lineages efficiently infected and survived in host phagocytes and epithelial cells without eliciting significant cellular cytotoxicity, suggesting a suppression of host immune response that may facilitate the persistence of Salmonella 4,[5],12:i:-.


Assuntos
Farmacorresistência Bacteriana Múltipla/genética , Evolução Molecular , Salmonella enterica/classificação , Salmonella enterica/genética , Sorogrupo , Antibacterianos/farmacologia , Austrália , Teorema de Bayes , Linhagem Celular , Farmacorresistência Bacteriana Múltipla/efeitos dos fármacos , Flagelina/genética , Humanos , Imunidade , Metais Pesados/farmacologia , Filogenia , Salmonella enterica/efeitos dos fármacos , Salmonella typhimurium , Células THP-1 , Sequenciamento Completo do Genoma
7.
Lancet Public Health ; 6(8): e547-e556, 2021 08.
Artigo em Inglês | MEDLINE | ID: mdl-34252365

RESUMO

BACKGROUND: A cornerstone of Australia's ability to control COVID-19 has been effective border control with an extensive supervised quarantine programme. However, a rapid recrudescence of COVID-19 was observed in the state of Victoria in June, 2020. We aim to describe the genomic findings that located the source of this second wave and show the role of genomic epidemiology in the successful elimination of COVID-19 for a second time in Australia. METHODS: In this observational, genomic epidemiological study, we did genomic sequencing of all laboratory-confirmed cases of COVID-19 diagnosed in Victoria, Australia between Jan 25, 2020, and Jan 31, 2021. We did phylogenetic analyses, genomic cluster discovery, and integrated results with epidemiological data (detailed information on demographics, risk factors, and exposure) collected via interview by the Victorian Government Department of Health. Genomic transmission networks were used to group multiple genomic clusters when epidemiological and genomic data suggested they arose from a single importation event and diversified within Victoria. To identify transmission of emergent lineages between Victoria and other states or territories in Australia, all publicly available SARS-CoV-2 sequences uploaded before Feb 11, 2021, were obtained from the national sequence sharing programme AusTrakka, and epidemiological data were obtained from the submitting laboratories. We did phylodynamic analyses to estimate the growth rate, doubling time, and number of days from the first local infection to the collection of the first sequenced genome for the dominant local cluster, and compared our growth estimates to previously published estimates from a similar growth phase of lineage B.1.1.7 (also known as the Alpha variant) in the UK. FINDINGS: Between Jan 25, 2020, and Jan 31, 2021, there were 20 451 laboratory-confirmed cases of COVID-19 in Victoria, Australia, of which 15 431 were submitted for sequencing, and 11 711 met all quality control metrics and were included in our analysis. We identified 595 genomic clusters, with a median of five cases per cluster (IQR 2-11). Overall, samples from 11 503 (98·2%) of 11 711 cases clustered with another sample in Victoria, either within a genomic cluster or transmission network. Genomic analysis revealed that 10 426 cases, including 10 416 (98·4%) of 10 584 locally acquired cases, diagnosed during the second wave (between June and October, 2020) were derived from a single incursion from hotel quarantine, with the outbreak lineage (transmission network G, lineage D.2) rapidly detected in other Australian states and territories. Phylodynamic analyses indicated that the epidemic growth rate of the outbreak lineage in Victoria during the initial growth phase (samples collected between June 4 and July 9, 2020; 47·4 putative transmission events, per branch, per year [1/years; 95% credible interval 26·0-85·0]), was similar to that of other reported variants, such as B.1.1.7 in the UK (mean approximately 71·5 1/years). Strict interventions were implemented, and the outbreak lineage has not been detected in Australia since Oct 29, 2020. Subsequent cases represented independent international or interstate introductions, with limited local spread. INTERPRETATION: Our study highlights how rapid escalation of clonal outbreaks can occur from a single incursion. However, strict quarantine measures and decisive public health responses to emergent cases are effective, even with high epidemic growth rates. Real-time genomic surveillance can alter the way in which public health agencies view and respond to COVID-19 outbreaks. FUNDING: The Victorian Government, the National Health and Medical Research Council Australia, and the Medical Research Future Fund.


Assuntos
COVID-19/prevenção & controle , SARS-CoV-2/genética , COVID-19/epidemiologia , Estudos Epidemiológicos , Genômica , Humanos , SARS-CoV-2/isolamento & purificação , Vitória/epidemiologia
8.
Genome Med ; 13(1): 121, 2021 07 28.
Artigo em Inglês | MEDLINE | ID: mdl-34321076

RESUMO

BACKGROUND: Pathogen whole genome sequencing (WGS) is being incorporated into public health surveillance and disease control systems worldwide and has the potential to make significant contributions to infectious disease surveillance, outbreak investigation and infection prevention and control. However, to date, there are limited data regarding (i) the optimal models for integration of genomic data into epidemiological investigations and (ii) how to quantify and evaluate public health impacts resulting from genomic epidemiological investigations. METHODS: We developed the Pathogen Genomics in Public HeAlth Surveillance Evaluation (PG-PHASE) Framework to guide examination of the use of WGS in public health surveillance and disease control. We illustrate the use of this framework with three pathogens as case studies: Listeria monocytogenes, Mycobacterium tuberculosis and SARS-CoV-2. RESULTS: The framework utilises an adaptable whole-of-system approach towards understanding how interconnected elements in the public health application of pathogen genomics contribute to public health processes and outcomes. The three phases of the PG-PHASE Framework are designed to support understanding of WGS laboratory processes, analysis, reporting and data sharing, and how genomic data are utilised in public health practice across all stages, from the decision to send an isolate or sample for sequencing to the use of sequence data in public health surveillance, investigation and decision-making. Importantly, the phases can be used separately or in conjunction, depending on the need of the evaluator. Subsequent to conducting evaluation underpinned by the framework, avenues may be developed for strategic investment or interventions to improve utilisation of whole genome sequencing. CONCLUSIONS: Comprehensive evaluation is critical to support health departments, public health laboratories and other stakeholders to successfully incorporate microbial genomics into public health practice. The PG-PHASE Framework aims to assist public health laboratories, health departments and authorities who are either considering transitioning to whole genome sequencing or intending to assess the integration of WGS in public health practice, including the capacity to detect and respond to outbreaks and associated costs, challenges and facilitators in the utilisation of microbial genomics and public health impacts.


Assuntos
Ciência da Implementação , Infecções/diagnóstico , Listeria monocytogenes/isolamento & purificação , Mycobacterium tuberculosis/isolamento & purificação , SARS-CoV-2/isolamento & purificação , Sequenciamento Completo do Genoma/métodos , Genoma Bacteriano , Genoma Viral , Humanos , Infecções/epidemiologia , Listeria monocytogenes/genética , Mycobacterium tuberculosis/genética , Vigilância da População , Saúde Pública , SARS-CoV-2/genética
10.
Infect Control Hosp Epidemiol ; 42(5): 573-581, 2021 05.
Artigo em Inglês | MEDLINE | ID: mdl-34008484

RESUMO

OBJECTIVES: To conduct a pilot study implementing combined genomic and epidemiologic surveillance for hospital-acquired multidrug-resistant organisms (MDROs) to predict transmission between patients and to estimate the local burden of MDRO transmission. DESIGN: Pilot prospective multicenter surveillance study. SETTING: The study was conducted in 8 university hospitals (2,800 beds total) in Melbourne, Australia (population 4.8 million), including 4 acute-care, 1 specialist cancer care, and 3 subacute-care hospitals. METHODS: All clinical and screening isolates from hospital inpatients (April 24 to June 18, 2017) were collected for 6 MDROs: vanA VRE, MRSA, ESBL Escherichia coli (ESBL-Ec) and Klebsiella pneumoniae (ESBL-Kp), and carbapenem-resistant Pseudomonas aeruginosa (CRPa) and Acinetobacter baumannii (CRAb). Isolates were analyzed and reported as routine by hospital laboratories, underwent whole-genome sequencing at the central laboratory, and were analyzed using open-source bioinformatic tools. MDRO burden and transmission were assessed using combined genomic and epidemiologic data. RESULTS: In total, 408 isolates were collected from 358 patients; 47.5% were screening isolates. ESBL-Ec was most common (52.5%), then MRSA (21.6%), vanA VRE (15.7%), and ESBL-Kp (7.6%). Most MDROs (88.3%) were isolated from patients with recent healthcare exposure.Combining genomics and epidemiology identified that at least 27.1% of MDROs were likely acquired in a hospital; most of these transmission events would not have been detected without genomics. The highest proportion of transmission occurred with vanA VRE (88.4% of patients). CONCLUSIONS: Genomic and epidemiologic data from multiple institutions can feasibly be combined prospectively, providing substantial insights into the burden and distribution of MDROs, including in-hospital transmission. This analysis enables infection control teams to target interventions more effectively.


Assuntos
Farmacorresistência Bacteriana Múltipla , Genômica , Farmacorresistência Bacteriana Múltipla/genética , Monitoramento Epidemiológico , Hospitais , Humanos , Projetos Piloto , Estudos Prospectivos
11.
Nat Commun ; 12(1): 2491, 2021 05 03.
Artigo em Inglês | MEDLINE | ID: mdl-33941780

RESUMO

Mycobacterium kansasii can cause serious pulmonary disease. It belongs to a group of closely-related species of non-tuberculous mycobacteria known as the M. kansasii complex (MKC). Here, we report a population genomics analysis of 358 MKC isolates from worldwide water and clinical sources. We find that recombination, likely mediated by distributive conjugative transfer, has contributed to speciation and on-going diversification of the MKC. Our analyses support municipal water as a main source of MKC infections. Furthermore, nearly 80% of the MKC infections are due to closely-related M. kansasii strains, forming a main cluster that apparently originated in the 1900s and subsequently expanded globally. Bioinformatic analyses indicate that several genes involved in metabolism (e.g., maintenance of the methylcitrate cycle), ESX-I secretion, metal ion homeostasis and cell surface remodelling may have contributed to M. kansasii's success and its ongoing adaptation to the human host.


Assuntos
Água Potável/microbiologia , Genoma Bacteriano/genética , Pneumopatias/epidemiologia , Infecções por Mycobacterium não Tuberculosas/epidemiologia , Mycobacterium kansasii/genética , Metabolismo Energético/genética , Variação Genética/genética , Genética Populacional/métodos , Genômica , Humanos , Pneumopatias/microbiologia , Infecções por Mycobacterium não Tuberculosas/microbiologia , Mycobacterium kansasii/isolamento & purificação , Virulência/genética , Microbiologia da Água
12.
Trends Microbiol ; 29(9): 788-797, 2021 09.
Artigo em Inglês | MEDLINE | ID: mdl-33736902

RESUMO

Phylodynamic methods have been essential to understand the interplay between the evolution and epidemiology of infectious diseases. To date, the field has centered on viruses. Bacterial pathogens are seldom analyzed under such phylodynamic frameworks, due to their complex genome evolution and, until recently, a paucity of whole-genome sequence data sets with rich associated metadata. We posit that the increasing availability of bacterial genomes and epidemiological data means that the field is now ripe to lay the foundations for applying phylodynamics to bacterial pathogens. The development of new methods that integrate more complex genomic and ecological data will help to inform public heath surveillance and control strategies for bacterial pathogens that represent serious threats to human health.

13.
Genome Med ; 13(1): 52, 2021 03 30.
Artigo em Inglês | MEDLINE | ID: mdl-33785076

RESUMO

BACKGROUND: The hospital-adapted A1 group of Enterococcus faecium remains an organism of significant concern in the context of drug-resistant hospital-associated infections. How this pathogen evolves and disseminates remains poorly understood. METHODS: A large, globally representative collection of short-read genomic data from the hospital-associated A1 group of Enterococcus faecium was assembled (n = 973). We analysed, using a novel analysis approach, global diversity in terms of both the dynamics of the accessory genome and homologous recombination among conserved genes. RESULTS: Two main modes of genomic evolution continue to shape E. faecium: the acquisition and loss of genes, including antimicrobial resistance genes, through mobile genetic elements including plasmids, and homologous recombination of the core genome. These events lead to new clones emerging at the local level, followed by the erosion of signals of clonality through recombination, and in some identifiable cases producing new clonal clusters. These patterns lead to new, emerging lineages which are able to spread globally over relatively short timeframes. CONCLUSIONS: The ability of A1 E. faecium to continually present new combinations of genes for potential selection suggests that controlling this pathogen will remain challenging but establishing a framework for understanding genomic evolution is likely to aid in tracking the threats posed by newly emerging lineages.

14.
J Clin Microbiol ; 59(5)2021 04 20.
Artigo em Inglês | MEDLINE | ID: mdl-33658263

RESUMO

Resistance-guided therapy (RGT) for gonorrhea may reduce unnecessary use of broad-spectrum antibiotics. When reflexed from the Aptima Combo 2 assay, the ResistancePlus GC assay demonstrated 94.8% sensitivity and 100.0% specificity for Neisseria gonorrhoeae detection. Of the 379 concordant N. gonorrhoeae-positive samples, 86.8% were found to possess the gyrA S91F mutation, which was highly predictive for ciprofloxacin resistance and stable across 3,144 publicly available N. gonorrhoeae genomes. Our work supports the feasibility of implementing RGT for gonorrhea into routine molecular workflows.


Assuntos
Gonorreia , Neisseria gonorrhoeae , Antibacterianos/farmacologia , Antibacterianos/uso terapêutico , Ciprofloxacina/farmacologia , Farmacorresistência Bacteriana , Gonorreia/diagnóstico , Gonorreia/tratamento farmacológico , Humanos , Testes de Sensibilidade Microbiana , Neisseria gonorrhoeae/genética , Reflexo
15.
Lancet Infect Dis ; 21(5): 647-656, 2021 05.
Artigo em Inglês | MEDLINE | ID: mdl-33676595

RESUMO

BACKGROUND: To address the increasing incidence of gonorrhoea and antimicrobial resistance, we compared the efficacy of Listerine and Biotène mouthwashes for preventing gonorrhoea among men who have sex with men (MSM). METHODS: The OMEGA trial was a multicentre, parallel-group, double-blind randomised controlled trial among MSM, done at three urban sexual health clinics and one general practice clinic in Australia. Men were eligible if they were diagnosed with oropharyngeal gonorrhoea by nucleic acid amplification test (NAAT) in the previous 30 days or were aged 16-24 years. They were randomly assigned to receive Listerine (intervention) or Biotène (control) via a computer-generated sequence (1:1 ratio, block size of four). Participants, clinicians, data collectors, data analysts, and outcome adjudicators were masked to the interventions after assignment. Participants were instructed to rinse and gargle with 20 mL of mouthwash for 60 s at least once daily for 12 weeks. Oropharyngeal swabs were collected by research nurses every 6 weeks, and participants provided saliva samples every 3 weeks, to be tested for Neisseria gonorrhoeae with NAAT and quantitative PCR. The primary outcome was proportion of MSM diagnosed with oropharyngeal N gonorrhoeae infection at any point over the 12-week period, defined as a positive result for either oropharyngeal swabs or saliva samples by NAAT, and the cumulative incidence of oropharyngeal gonorrhoea at the week 12 visit. A modified intention-to-treat analysis for the primary outcome was done that included men who provided at least one follow-up specimen over the 12-week study period. The trial was registered on the Australian and New Zealand Clinical Trials Registry (ACTRN12616000247471). FINDINGS: Between March 30, 2016, and Oct 26, 2018, 786 MSM were screened and 256 were excluded. 264 MSM were randomly assigned to the Biotène group and 266 to the Listerine group. The analysis population included 227 (86%) men in the Biotène group and 219 (82%) in the Listerine group. Oropharyngeal gonorrhoea was detected in ten (4%) of 227 of MSM in the Biotène group and in 15 (7%) of 219 in the Listerine group (adjusted risk difference 2·5%, 95% CI -1·8 to 6·8). The cumulative incidence of oropharyngeal gonorrhoea at the week 12 visit did not differ between the two mouthwash groups (adjusted risk difference 3·1%, 95% CI -1·4 to 7·7). INTERPRETATION: Listerine did not reduce the incidence of oropharyngeal gonorrhoea compared with Biotène. However, previous research suggests that mouthwash might reduce the infectivity of oropharyngeal gonorrhoea; therefore, further studies of mouthwash examining its inhibitory effect on N gonorrhoeae are warranted to determine if it has a potential role for the prevention of transmission. FUNDING: Australian National Health and Medical Research Council.


Assuntos
Anti-Infecciosos Locais/uso terapêutico , Gonorreia/prevenção & controle , Antissépticos Bucais/uso terapêutico , Adulto , Austrália , Método Duplo-Cego , Combinação de Medicamentos , Glucose Oxidase , Homossexualidade Masculina , Humanos , Lactoperoxidase , Masculino , Estudos Multicêntricos como Assunto , Muramidase , Neisseria gonorrhoeae/efeitos dos fármacos , Nova Zelândia , Infecções Respiratórias/prevenção & controle , Salicilatos/uso terapêutico , Minorias Sexuais e de Gênero , Inquéritos e Questionários , Terpenos/uso terapêutico , Adulto Jovem
16.
Artigo em Inglês | MEDLINE | ID: mdl-33593834

RESUMO

Topical antibiotic preparations, such as fusidic acid (FA) or mupirocin, are used in the prevention and treatment of superficial skin infections caused by staphylococci. Previous genomic epidemiology work has suggested an association between the widespread use of topical antibiotics and the emergence of methicillin resistant Staphylococcus aureus in some settings. In this study, we provide experimental proof of co-selection for multidrug resistance in S. aureus following exposure to FA or mupirocin. Through targeted mutagenesis and phenotypic analyses, we confirmed that fusC carriage confers resistance to FA, and mupA carriage confers high-level resistance to mupirocin in multiple S. aureus genetic backgrounds. In vitro experiments demonstrated that carriage of fusC and mupA confer a competitive advantage in the presence of sub-inhibitory concentrations of FA and mupirocin, respectively. Further, we used a porcine skin colonisation model to show that clinically relevant concentrations of topical antibiotics can co-select for presence of unrelated antimicrobial resistance determinants, such as mecA, blaZ, and qacA, in fusC or mupA harbouring S. aureus These findings provide valuable insights on the role of acquired FA or mupirocin resistance in co-selecting for broader antibiotic resistance in S. aureus, prompting greater need for judicious use of topical antibiotics.

17.
Int J Food Microbiol ; 340: 109042, 2021 Feb 16.
Artigo em Inglês | MEDLINE | ID: mdl-33461002

RESUMO

The development of antimicrobial resistance in foodborne pathogens is a growing public health concern. This study was undertaken to determine the antimicrobial susceptibility of Salmonella enterica subspecies enterica isolated from the Australian commercial egg layer industry. S. enterica subspecies enterica (n=307) isolated from Australian commercial layer flock environments (2015-2018) were obtained from reference, research and State Government laboratories from six Australian states. All Salmonella isolates were serotyped. Antimicrobial susceptibility testing (AST) for 16 antimicrobial agents was performed by broth microdilution. Antimicrobial resistance genes and sequence types (STs) were identified in significant isolates by whole genome sequencing (WGS). Three main serotypes were detected, S. Typhimurium (n=61, 19.9%), S. Senftenburg (n=45, 14.7%) and S. Agona (n=37, 12.1%). AST showed 293/307 (95.4%) isolates were susceptible to all tested antimicrobial agents and all isolates were susceptible to amoxicillin-clavulanate, azithromycin, ceftiofur, ceftriaxone, ciprofloxacin, colistin, florfenicol, gentamicin, kanamycin and trimethoprim-sulfamethoxazole. Low levels of non-susceptibility were observed to streptomycin (2.3%, n=7), sulfisoxazole (2.0%, n=6), chloramphenicol (1.3%, n=4) and tetracycline (1.0%, n=3). Very low levels of non-susceptibility were observed to ampicillin (2/307; 0.7%) and cefoxitin (2/307; 0.7%). Two isolates (S. Havana and S. Montevideo), exhibited multidrug-resistant phenotypes to streptomycin, sulfisoxazole and tetracycline and possessed corresponding antimicrobial resistance genes (aadA4, aac(6')-Iaa, sul1, tetB). One S. Typhimurium isolate was resistant to ampicillin and tetracycline, and possessed both tetA and blaTEM-1B. WGS also identified these isolates as belonging to ST4 (S. Montevideo), ST578 (S. Havana) and ST19 (S. Typhimurium). The absence of resistance to highest priority critically important antimicrobials as well as the extremely low level of AMR generally among Australian commercial egg layer Salmonella isolates likely reflect Australia's conservative antimicrobial registration policy in food-producing animals and low rates of antimicrobial use within the industry.


Assuntos
Antibacterianos/farmacologia , Galinhas/microbiologia , Farmacorresistência Bacteriana , Abrigo para Animais , Aves Domésticas/microbiologia , Salmonella enterica/efeitos dos fármacos , Animais , Austrália , Ciprofloxacina/farmacologia , Farmacorresistência Bacteriana/genética , Farmacorresistência Bacteriana Múltipla/genética , Genoma Bacteriano , Testes de Sensibilidade Microbiana , Salmonella enterica/isolamento & purificação , Sorogrupo , Sequenciamento Completo do Genoma
18.
Hepatology ; 73(5): 1652-1670, 2021 May.
Artigo em Inglês | MEDLINE | ID: mdl-32780526

RESUMO

BACKGROUND AND AIMS: We conducted haplotype analysis of complete hepatitis B virus (HBV) genomes following deep sequencing from 368 patients across multiple phases of chronic hepatitis B (CHB) infection from four major genotypes (A-D), analyzing 4,110 haplotypes to identify viral variants associated with treatment outcome and disease progression. APPROACH AND RESULTS: Between 18.2% and 41.8% of nucleotides and between 5.9% and 34.3% of amino acids were 100% conserved in all genotypes and phases examined, depending on the region analyzed. Hepatitis B e antigen (HBeAg) loss by week 192 was associated with different haplotype populations at baseline. Haplotype populations differed across the HBV genome and CHB history, this being most pronounced in the precore/core gene. Mean number of haplotypes (frequency) per patient was higher in immune-active, HBeAg-positive chronic hepatitis phase 2 (11.8) and HBeAg-negative chronic hepatitis phase 4 (16.2) compared to subjects in the "immune-tolerant," HBeAg-positive chronic infection phase 1 (4.3, P< 0.0001). Haplotype frequency was lowest in genotype B (6.2, P< 0.0001) compared to the other genotypes (A = 11.8, C = 11.8, D = 13.6). Haplotype genetic diversity increased over the course of CHB history, being lowest in phase 1, increasing in phase 2, and highest in phase 4 in all genotypes except genotype C. HBeAg loss by week 192 of tenofovir therapy was associated with different haplotype populations at baseline. CONCLUSIONS: Despite a degree of HBV haplotype diversity and heterogeneity across the phases of CHB natural history, highly conserved sequences in key genes and regulatory regions were identified in multiple HBV genotypes that should be further investigated as targets for antiviral therapies and predictors of treatment response.

19.
Intern Med J ; 51(3): 390-397, 2021 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-32043702

RESUMO

BACKGROUND: Historically, Australian cases of invasive meningococcal disease (IMD) have been most frequently caused by Neisseria meningitidis serogroup B, but recently an increase in cases due to serogroup W (MenW) and serogroup Y (MenY) has occurred. AIM: To determine whether clinical manifestations of IMD have changed due to increased incidence of MenW and MenY. METHODS: We performed a retrospective review of IMD cases notified to the Department of Health and Human Services in Victoria, Australia. We compared the period between January 2013 and June 2015 (defined as P1) immediately before the increase in MenW and MenY was noted, with the equal time period of July 2015 to December 2017 (P2), when this increase was observed. RESULTS: IMD was notified more frequently in P2 than P1 (1.24 vs 0.53 per 100 000 person-years, P < 0.001). IMD cases in P2 were older (46 vs 19 years, P < 0.001), and more likely due to MenW (92/187, 49.2% vs 11/80, 13.8%, P < 0.001) or MenY (31/187, 16.6% vs 4/80, 5.0%, P = 0.01). IMD cases from P2 were more likely bacteraemic (151/187, 80.7% vs 55/80, 68.8%, P = 0.04), while meningitis (68/187, 36.4% vs 41/80, 51.3%, P = 0.03) and rash (65/181, 35.9% vs 45/78, 57.7%, P = 0.002) were less frequent. Intensive care unit admission rates and in-hospital mortality were unchanged. CONCLUSION: Alongside an increase in IMD in Victoria, the proliferation of cases of MenW and MenY occurred in older patients, and were more often identified through bacteraemia rather than meningitis or purpura fulminans. Clinicians should be aware of these changes to facilitate earlier identification and treatment of IMD.


Assuntos
Infecções Meningocócicas , Neisseria meningitidis , Idoso , Humanos , Incidência , Infecções Meningocócicas/diagnóstico , Infecções Meningocócicas/epidemiologia , Neisseria meningitidis Sorogrupo Y , Estudos Retrospectivos , Sorogrupo , Vitória/epidemiologia
20.
J Med Microbiol ; 70(2)2021 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-33270005

RESUMO

Saliva has recently been proposed as a suitable specimen for the diagnosis of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Use of saliva as a diagnostic specimen may present opportunities for SARS-CoV-2 reverse transcription polymerase chain reaction (RT-PCR) testing in remote and low-resource settings. Determining the stability of SARS-CoV-2 RNA in saliva over time is an important step in determining optimal storage and transport times. We undertook an in vitro study to assess whether SARS-CoV-2 could be detected in contrived saliva samples. The contrived saliva samples comprised 10 ml pooled saliva spiked with gamma-irradiated SARS-CoV-2 to achieve a concentration of 2.58×104 copies ml SARS-CoV-2, which was subsequently divided into 2 ml aliquots comprising: (i) neat saliva; and a 1 : 1 dilution with (ii) normal saline; (iii) viral transport media, and (iv) liquid Amies medium. Contrived samples were made in quadruplicate, with two samples of each stored at either: (i) room temperature or (ii) 4 °C. SARS-CoV-2 was detected in all SARS-CoV-2 spiked samples at time point 0, day 1, 3 and 7 at both storage temperatures using the N gene RT-PCR assay and time point 0, day 1 and day 7 using the Xpert Xpress SARS-CoV-2 (Cepheid, Sunnyvale, USA) RT-PCR assay. The ability to detect SARS-CoV-2 in saliva over a 1 week period is an important finding that presents further opportunities for saliva testing as a diagnostic specimen for the diagnosis of SARS-CoV-2.


Assuntos
Teste para COVID-19/métodos , COVID-19/diagnóstico , RNA Viral/análise , SARS-CoV-2/genética , Saliva/virologia , Humanos , Técnicas de Diagnóstico Molecular , Reação em Cadeia da Polimerase Via Transcriptase Reversa , SARS-CoV-2/isolamento & purificação , Manejo de Espécimes
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