Toni Chapman

Comparative analysis of virulence genes, genetic diversity, and phylogeny of commensal and enterotoxigenic Escherichia coli isolates from weaned pigs

Publisher: American Society for Microbiology

Date: 2007

DOI: 10.1128/AEM.00990-06

Abstract: If the acquisition of virulence genes (VGs) for pathogenicity were not solely acquired through horizontal gene transfers of pathogenicity islands, transposons, and phages, then clonal clusters of enterotoxigenic Escherichia coli (ETEC) would contain few or even none of the VGs found in strains responsible for extraintestinal infections. To evaluate this possibility, 47 postweaning diarrhea (PWD) ETEC strains from different geographical origins and 158 commensal E. coli isolates from the gastrointestinal tracts of eight group-housed healthy pigs were screened for 36 extraintestinal and 18 enteric VGs using multiplex PCR assays. Of 36 extraintestinal VGs, only 8 were detected ( fimH , traT , fyuA , hlyA , kpsMtII , k5 , iha , and ompT ) in the ETEC collection. Among these, hlyA (α-hemolysin) and iha (nonhemagglutinating adhesin) occurred significantly more frequently among the ETEC isolates than in the commensal isolates. Clustering analysis based on the VG profiles separated commensal and ETEC isolates and even differentiated serogroup O141 from O149. On the other hand, pulsed-field gel electrophoresis (PFGE) successfully clustered ETEC isolates according to both serotype and geographical origin. In contrast, the commensal isolates were heterogeneous with respect to both serotype and DNA fingerprint. This study has validated the use of VG profiling to examine pathogenic relationships between porcine ETEC isolates. The clonal relationships of these isolates can be further clarified by PFGE fingerprinting. The presence of extraintestinal VGs in porcine ETEC confirmed the hypothesis that in idual virulence gene acquisitions can occur concurrently against a background of horizontal gene transfers of pathogenicity islands. Over time, this could enable specific clonotypes to respond to host selection pressure and to evolve into new strains with increased virulence.

Multilocus sequence typing of Australian Streptococcus suis type 2 by MALDI-TOF mass spectrometry analysis of PCR amplicons

Publisher: Elsevier BV

Date: 06-2015

DOI: 10.1016/J.VETMIC.2015.03.006

Abstract: Streptococcus suis serotype 2 is a ubiquitous pathogen of swine and is known to cause severe disease in humans. Multilocus sequence typing (MLST) is ideal for characterising this organism because it permits isolates to be compared on a national and international scale. A novel approach to MLST using matrix-assisted laser desorption ionisation-time of flight mass spectrometry (MS-MLST) provides a more rapid alternative to dideoxy sequencing. This study used MS-MLST to define the multilocus sequence types (STs) present among a collection of Australian S. suis type 2, and thus, delivered a basis for comparison of Australian isolates with international strains already well characterised for virulence attributes. A collection of 45 isolates recovered from infected humans (n=3) and diseased pigs (n=42) was genotyped using MS-MLST and conventional MLST. Both methods were 100% concordant in their classification of sequence types (STs), although MS-MLST permitted much quicker analysis of sequence data. The collection contained ST25 (n=31), ST1 (n=10), ST28 (n=3) and ST369 (n=1). These results are consistent with the population structure of S. suis type 2 observed in diseased pigs and humans in Canada and the United Kingdom. MS-MLST may have utility for studying the population structure and epidemiology of S. suis in countries where the ersity of S. suis is greater and human disease is more common.

Molecular serogrouping of porcine enterotoxigenic Escherichia coli from Australia

Publisher: Elsevier BV

Date: 2012

DOI: 10.1016/J.MIMET.2011.10.016

Abstract: Enterotoxigenic Escherichia coli (ETEC) is a common etiological agent of neonatal, pre and post weaning diarrhoea in piglets. One of the most important steps in the diagnosis and epidemiological understanding of this organism is accurate serogrouping. In many instances, however, conventional serogrouping fails to produce accurate identification of serogroups. In this communication we report a modified and simplified molecular serogrouping method (rfb-RFLP) for the accurate identification of the most common porcine ETEC strains that cause neonatal, pre and post weaning diarrhoea in Australia.

Population structure of Bactrocera dorsalis s.s., B. papayae and B. philippinensis (diptera: Tephritidae) in southeast asia: Evidence for a single species hypothesis using mitochondrial DNA and wing-shape data

Publisher: Springer Science and Business Media LLC

Date: 2012

DOI: 10.1186/1471-2148-12-130

Molecular Characterization of Commensal Escherichia coli Adapted to Different Compartments of the Porcine Gastrointestinal Tract

Publisher: American Society for Microbiology

Date: 10-2012

DOI: 10.1128/AEM.01688-12

Abstract: The role of Escherichia coli as a pathogen has been the focus of considerable study, while much less is known about it as a commensal and how it adapts to and colonizes different environmental niches within the mammalian gut. In this study, we characterize Escherichia coli organisms ( n = 146) isolated from different regions of the intestinal tracts of eight pigs (dueodenum, ileum, colon, and feces). The isolates were typed using the method of random lified polymorphic DNA (RAPD) and screened for the presence of bacteriocin genes and plasmid replicon types. Molecular analysis of variance using the RAPD data showed that E. coli isolates are nonrandomly distributed among different gut regions, and that gut region accounted for 25% ( P 0.001) of the observed variation among strains. Bacteriocin screening revealed that a bacteriocin gene was detected in 45% of the isolates, with 43% carrying colicin genes and 3% carrying microcin genes. Of the bacteriocins observed (H47, E3, E1, E2, E7, Ia/Ib, and B/M), the frequency with which they were detected varied with respect to gut region for the colicins E2, E7, Ia/Ib, and B/M. The plasmid replicon typing gave rise to 25 profiles from the 13 Inc types detected. Inc F types were detected most frequently, followed by Inc HI1 and N types. Of the Inc types detected, 7 were nonrandomly distributed among isolates from the different regions of the gut. The results of this study indicate that not only may the different regions of the gastrointestinal tract harbor different strains of E. coli but also that strains from different regions have different characteristics.

Safety of raw meat and shellfish in Vietnam: An analysis of Escherichia coli isolations for antibiotic resistance and virulence genes

Publisher: Elsevier BV

Date: 06-2008

DOI: 10.1016/J.IJFOODMICRO.2008.03.029

Abstract: This study was conducted to examine a current baseline profile of antimicrobial resistance and virulence of Escherichia coli isolated from foods commonly sold in the market place in Vietnam. E. coli were isolated from 180 s les of raw meat, poultry and shellfish and also isolated from 43 chicken faeces s les. Ninety-nine E. coli isolates recovered from all sources were selected for the investigation of their susceptibility to 15 antimicrobial agents by the disk diffusion method. Eighty-four percent of the isolates were resistant to one or more antibiotics, and multi-resistance, defined as resistance to at least 3 different classes of antibiotics, was detected in all sources. The rates of multi-resistance were up to 89.5% in chicken, 95% in chicken faeces and 75% in pork isolates. Resistance was most frequently observed to tetracycline (77.8%), sulfafurazole (60.6%), icillin (50.5%), amoxicillin (50.5%), trimethoprim (51.5%), chlor henicol (43.4%), streptomycin (39.4%), nalidixic acid (34.3%) and gentamicin (24.2%). In addition, the isolates also displayed resistance to fluoroquinolones (ciprofloxacin 16.2%, norfloxacin 17.2%, and enrofloxacin 21.2%), with chicken isolates showing the highest rates of resistance to these antibiotics (52.6-63.2%). Thirty-eight multi-resistant isolates were selected for further the examination of antibiotic resistance genes and were also evaluated for virulence gene profiles by multiplex and uniplex polymerase chain reaction. The beta-lactam TEM gene and tetracycline resistance tetA, tetB genes were frequently detected in the tested isolates (84.2% and 89.5% respectively). Genes which are responsible for resistance to streptomycin (aadA) (68.4%), chlor henicol (cmlA) (42.1%), sulfonamides (sulI) (39.5%), trimethoprim (dhfrV) (26.3%) and kanamycin (aphA-1) (23.7%) were also widely distributed. Plasmid-mediated C genes were detected in E. coli isolates from chicken and pork. The isolates were tested for the presence of 58 virulence genes for adhesins, toxins, capsule synthesis, siderophores, invasins and others from different E. coli pathotypes. All of the tested isolates contained at least one virulence gene and there were 16 genes detected. Virulence genes detected were fimH (92.1%), bmaE (84.2%), TSPE4.C2 (42.1%), aidA AIDA-I (orfB) (31.6%), east1 (26.3%), traT (23.7%), and others including fyuA, iutA, chuA, yjaA, iss, iroN(E. coli), ibeA, aah (orfA), iha and papG allele III (10.5-2.6%). Typical toxin genes produced by enterohemorrhagic and enterotoxigenic E. coli pathotypes (a heat-stable toxin (ST), heat-labile toxin (LT) and Shiga toxin stx1, stx2) were not detected in any of these 38 isolates. The study has revealed that E. coli in raw foods is a significant reservoir of resistance and virulence genes.

Staphylococcus aureus ST398 detected in pigs in Australia

Publisher: Oxford University Press (OUP)

Date: 19-01-2014

DOI: 10.1093/JAC/DKT526

Fluoroquinolone-resistant extraintestinal pathogenic Escherichia coli, including O25b-ST131, isolated from faeces of hospitalized dogs in an Australian veterinary referral centre

Publisher: Oxford University Press (OUP)

Date: 08-01-2013

DOI: 10.1093/JAC/DKS515

Abstract: To determine rates of carriage of fluoroquinolone-resistant Escherichia coli and extraintestinal pathogenic E. coli (ExPEC) among dogs in a specialist referral hospital and to examine the population structure of the isolates. Fluoroquinolone-resistant faecal E. coli isolates (n = 232, from 23 of 123 dogs) recovered from hospitalized dogs in a veterinary referral centre in Sydney, Australia, over 140 days in 2009 were characterized by phylogenetic grouping, virulence genotyping and random lified polymorphic DNA (RAPD) analysis. The RAPD dendrogram for representative isolates showed one group B2-associated cluster and three group D-associated clusters each contained isolates with closely related ExPEC-associated virulence profiles. All group B2 faecal isolates represented the O25b-ST131 clonal group and were closely related to recent canine extraintestinal ST131 clinical isolates from the east coast of Australia by RAPD analysis. Hospitalized dogs may carry fluoroquinolone-resistant ExPEC in their faeces, including those representing O25b-ST131.

Green Fluorescent Protein-Based Biosensor To Detect and Quantify Stress Responses Induced by DNA-Degrading Colicins

Publisher: American Society for Microbiology

Date: 15-09-2011

DOI: 10.1128/AEM.00534-11

Abstract: Here we report the development of a whole-cell biosensor to detect and quantify the induction of the SOS response activated by DNA-degrading colicins. This biosensor utilizes the SOS-responsive cda promoter to regulate the expression of green fluorescent protein. The biosensor assay revealed induction of stress for all DNA-degrading reference colicins (E2, E7, and E8).

Genomic Characterisation of a Multiple Drug Resistant IncHI2 ST4 Plasmid in Escherichia coli ST744 in Australia

Publisher: MDPI AG

Date: 14-06-2020

DOI: 10.3390/MICROORGANISMS8060896

Abstract: Antibiotic resistance genes (ARGs) including those from the blaCTX-M family and mcr-1 that encode resistance to extended spectrum β–lactams and colistin, respectively, have been linked with IncHI2 plasmids isolated from swine production facilities globally but not in IncHI2 plasmids from Australia. Here we describe the first complete sequence of a multiple drug resistance Australian IncHI2-ST4 plasmid, pTZ41_1P, from a commensal E. coli from a healthy piglet. pTZ41_1P carries genes conferring resistance to heavy-metals (copper, silver, tellurium and arsenic), β-lactams, aminoglycosides and sulphonamides. The ARGs reside within a complex resistance locus (CRL) that shows considerable sequence identity to a CRL in pSDE_SvHI2, an IncHI2:ST3 plasmid from an enterotoxigenic E. coli with serotype O157:H19 of porcine origin that caused substantial losses to swine production operations in Australia in 2007. pTZ41_1P is closely related to IncHI2 plasmids found in E. coli and Salmonella enterica from porcine, avian and human sources in Europe and China but it does not carry genes encoding resistance to clinically-important antibiotics. We identified regions of IncHI2 plasmids that contribute to the genetic plasticity of this group of plasmids and highlight how they may readily acquire new resistance gene cargo. Genomic surveillance should be improved to monitor IncHI2 plasmids.

Microbiome of the Queensland fruit fly through metamorphosis

Publisher: MDPI AG

Date: 26-05-2020

DOI: 10.3390/MICROORGANISMS8060795

Abstract: Bactrocera tryoni (Froggatt) (Queensland fruit fly, or “Qfly”) is a highly polyphagous tephritid fruit fly and a serious economic pest in Australia. Qfly biology is intimately linked to the bacteria and fungi of its microbiome. While there are numerous studies of the microbiome in larvae and adults, the transition of the microbiome through the pupal stage remains unknown. To address this knowledge gap, we used high-throughput Next-Generation Sequencing (NGS) to examine microbial communities at each developmental stage in the Qfly life cycle, targeting the bacterial 16S rRNA and fungal ITS regions. We found that microbial communities were similar at the larval and pupal stage and were also similar between adult males and females, yet there were marked differences between the larval and adult stages. Specific bacterial and fungal taxa are present in the larvae and adults (fed hydrolyzed yeast with sugar) which is likely related to differences in nutritional biology of these life stages. We observed a significant abundance of the Acetobacteraceae at the family level, both in the larval and pupal stages. Conversely, Enterobacteriaceae was highly abundant ( %) only in the adults. The majority of fungal taxa present in Qfly were yeasts or yeast-like fungi. In addition to elucidating changes in the microbiome through developmental stages, this study characterizes the Qfly microbiome present at the establishment of laboratory colonies as they enter the domestication process.

Virulence characteristics of translocating Escherichia coli and the interleukin-8 response to infection

Publisher: Elsevier BV

Date: 02-2011

DOI: 10.1016/J.MICPATH.2010.11.003

Abstract: Four efficiently translocating Escherichia coli (TEC) strains isolated from the blood of humans (HMLN-1), pigs (PC-1) and rats (KIC-1 and KIC-2) were tested for their ability to adhere and translocate across human gut epithelial Caco-2 and HT-29 cells, to elicit a proinflammatory response and for the presence of 47 pathogenic E. coli virulence genes. HMLN-1 and PC-1 were more efficient in adhesion and translocation than rat strains, had identical biochemical phenotype (BPT) and serotype (O77:H18) and phylogenetic group (D). KIC-2 adhered more than KIC-1, belonged to different BPT and serotype but the same phylogenetic group as KIC-1. TEC strains elicited significantly higher IL-8 response in both cell lines (P < 0.05) and monocytic THP-1 (P < 0.0001) cells than non-TEC strains. KIC-2 induced the highest IL-8 response which may be associated with its immunostimulatory flagellin. Apart from adhesin genes fimH and bmaE that were carried by all strains, HMLN-1 and PC-1 carried capsule synthesis gene kpsMT III and KIC-2 carried the EAST1 toxin gene. The lack of known virulence genes and the ability of TEC to efficiently adhere and translocate whilst causing proinflammatory response suggests that these strains may carry as yet unidentified genes that enable their translocating ability.

Using Genomics to Design a Pathovar-Specific Loop-Mediated Isothermal Amplification (LAMP) Assay, for the Improved Detection of Xanthomonas citri pv. citri

Publisher: MDPI AG

Date: 02-06-2022

DOI: 10.3390/MICROORGANISMS10061153

Abstract: The ability to swiftly respond to pathogen incursions relies heavily on fast and accurate diagnostics. Current published assays for citrus bacterial canker do not target Xanthomonas citri pv. citri, the causative agent, with high specificity when testing Australian s les. While the current diagnostics are useful in countries where canker is endemic, the detection of canker in Australia requires an emergency response. Close relatives to X. citri pv. citri found in Australia may generate false positives with the current recommended diagnostic assays. Therefore, we developed a more specific detection tool for citrus bacterial canker to provide greater diagnostic confidence for surveillance and eradication efforts. We used genomic comparisons of 161 Xanthomonad genomes and identified and confirmed genomic regions specific for X. citri pv. citri by performing local alignments of unique regions to reference genomes. We then developed loop-mediated isothermal lification primers and validated them against a panel of 190 isolates to confirm specificity. Our diagnostic assay showed 100% corroboration with the concurrently developed multiplex primers and represents an improved diagnostic method capable of effective citrus bacterial canker identification.

Porcine commensalEscherichia coli: A reservoir for class 1 integrons associated with IS26

Publisher: Cold Spring Harbor Laboratory

Date: 03-07-2017

DOI: 10.1101/158808

Abstract: Porcine faecal waste is a serious environmental pollutant. Carriage of antimicrobial resistance and virulence-associated genes (VAGs) and the zoonotic potential of commensal Escherichia coli from swine is largely unknown. Furthermore, little is known about the role of commensal E. coli as contributors to the mobilisation of antimicrobial resistance genes between food animals and the environment. Here, we report whole genome sequence analysis of 141 E. coli from the faeces of healthy pigs. Most strains belonged to phylogroups A and B1 and carried i) a class 1 integron ii) VAGs linked with extraintestinal infection in humans iii) antimicrobial resistance genes bla TEM , aphAl, cmlA, strAB, tet(A) A, dfrA12, dfrA5, sul1, sul2, sul3 iv) IS26 and v) heavy metal resistance genes ( merA, cusA, terA ). Carriage of the sulphonamide resistance gene sul3 was notable in this study. The 141 strains belonged to 42 multilocus sequence types, but clonal complex 10 featured prominently. Structurally erse class 1 integrons that were frequently associated with IS26 carried unique genetic features that were also identified in extraintestinal pathogenic E. coli (ExPEC) from humans. This study provides the first detailed genomic analysis and point of reference for commensal E. coli of porcine origin, facilitating tracking of specific lineages and the mobile resistance genes they carry. None to declare.

Selected culturable enteric bacterial populations are modified by diet acidification and the growth promotant Tylosin

Publisher: Oxford University Press (OUP)

Date: 08-2005

DOI: 10.1111/J.1472-765X.2005.01743.X

Post-weaning shifts in microbiome composition and metabolism revealed by over 25,000 pig gut metagenome assembled genomes

Publisher: Cold Spring Harbor Laboratory

Date: 17-08-2020

DOI: 10.1101/2020.08.17.253872

Abstract: Using a previously described metagenomics dataset of 27 billion reads, we reconstructed over 50,000 metagenome-assembled genomes (MAGs) of organisms resident in the porcine gut, 46.5% of which were classified as % complete with a % contamination rate, and 24.4% were nearly complete genomes. Here we describe the generation and analysis of those MAGs using time-series s les. The gut microbial communities of piglets appear to follow a highly structured developmental program in the weeks following weaning, and this development is robust to treatments including an intramuscular antibiotic treatment and two probiotic treatments. The high resolution we obtained allowed us to identify specific taxonomic “signatures” that characterize the microbiome development immediately after weaning. Additionally, we characterized the carbohydrate repertoire of the organisms resident in the porcine gut, identifying 294 carbohydrate active enzymes. We tracked the shifts in abundance of these enzymes across time, and identified the species and higher-level taxonomic groups carrying each of these enzymes in their MAGs, raising the possibility of modifying the piglet microbiome through the tailored provision of carbohydrate substrates.

Genetic relatedness and virulence gene profiles of Escherichia coli strains isolated from septicaemic and uroseptic patients

Publisher: Springer Science and Business Media LLC

Date: 08-09-2009

DOI: 10.1007/S10096-009-0809-2

Abstract: We investigated the relationship between clonality and virulence factors (VFs) of a collection of Escherichia coli strains isolated from septicaemic and uroseptic patients with respect to their origin of translocation. Forty septicaemic and 30 uroseptic strains of E. coli were tested for their phylogenetic groupings, genetic relatedness using randomly lified polymorphic DNA (RAPD), biochemical fingerprinting method (biochemical phenotypes [BPTs]), adherence to HT-29 cells and the presence of 56 E. coli VF genes. Strains belonging to phylogenetic groups B2 and D constituted 93% of all strains. Fifty-four (77%) strains belonged to two major BPT/RAPD clusters (A and B), with cluster A carrying significantly (P = 0.0099) more uroseptic strains. The degree of adhesion to HT-29 cells of uroseptic strains was significantly (P = 0.0012) greater than that of septicaemic strains. Of the 56 VF genes tested, pap genes was the only group that were found significantly (P < 0.0001) more often among uroseptic isolates. Phylogenetic group B2 contained a significantly higher number of strains carrying pap genes than those in group D. We conclude that uroseptic E. coli are clonally different from septicaemic strains, carry more pap genes and predominantly adhere more to the HT-29 cell model of the gut.

Comparative genomic analysis of a multiple antimicrobial resistant enterotoxigenic E. coli O157 lineage from Australian pigs

Publisher: Springer Science and Business Media LLC

Date: 10-03-2015

DOI: 10.1186/S12864-015-1382-Y

Molecular characterization of Escherichia coli strains that cause symptomatic and asymptomatic urinary tract infections

Publisher: American Society for Microbiology

Date: 03-2012

DOI: 10.1128/JCM.06671-11

Abstract: The differences between Escherichia coli strains associated with symptomatic and asymptomatic urinary tract infections (UTIs) remain to be properly determined. Here we examined the prevalence of plasmid types and bacteriocins, as well as genetic relatedness, in a defined collection of E. coli strains that cause UTIs. Comparative analysis identified a subgroup of strains with a high number of virulence genes (VGs) and microcins M/H47. We also identified associations between microcin genes, VGs, and specific plasmid types.

Phylogenetic diversity analysis of shotgun metagenomic reads describes gut microbiome development and treatment effects in the post-weaned pig

Publisher: Public Library of Science (PLoS)

Date: 24-06-2022

DOI: 10.1371/JOURNAL.PONE.0270372

Abstract: Intensive farming practices can increase exposure of animals to infectious agents against which antibiotics are used. Orally administered antibiotics are well known to cause dysbiosis. To counteract dysbiotic effects, numerous studies in the past two decades sought to understand whether probiotics are a valid tool to help re-establish a healthy gut microbial community after antibiotic treatment. Although dysbiotic effects of antibiotics are well investigated, little is known about the effects of intramuscular antibiotic treatment on the gut microbiome and a few studies attempted to study treatment effects using phylogenetic ersity analysis techniques. In this study we sought to determine the effects of two probiotic- and one intramuscularly administered antibiotic treatment on the developing gut microbiome of post-weaning piglets between their 3 rd and 9 th week of life. Shotgun metagenomic sequences from over 800 faecal time-series s les derived from 126 post-weaning piglets and 42 sows were analysed in a phylogenetic framework. Differences between in idual hosts such as breed, litter, and age, were found to be important contributors to variation in the community composition. Host age was the dominant factor in shaping the gut microbiota of piglets after weaning. The post-weaning pig gut microbiome appeared to follow a highly structured developmental program with characteristic post-weaning changes that can distinguish hosts that were born as little as two days apart in the second month of life. Treatment effects of the antibiotic and probiotic treatments were found but were subtle and included a higher representation of Mollicutes associated with intramuscular antibiotic treatment, and an increase of Lactobacillus associated with probiotic treatment. The discovery of correlations between experimental factors and microbial community composition is more commonly addressed with OTU-based methods and rarely analysed via phylogenetic ersity measures. The latter method, although less intuitive than the former, suffers less from library size normalization biases, and it proved to be instrumental in this study for the discovery of correlations between microbiome composition and host-, and treatment factors.

Phylogenetic and molecular insights into the evolution of multidrug-resistant porcine enterotoxigenic Escherichia coli in Australia

Publisher: Elsevier BV

Date: 08-2014

DOI: 10.1016/J.IJANTIMICAG.2014.04.011

Abstract: This study investigated the phylogeny and molecular epidemiology of Australian porcine enterotoxigenic Escherichia coli (ETEC) isolates (n=70) by performing multilocus sequence typing (MLST), random lified polymorphic DNA (RAPD) analysis, virulence gene analysis, plasmid, bacteriocin, integron and antimicrobial resistance gene typing, and antimicrobial susceptibility phenotyping. Isolates of the most commonly observed O serogroup (O149) were highly clonal with a lower frequency of antimicrobial resistance compared with the less common O141 serogroup isolates, which were more genetically erse and resistant to a greater array of antimicrobials. The O149 and O141 isolates belonged to sequence types (STs) ST100 and ST1260, respectively. A small number of new STs were identified for the least common serogroups, including O157 (ST4245), O138 (ST4244), O139 (ST4246) and O8 (ST4247). A high frequency of plasmid replicons was observed among all ETEC isolates. However, O149 isolates predominantly carried IncFIB, I1, HI1 and FIC, whereas O141 isolates carried a more varied array, including IncI1, FIB, FIC, HI1, I1, Y and, most significantly, A/C. O141 isolates also possessed a greater ersity of bacteriocins, with almost one-half of the isolates carrying colicin E3 (44.4% 12/27) and E7 (48.1% 13/27). This study shows that Australian porcine ETEC are distinct from isolates obtained in other parts of the world with respect to the MLST profile and the absence of resistance to critically important antimicrobials, including third-generation cephalosporins and fluoroquinolones.

Comparative genomic analysis of toxin-negative strains of Clostridium difficile from humans and animals with symptoms of gastrointestinal disease

Publisher: Springer Science and Business Media LLC

Date: 12-03-2016

DOI: 10.1186/S12866-016-0653-3

Probiotic Microorganisms

Publisher: Wiley

Date: 14-07-2008

DOI: 10.1002/9780470432624.CH1

The genome of Clostridium difficile 5.3

Publisher: Springer Science and Business Media LLC

Date: 2014

DOI: 10.1186/1757-4749-6-4

Colonisation dynamics and virulence of two clonal groups of multidrug-resistant Escherichia coli isolated from dogs

Publisher: Elsevier BV

Date: 2009

DOI: 10.1016/J.MICINF.2008.10.014

Abstract: The study established the virulence potential of multidrug-resistant Escherichia coli (MDREC) isolates from nosocomial infections in hospitalised dogs. The isolates were resistant to fluoroquinolones, belonged to two distinct clonal groups (CG1 and CG2) and contained a plasmid-mediated AmpC (CMY-7) beta-lactamase. CG1 isolates (n=14) possessed two of 36 assayed extraintestinal virulence genes (iutA and traT) and belonged to phylogenetic group A, whereas CG2 isolates (n=19) contained four such genes (iutA, ibeA, fimH and kpsMT K5) and belonged to group D. In a mouse gastrointestinal tract colonisation model, colonisation by index CG1 strain C1 was transient, in contrast to the index CG2 strain C2b, which persisted up to 40days post-inoculation. In a mouse subcutaneous challenge model, both strains were less virulent than archetypal group B2 extraintestinal pathogenic E. coli (ExPEC) strain CFT073 strain C1 caused no systemic signs and strain C2b was lethal to only one of six mice. In a mouse urinary tract infection model, strain C2b colonised the mouse bladder over 2 logs higher compared to strain C1. Whilst both groups of canine MDREC appear less virulent than a reference human ExPEC strain, CG2 strains have greater capacity for colonisation and virulence.

Building More Resilient Culture Collections: A Call for Increased Deposits of Plant-Associated Bacteria

Publisher: MDPI AG

Date: 30-03-2022

DOI: 10.3390/MICROORGANISMS10040741

Abstract: Biological collections preserve our past, while helping protect our future and increase future knowledge. Plant bacterial culture collections are our security for domestic and global biosecurity. This feature article will provide an introduction to the global position of plant bacterial collections. The role of collections in monitoring plant pathogenic bacteria will be explored through the presentation of five cases studies. These case studies demonstrate why culture collections were imperative for the outcome in each situation. We discuss what we believe should be the best practices to improve microbial preservation and accessioning rates, and why plant bacterial culture collections must increase deposits to be prepared for future emerging pathogens. This is not only the case for global culture collections, but on a much bigger scale, our future scientific successes, our biosecurity decisions and responses, and our knowledge are contingent upon preserving our valuable bacterial strains. It is hoped that once you read this article, you will see the need to deposit your strains in registered public collections and make a concerted effort to build better bacterial culture collections with us.

Post-weaning shifts in microbiome composition and metabolism revealed by over 25 000 pig gut metagenome-assembled genomes

Publisher: Microbiology Society

Date: 09-08-2021

DOI: 10.1099/MGEN.0.000501

Abstract: Using a previously described metagenomics dataset of 27 billion reads, we reconstructed over 50 000 metagenome-assembled genomes (MAGs) of organisms resident in the porcine gut, 46.5 % of which were classified as % complete with a % contamination rate, and 24.4 % were nearly complete genomes. Here, we describe the generation and analysis of those MAGs using time-series s les. The gut microbial communities of piglets appear to follow a highly structured developmental programme in the weeks following weaning, and this development is robust to treatments including an intramuscular antibiotic treatment and two probiotic treatments. The high resolution we obtained allowed us to identify specific taxonomic ‘signatures’ that characterize the gut microbial development immediately after weaning. Additionally, we characterized the carbohydrate repertoire of the organisms resident in the porcine gut. We tracked the abundance shifts of 294 carbohydrate active enzymes, and identified the species and higher-level taxonomic groups carrying each of these enzymes in their MAGs. This knowledge can contribute to the design of probiotics and prebiotic interventions as a means to modify the piglet gut microbiome.

Phylogenetic diversity analysis of shotgun metagenomic reads describes gut microbiome development and treatment effects in the post-weaned pig

Publisher: Cold Spring Harbor Laboratory

Date: 20-07-2020

DOI: 10.1101/2020.07.20.211326

Abstract: Intensive farming practices can increase exposure of animals to infectious agents against which antibiotics are used. Besides leading to antimicrobial resistance (AMR), orally administered antibiotics are well known to cause dysbiosis. To counteract dysbiotic effects, numerous studies in the past two decades sought to understand whether probiotics are a valid tool to help re-establish a healthy gut microbial community after antibiotic treatment. However, although dysbiotic effects of antibiotics are well investigated, little is known about the effects of intramuscular antibiotic treatment on the gut microbiome and a few studies attempted to study treatment effects using phylogenetic ersity analysis techniques. In this study we sought to determine the effects of two probiotic- and one intramuscularly administered antibiotic treatment on the developing gut microbiome of post-weaning piglets between their 3 rd and 9 th week of life. Shotgun metagenomic sequences from over 800 faecal time-series s les derived from 126 piglets and 42 sows were analysed in a phylogenetic framework to characterise the developing gut microbial community composition of post-weaning piglets. We assessed the effects of intramuscular antibiotic treatment and probiotic oral treatment on the ersity of these gut microbial communities using alpha and beta ersity measures. Differences between in idual hosts such as breed, litter, and age, were found to be important contributors to variation in the community composition. Host age was the dominant factor in shaping the gut microbiota of piglets after weaning. The post-weaning pig gut microbiome appeared to follow a highly structured developmental program with characteristic post-weaning changes that can distinguish hosts that were born as little as two days apart in the second month of life. Treatment effects of the antibiotic and probiotic treatments were found but were subtle and included a higher representation of Mollicutes associated with intramuscular antibiotic treatment, and an increase of Lactobacillus associated with probiotic treatment. The discovery of correlations between experimental factors and microbial community composition is more commonly addressed with OTU-based methods and rarely analysed via phylogenetic ersity measures. The latter method, although less intuitive than the former, suffers less from library size normalization biases, and it proved to be instrumental in this study for the discovery of correlations between microbiome composition and host-, and treatment factors.

Whole Genome Sequencing Analysis of Porcine Faecal Commensal Escherichia coli Carrying Class 1 Integrons from Sows and Their Offspring

Publisher: MDPI AG

Date: 04-06-2020

DOI: 10.3390/MICROORGANISMS8060843

Abstract: Intensive pig production systems often rely on the use of antimicrobials and heavy metal feed additives to maintain animal health and welfare. To gain insight into the carriage of antimicrobial resistance genes (ARGs) in the faecal flora of commercially reared healthy swine, we characterised the genome sequences of 117 porcine commensal E. coli that carried the class 1 integrase gene (intI1+). Isolates were sourced from 42 healthy sows and 126 of their offspring from a commercial breeding operation in Australia in 2017. intI1+ E. coli was detected in 28/42 (67%) sows and 90/126 (71%) piglets. Phylogroup A, particularly clonal complex 10, and phylogroup B1 featured prominently in the study collection. ST10, ST20, ST48 and ST361 were the dominant sequence types. Notably, 113/117 isolates (96%) carried three or more ARGs. Genes encoding resistance to β-lactams, aminoglycosides, trimethoprim, sulphonamides, tetracyclines and heavy metals were dominant. ARGs encoding resistance to last-line agents, such as carbapenems and third generation cephalosporins, were not detected. IS26, an insertion sequence noted for its ability to capture and mobilise ARGs, was present in 108/117 (92%) intI1+ isolates, and it played a role in determining class 1 integron structure. Our data shows that healthy Australian pig faeces are an important reservoir of multidrug resistant E. coli that carry genes encoding resistance to multiple first-generation antibiotics and virulence-associated genes.

Molecular techniques for the detection and differentiation of host and parasitoid species and the implications for fruit fly management

Publisher: MDPI AG

Date: 14-08-2012

DOI: 10.3390/INSECTS3030763

Comparative Genomics of Xanthomonas citri pv. citri A* Pathotype Reveals Three Distinct Clades with Varying Plasmid Distribution

Publisher: MDPI AG

Date: 08-12-2020

DOI: 10.3390/MICROORGANISMS8121947

Abstract: Citrus bacterial canker (CBC) is an important disease of citrus cultivars worldwide that causes blister-like lesions on host plants and leads to more severe symptoms such as plant defoliation and premature fruit drop. The causative agent, Xanthomonas citri pv. citri, exists as three pathotypes—A, A*, and Aw—which differ in their host range and elicited host response. To date, comparative analyses have been h ered by the lack of closed genomes for the A* pathotype. In this study, we sequenced and assembled six CBC isolates of pathotype A* using second- and third-generation sequencing technologies to produce complete, closed assemblies. Analysis of these genomes and reference A, A*, and Aw sequences revealed genetic groups within the A* pathotype. Investigation of accessory genomes revealed virulence factors, including type IV secretion systems and heavy metal resistance genes, differentiating the genetic groups. Genomic comparisons of closed genome assemblies also provided plasmid distribution information for the three genetic groups of A*. The genomes presented here complement existing closed genomes of A and Aw pathotypes that are publicly available and open opportunities to investigate the evolution of X. citri pv. citri and the virulence factors that contribute to this serious pathogen.

Dynamics of the Queensland Fruit Fly Microbiome through the Transition from Nature to an Established Laboratory Colony

Publisher: MDPI AG

Date: 26-01-2022

DOI: 10.3390/MICROORGANISMS10020291

Abstract: The transition from nature to laboratory or mass rearing can impose significant physiological and evolutionary impact on insects. The Queensland fruit fly (also known as ‘Qfly’), Bactrocera tryoni (Froggatt) (Diptera: Tephritidae), is a serious economic pest that presents major challenges for horticulture industries in Australia. The sterile insect technique (SIT) is being developed to manage outbreaks in regions that remain free of Qfly and to suppress populations in regions where this species is endemic. The biology of Qfly is intimately connected to its microbiome. Therefore, changes in the microbiome that occur through domestication have implications for SIT. There are numerous studies of the microbiome in Qfly larvae and adults, but there is little information on how the microbiome changes as Qfly laboratory colonies are established. In this study, high-throughput Illumina sequencing was used to assess the Qfly microbiome in colonies reared from wild larvae, collected from fruit, for five generations, on a gel-based larval diet. Beta ersity analysis showed that the bacterial communities from Generation 5 (G5) clustered separately from earlier generations. At the genus level, bacterial communities were significantly different between the generations and mostly altered at G5. However, communities were found similar at phyla to family taxonomic levels. We observed high abundance of Morganella and Burkholderia at the genus level in the larval and pupal stages respectively at G5, but these were not detected in earlier generations. Overall, our findings demonstrate that the domestication process strongly affects the Qfly microbiome and prompts questions about the functional relationship between the Qfly and its microbiome, as well as implications for the performance of insects that have been domesticated and mass-reared for SIT programs.

Comparison of Virulence Gene Profiles of Escherichia coli Strains Isolated from Healthy and Diarrheic Swine

Publisher: American Society for Microbiology

Date: 07-2006

DOI: 10.1128/AEM.02885-05

Abstract: A combination of uni- and multiplex PCR assays targeting 58 virulence genes (VGs) associated with Escherichia coli strains causing intestinal and extraintestinal disease in humans and other mammals was used to analyze the VG repertoire of 23 commensal E. coli isolates from healthy pigs and 52 clinical isolates associated with porcine neonatal diarrhea (ND) and postweaning diarrhea (PWD). The relationship between the presence and absence of VGs was interrogated using three statistical methods. According to the generalized linear model, 17 of 58 VGs were found to be significant ( P 0.05) in distinguishing between commensal and clinical isolates. Nine of the 17 genes represented by iha , hlyA , aidA , east1 , aah , fimH , iroN E. coli , traT , and saa have not been previously identified as important VGs in clinical porcine isolates in Australia. The remaining eight VGs code for fimbriae (F4, F5, F18, and F41) and toxins (STa, STb, LT, and Stx2), normally associated with porcine enterotoxigenic E. coli . Agglomerative hierarchical algorithm analysis grouped E. coli strains into subclusters based primarily on their serogroup. Multivariate analyses of clonal relationships based on the 17 VGs were collapsed into two-dimensional space by principal coordinate analysis. PWD clones were distributed in two quadrants, separated from ND and commensal clones, which tended to cluster within one quadrant. Clonal subclusters within quadrants were highly correlated with serogroups. These methods of analysis provide different perspectives in our attempts to understand how commensal and clinical porcine enterotoxigenic E. coli strains have evolved and are engaged in the dynamic process of losing or acquiring VGs within the pig population.

Human-associated fluoroquinolone-resistant Escherichia coli clonal lineages, including ST354, isolated from canine feces and extraintestinal infections in Australia

Publisher: Elsevier BV

Date: 04-2015

DOI: 10.1016/J.MICINF.2014.12.016

Abstract: Phylogenetic group D extraintestinal pathogenic Escherichia coli (ExPEC), including O15:K52:H1 and clonal group A, have spread globally and become fluoroquinolone-resistant. Here we investigated the role of canine feces as a reservoir of these (and other) human-associated ExPEC and their potential as canine pathogens. We characterized and compared fluoroquinolone-resistant E. coli isolates originally identified as phylogenetic group D from either the feces of hospitalized dogs (n = 67 14 dogs) or extraintestinal infections (n = 53 33 dogs). Isolates underwent phylogenetic grouping, random lified polymorphic DNA (RAPD) analysis, virulence genotyping, resistance genotyping, human-associated ExPEC O-typing, and multi-locus sequence typing. Five of seven human-associated sequence types (STs) exhibited ExPEC-associated O-types, and appeared in separate RAPD clusters. The largest subgroup (16 fecal, 26 clinical isolates) were ST354 (phylogroup F) isolates. ST420 (phylogroup B2) O1-ST38, O15:K52:H1-ST393, and O15:K1-ST130 (phylogroup D) and O7-ST457, and O1-ST648 (phylogroup F) were also identified. Three ST-specific RAPD sub-clusters (ST354, ST393, and ST457) contained closely related isolates from both fecal or clinical sources. Genes encoding CTX-M and AmpC β-lactamases were identified in isolates from five STs. Major human-associated fluoroquinolone-resistant ± extended-spectrum cephalosporin-resistant ExPEC of public health importance may be carried in dog feces and cause extraintestinal infections in some dogs.

A large-scale metagenomic survey dataset of the post-weaning piglet gut lumen

Publisher: Oxford University Press (OUP)

Date: 06-2021

DOI: 10.1093/GIGASCIENCE/GIAB039

Abstract: Early weaning and intensive farming practices predispose piglets to the development of infectious and often lethal diseases, against which antibiotics are used. Besides contributing to the build-up of antimicrobial resistance, antibiotics are known to modulate the gut microbial composition. As an alternative to antibiotic treatment, studies have previously investigated the potential of probiotics for the prevention of postweaning diarrhea. In order to describe the post-weaning gut microbiota, and to study the effects of two probiotics formulations and of intramuscular antibiotic treatment on the gut microbiota, we s led and processed over 800 faecal time-series s les from 126 piglets and 42 sows. Here we report on the largest shotgun metagenomic dataset of the pig gut lumen microbiome to date, consisting of & Tbp of shotgun metagenomic sequencing data. The animal trial, the workflow from s le collection to s le processing, and the preparation of libraries for sequencing, are described in detail. We provide a preliminary analysis of the dataset, centered on a taxonomic profiling of the s les, and a 16S-based beta ersity analysis of the mothers and the piglets in the first 5 weeks after weaning. This study was conducted to generate a publicly available databank of the faecal metagenome of weaner piglets aged between 3 and 9 weeks old, treated with different probiotic formulations and intramuscular antibiotic treatment. Besides investigating the effects of the probiotic and intramuscular antibiotic treatment, the dataset can be explored to assess a wide range of ecological questions with regards to antimicrobial resistance, host-associated microbial and phage communities, and their dynamics during the aging of the host.

Rapid identification of virulence genes in enterotoxigenic Escherichia coli isolates associated with diarrhoea in Queensland piggeries

Publisher: Wiley

Date: 05-2005

DOI: 10.1111/J.1751-0813.2005.TB12745.X

Abstract: To identify virulence genes in enterotoxigenic E coli (ETEC) isolates associated with diarrhoea in neonatal, 1 to 3 week-old and weaned pigs in southeast Queensland. Multiplex PCR and serotyping were applied to E coli isolates obtained over a 5-year period (1998-2002) from cases diagnosed at Toowoomba Veterinary Laboratory. A total of 126 isolates from 25 different Queensland piggeries were tested for haemolytic activity on 5% sheep blood agar and by multiplex PCR for the presence of five commonly recognised fimbrial (F4, F5, F6, F41 and F18) and three enterotoxin genes (STa, STb, LT). A subset of 62 representative isolates were serotyped by slide agglutination. For comparative purposes, multiplex PCR was also performed on the DNA of 31 ETEC isolates from 9 serotypes originating from piggeries in southern New South Wales. A total of 113 (89.7%) of the isolates from Queensland possessed ETEC virulence genes, including 14 of 15 isolates from neonatal pigs (93.3%), 18 of 23 isolates from 1 to 3 week old pigs (78.3%) and 81 of 88 isolates from weaned pigs (92.1%). F4:STa:STb:LT (serotype O149) was the most prevalent pathotype in neonatal and 1-3 week old pigs and F4:STa:STb:LT (serotype O149) and F18:STa:STb:LT (serotype O141) were most prevalent in weaned pigs. In comparison, isolates obtained from neonatal pigs from New South Wales belonged to a more erse range of pathotypes and serotypes. Multiplex PCR was a rapid and specific method for detecting the presence of ETEC virulence genes in porcine E coli isolates. For isolates obtained from cases of suspected colibacillosis in Queensland, growth of a heavy pure culture of haemolytic E coli was a sensitive prognostic indicator of the presence of ETEC virulence genes in the isolate. ETEC pathotypes and serotypes remained stable in Queensland piggeries over the five-year study period and appear to have changed little over the last three decades.

Salmonella enterica isolated from infections in Australian livestock remain susceptible to critical antimicrobials

Publisher: Elsevier BV

Date: 02-2014

DOI: 10.1016/J.IJANTIMICAG.2013.10.014

Abstract: Salmonella enterica is a zoonotic pathogen causing a variety of diseases in humans and animals. Many countries are reporting an increase in the prevalence of multidrug-resistant (MDR) S. enterica in food animals. The aim of this study was to determine whether S. enterica isolated from livestock in New South Wales, Australia, have similar resistance traits to those reported internationally. Salmonella enterica (n=165) from clinical infections in food animals between 2007 and 2011 were serotyped and tested for susceptibility to 18 antimicrobials. Also, 22 antimicrobial resistance genes (ARGs), 3 integrons and 18 plasmid replicon types were screened for using PCR. Most isolates (66.1%) remained susceptible to all antimicrobials 8.5% of the isolates were resistant to four or more antimicrobials. Antimicrobials with the highest prevalence of resistance were sulfafurazole (28.5%), icillin (17.0%), tetracycline (15.8%) and trimethoprim (8.5%). There was no resistance to fluoroquinolones or third-generation cephalosporins. The most common ARGs were blaTEM (15.2%), sul2 (10.3%), tetB (9.1%), tetA (5.5%), aphA1 (4.8%) and dhfrV (4.8%). Class 1 integrons (7.9%) and IncFIIA (69.7%) were the most commonly detected integron and plasmid replicon types, respectively. Class 1 integrons were positively associated with MDR phenotypes and ARG carriage (P≤0.001). Internationally prominent MDR serovars associated with severe disease in humans (e.g. AmpC-positive Salmonella Newport) were not detected. Overall, the comparatively favourable resistance status of S. enterica in Australian livestock represents minimal public health risk associated with MDR strains and supports a conservative approach to the registration of antimicrobial drug classes in food-producing animals.

New South Wales Department Of Primary Industries

Location: Australia

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Western Sydney University - Hawkesbury Campus

Location: Australia

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University Of Wollongong

Location: Australia

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Linkage Projects - Grant ID: LP180100593

Start Date: 2020

End Date: 09-2024

Amount: $496,349.00

Funder: Australian Research Council

Industrial Transformation Training Centres - Grant ID: IC150100026

Start Date: 06-2016

End Date: 12-2022

Amount: $3,732,019.00

Funder: Australian Research Council

Linkage Projects - Grant ID: LP150100912

Start Date: 03-2016

End Date: 04-2020

Amount: $520,683.00

Funder: Australian Research Council