ORCID Profile
0000-0001-7013-7326
Current Organisation
New South Wales Department of Primary Industries
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Veterinary epidemiology | Bioinformatics Software | Phylogeny and Comparative Analysis | Genomics | Animal production | Animal protection (incl. pests and pathogens) | Genetics | Microbial genetics |
Control of Pests, Diseases and Exotic Species at Regional or Larger Scales | Expanding Knowledge in the Biological Sciences | Food Safety
Publisher: MDPI AG
Date: 22-10-2021
DOI: 10.3390/PATHOGENS10111367
Abstract: Chlamydia pecorum is a common gastrointestinal inhabitant of livestock but infections can manifest in a broad array of clinical presentations and in a range of host species. While C. pecorum is a known cause of ovine abortion, clinical cases have only recently been described in detail. Here, the prevalence and sequence types (STs) of C. pecorum in ewes from a property experiencing high levels of perinatal mortality (PNM) in New South Wales (NSW), Australia, were investigated using serological and molecular methods. Ewes that were PNM+ were statistically more likely to test seropositive compared to PNM− ewes and displayed higher antibody titres however, an increase in chlamydial shedding from either the rectum, vagina or conjunctiva of PNM+ ewes was not observed. Multilocus sequence typing (MLST) indicated that C. pecorum ST23 was the major ST shed by ewes in the flock, was the only ST identified from the vaginal site, and was the same ST detected within aborted foetal tissues. Whole genome sequencing of C. pecorum isolated from one abortion case revealed that the C. pecorum plasmid (pCpec) contained a unique deletion in coding sequence 1 (CDS1) that was also present in C. pecorum ST23 shed from the ewes. A further unique deletion was noted in a polymorphic membrane protein gene (pmpG) of the C. pecorum chromosome, which warrants further investigation given the role of PmpG in host cell adherence and tissue tropism.This study describes novel infection parameters in a sheep flock experiencing C. pecorum-associated perinatal mortality, provides the first genomic data from an abortigenic C. pecorum strain, and raises questions about possible links between unique genetic features of this strain and C. pecorum abortion.
Publisher: Microbiology Society
Date: 2020
Publisher: Springer Science and Business Media LLC
Date: 25-02-2016
Publisher: Elsevier BV
Date: 06-2015
DOI: 10.1016/J.MEEGID.2015.03.017
Abstract: In Australia, outbreaks of clinical theileriosis caused by Theileria orientalis have been largely associated with the Ikeda genotype which can occur as a sole infection, or more commonly, as a mixture of genotypes. The most prevalent genotype, Chitose, frequently co-occurs with type Ikeda, however the role of this genotype in clinical disease has not been clearly established. Furthermore, the dynamics of in idual genotypes in field infection of cattle have not been examined. In this study we developed quantitative PCR (qPCR) and genotyping methods to examine the role of the Chitose genotype in clinical disease and to investigate the temporal dynamics of T. orientalis Ikeda, Chitose and Buffeli genotypes in naïve animals introduced to a T. orientalis-endemic area. Analysis of the major piroplasm surface protein (MPSP) genes of Chitose isolates revealed the presence of two distinct phylogenetic clusters, Chitose A and Chitose B. A genotyping assay aimed at determining Chitose A/B allele frequency revealed that the Chitose A phylogenetic cluster is strongly associated with clinical disease but nearly always co-occurs with the Ikeda genotype. qPCR revealed that the Chitose genotype (particularly Chitose A), undergoes temporal switching in conjunction with the Ikeda genotype and contributes substantially to the overall parasite burden. The benign Buffeli genotype can also undergo temporal switching but levels of this genotype appear to remain low relative to the Ikeda and Chitose types. Interplay between vector and host immunological factors is presumed to be critical to the population dynamics observed in this study. Genotypic switching likely contributes to the persistence of T. orientalis in the host.
Publisher: Springer Science and Business Media LLC
Date: 07-2018
Publisher: Elsevier BV
Date: 04-2018
Publisher: Springer Science and Business Media LLC
Date: 22-01-2016
Publisher: American Society for Microbiology
Date: 10-2020
DOI: 10.1128/MRA.00897-20
Abstract: The draft genome sequence of a novel “ Candidatus Liberibacter” species detected in an unidentified species of Zanthoxylum (Rutaceae) collected in Bhutan is reported. The total length is 1,408,989 bp with 1,169 coding sequences in 96 contigs, a GC content of 37.3%, and 76 to 77% average nucleotide identity with several other “ Ca . Liberibacter” species.
Publisher: Wiley
Date: 28-03-2016
DOI: 10.1111/AVJ.12415
Abstract: To estimate the prevalence of Theileria orientalis infection for Chitose, Ikeda and Buffeli major piroplasm surface protein (MPSP) types at a herd- and animal-level in beef cattle in the North Coast Livestock Health and Pest Authority (NCLHPA) region of New South Wales (NSW). A total of 24 beef herds in the NCLHPA region containing more than 100 cattle were randomly selected. Blood s les were collected from five animals per herd and tested using Theileria PCR for Chitose, Buffeli and Ikeda. S les were only taken from female cattle older than 2 years, born in the NCLHPA region and apparently healthy at the time of testing. The herd-level prevalence for all MPSP types (Chitose, Ikeda and Buffeli) was 100%, with a 95% confidence interval of 86.3-99.9%. The mean prevalence at an animal level was 83.3%, 92.5% and 95.0% for Theileria Chitose, Buffeli and Ikeda, respectively. Quantitative PCR testing showed that 81.9% of animals had a low-level infection, while 17.0% had a moderate level of infection, and only 1.0% had a high level of infection. The majority of animals had a mixed infection of two or three MPSP types and few animals showed single infection. The results indicate endemicity of T. orientalis, especially the Ikeda type, in the NCLHPA region of Australia.
Publisher: Public Library of Science (PLoS)
Date: 10-03-2014
Publisher: American Chemical Society (ACS)
Date: 11-09-2009
DOI: 10.1021/PR900544X
Abstract: C ylobacter jejuni is the most common cause of bacterial gastroenteritis in the developed world. Immunoproteomics highlighted a 42-45 kDa antigen that comigrated on two-dimensional (2-DE) gels with the C. jejuni major outer membrane protein (MOMP). Predictive analysis revealed two candidates for the identity of the antigen, the most likely of which was the surface-associated lipoprotein, JlpA. Recombinant JlpA (rJlpA) reacted with patient sera, confirming that JlpA is antigenic. Polyclonal antibodies raised against rJlpA reacted against 3 JlpA mass variants from multiple C. jejuni. These variants differed by approximately 1.5 kDa, suggesting the presence of the N-linked C. jejuni glycan on two sites. Soybean agglutinin affinity and 2-DE purified 2 JlpA glycoforms (43.5 and 45 kDa). Their identities were confirmed using mass spectrometry following trypsin digest. Glycopeptides within JlpA variants were identified by proteinase-K digestion, graphite micropurification and MS-MS. Sites of glycosylation were confirmed as asparagines 107 and 146, both of which are flanked by the N-linked sequon. Sequence analysis confirmed that the N146 sequon is conserved in all C. jejuni genomes examined to date, while the N107 sequon is absent in the reference strain NCTC 11168. Western blotting confirmed the presence of only a single JlpA glycoform in both virulent (O) and avirulent (GS) isolates of NCTC 11168. MS analysis showed that JlpA exists as 3 discrete forms, unmodified, glycosylated at N146, and glycosylated at both N(146/107), suggesting glycan addition at N146 is necessary for N107 glycosylation. Glycine extracts and Western blotting revealed that doubly glycosylated JlpA was the predominant form on the C. jejuni JHH1 surface however, glycosylation is not required for antigenicity. This is the first study to identify N-linked glycosylation of a surface-exposed C. jejuni virulence factor and to show strain variation in glycosylation sites.
Publisher: Hindawi Limited
Date: 09-11-2012
DOI: 10.1111/J.1462-5822.2011.01702.X
Abstract: Mycoplasma hyopneumoniae is a major, economically damaging respiratory pathogen. Although M. hyopneumoniae cells bind plasminogen, the identification of plasminogen-binding surface proteins and the biological ramifications of acquiring plasminogen requires further investigation. mhp182 encodes a highly expressed 102 kDa protein (P102) that undergoes proteolytic processing to generate surface-located N-terminal 60 kDa (P60) and C-terminal 42 kDa (P42) proteins of unknown function. We show that recombinant P102 (rP102) binds plasminogen at physiologically relevant concentrations (K(D) ~ 76 nM) increasing the susceptibility of plasmin(ogen) to activation by tissue-specific plasminogen activator (tPA). Recombinant proteins constructed to mimic P60 (rP60) and P42 (rP42) also bound plasminogen at physiologically significant levels. M. hyopneumoniae surface-bound plasminogen was activated by tPA and is able to degrade fibrinogen, demonstrating the biological functionality of M. hyopneumoniae-bound plasmin(ogen) upon activation. Plasmin(ogen) was readily detected in porcine ciliated airways and plasmin levels were consistently higher in bronchoalveolar lavage fluid from M. hyopneumoniae-infected animals. Additionally, rP102 and rP42 bind fibronectin with K(D) s of 26 and 33 nM respectively and recombinant P102 proteins promote adherence to porcine kidney epithelial-like cells. The multifunctional binding ability of P102 and activation of M. hyopneumoniae-sequestered plasmin(ogen) by an exogenous activator suggests P102 plays an important role in virulence.
Publisher: Wiley
Date: 29-08-2013
DOI: 10.1111/AEN.12047
Publisher: MDPI AG
Date: 23-01-2023
DOI: 10.3390/MICROORGANISMS11020297
Abstract: Erysipelothrix rhusiopathiae is a bacterial pathogen that is the causative agent of erysipelas in a variety of animals, including swine, emus, turkeys, muskox, caribou, moose, and humans. This study aims to investigate the population structure and genomic features of Australian isolates of E. rhusiopathiae in the Australian pig industry and compare them to the broader scope of isolates worldwide. A total of 178 isolates (154 Australian, seven vaccine isolates, six international isolates, and 11 of unknown origin) in this study were screened against an MLST scheme and publicly available reference isolates, identifying 59 new alleles, with isolates separating into two main single locus variant groups. Investigation with BLASTn revealed the presence of the spaA gene in 171 (96%) of the isolates, with three main groups of SpaA protein sequences observed amongst the isolates. Novel SpaA protein sequences, categorised here as group 3 sequences, consisted of two sequence types forming separate clades to groups 1 and 2, with amino acid variants at positions 195 (D/A), 303 (G/E) and 323(P/L). In addition to the newly identified groups, five new variant positions were identified, 124 (S/N), 307 (Q/R), 323 (P/L), 379 (M/I), and 400 (V/I). Resistance screening identified genes related to lincomycin, streptomycin, erythromycin, and tetracycline resistance. Of the 29 isolates carrying these resistance genes, 82% belonged to SpaA group 2-N101S (n = 22) or 2-N101S-I257L (n = 2). In addition, 79% (n = 23) of these 29 isolates belonged to MLST group ST 5. Our results illustrate that Australia appears to have a unique ersity of E. rhusiopathiae isolates in pig production industries within the wider global context of isolates.
Publisher: Hindawi Limited
Date: 19-01-2020
DOI: 10.1155/2020/9785861
Abstract: We describe three cases of osteoarticular infection (OAI) in young thoroughbred horses in which the causative organism was identified by MALDI-TOF as Kingella species . The pattern of OAI resembled that reported with Kingella infection in humans. Analysis by 16S rRNA PCR enabled construction of a phylogenetic tree that placed the isolates closer to Simonsiella and Alysiella species , rather than Kingella species . Average nucleotide identity (ANI) comparison between the new isolate and Kingella kingae and Alysiella crassa however revealed low probability that the new isolate belonged to either of these species. This preliminary analysis suggests the organism isolated is a previously unrecognised species.
Publisher: MDPI AG
Date: 15-07-2022
DOI: 10.3390/PATHOGENS11070801
Abstract: Theileria orientalis causes losses to cattle producers in Eastern Asia, Oceania and, more recently, North America. One pathogenic genotype (Ikeda) has been sequenced to the chromosomal level, while only draft genomes exist for globally distributed Chitose and Buffeli genotypes. To provide an accurate comparative gene-level analysis and help further understand their pathogenicity, we sequenced isolates of the Chitose and Buffeli genotypes of T. orientalis using long-read sequencing technology. A combination of several long-read assembly methods and short reads produced chromosomal-level assemblies for both Fish Creek (Chitose) and Goon Nure (Buffeli) isolates, including the first complete and circular apicoplast genomes generated for T. orientalis. Comparison with the Shintoku (Ikeda) reference sequence showed both large and small translocations in T. orientalis Buffeli, between chromosomes 2 and 3 and chromosomes 1 and 4, respectively. Ortholog clustering showed expansion of ABC transporter genes in Chitose and Buffeli. However, differences in several genes of unknown function, including DUF529/FAINT-domain-containing proteins, were also identified and these genes were more prevalent in Ikeda and Chitose genotypes. Phylogenetics and similarity measures were consistent with previous short-read genomic analysis. The generation of chromosomal sequences for these highly prevalent T. orientalis genotypes will also support future studies of population genetics and mixed genotype infections.
Publisher: Microbiology Society
Date: 07-12-2022
Abstract: The Pacific oyster (PO), Crassostrea gigas , is an important commercial marine species but periodically experiences large stock losses due to disease events known as summer mortality. Summer mortality has been linked to environmental perturbations and numerous viral and bacterial agents, indicating this disease is multifactorial in nature. In 2013 and 2014, several summer mortality events occurred within the Port Stephens estuary (NSW, Australia). Extensive culture and molecular-based investigations were undertaken and several potentially pathogenic Vibrio species were identified. To improve species identification and genomically characterise isolates obtained from this outbreak, whole-genome sequencing (WGS) and subsequent genomic analyses were performed on 48 bacterial isolates, as well as a further nine isolates from other summer mortality studies using the same batch of juveniles. Average nucleotide identity (ANI) identified most isolates to the species level and included members of the Photobacterium , Pseudoalteromonas , Shewanella and Vibrio genera, with Vibrio species making up more than two-thirds of all species identified. Construction of a phylogenomic tree, ANI analysis, and pan-genome analysis of the 57 isolates represents the most comprehensive culture-based phylogenomic survey of Vibrios during a PO summer mortality event in Australian waters and revealed large genomic ersity in many of the identified species. Our analysis revealed limited and inconsistent associations between isolate species and their geographical origins, or host health status. Together with ANI and pan-genome results, these inconsistencies suggest that to determine the role that microbes may have in Pacific oyster summer mortality events, isolate identification must be at the taxonomic level of strain. Our WGS data (specifically, the accessory genomes) differentiated bacterial strains, and coupled with associated metadata, highlight the possibility of predicting a strain’s environmental niche and level of pathogenicity.
Publisher: Springer Science and Business Media LLC
Date: 27-04-2018
Publisher: Elsevier BV
Date: 09-2015
DOI: 10.1016/J.VETPAR.2015.08.014
Abstract: Theileria orientalis is an emerging bovine pathogen in Australasia. PCR-based detection methods for this parasite are sensitive but relatively expensive, partly due to costs associated with DNA extraction. An inexpensive and efficient technique was developed for the extraction of T. orientalis DNA from blood based on hypotonic erythrocyte lysis and detergent-proteinase K treatment (DPK method). The DPK method compares favourably to a commercial extraction kit when paired with a T. orientalis multiplex qPCR.
Publisher: Springer Science and Business Media LLC
Date: 16-04-2015
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.
Publisher: Springer Science and Business Media LLC
Date: 25-02-2015
Publisher: Cold Spring Harbor Laboratory
Date: 27-09-2018
DOI: 10.1101/428599
Abstract: ST73 has emerged as one of the most frequently isolated extraintestinal pathogenic E. coli (ExPEC). To examine the localised ersity of ST73 clonal groups including their mobile genetic elements profile, we sequenced the genomes of 16 multiple drug-resistant ST73 isolates from patients with urinary tract infection from a single hospital in Sydney, Australia between 2009 and 2011. Genome sequences were used to generate a SNP-based phylogenetic tree to determine the relationship of these isolates in a global context with ST73 sequences (n=210) from public databases. There was no evidence of a dominant outbreak strain of ST73 in patients from this hospital, rather we identified at least eight separate groups, several of which reoccur, over a two-year period. The inferred phylogeny of all ST73 strains (n=226) including the ST73 Clone D i2 reference genome shows high bootstrap support and clusters into four major groups which correlate with serotype. The Sydney ST73 strains carry a wide variety of virulence-associated genes but the presence of iss , pic and several iron acquisition operons was notable. ST73 is a major clonal lineage of ExPEC that causes urinary tract infections often with uroseptic sequelae but has not garnered substantial scientific interest as the globally disseminated ST131. Isolation of multiple antimicrobial resistant variants of ExPEC ST73 have increased in frequency, but little is known about the carriage of class 1 integrons in this sequence type and the plasmids that are likely to mobilise them. This pilot study examines the ST73 isolates within a single hospital in Sydney Australia and provides the first large-scale core-genome phylogenetic analysis of ST73 utilizing public sequence read datasets. We used this analysis to identify at least 8 sub-groups of ST73 within this single hospital. Mobile genetic elements associated with antibiotic resistance were less erse and only three class 1 integron structures were identified, all sharing the same basic structure suggesting that the acquisition of drug resistance is a recent event. Genomic epidemiological studies are needed to further characterise established and emerging clonal populations of multiple drug resistant ExPEC to identify sources and aid outbreak investigations.
Publisher: American Society for Microbiology
Date: 03-2015
DOI: 10.1128/JCM.03387-14
Abstract: Theileria orientalis is an emerging pathogen of cattle in Asia, Australia, and New Zealand. This organism is a vector-borne hemoprotozoan that causes clinical disease characterized by anemia, abortion, and death, as well as persistent subclinical infections. Molecular methods of diagnosis are preferred due to their sensitivity and utility in differentiating between pathogenic and apathogenic genotypes. Conventional PCR (cPCR) assays for T. orientalis detection and typing are laborious and do not provide an estimate of parasite load. Current real-time PCR assays cannot differentiate between clinically relevant and benign genotypes or are only semiquantitative without a defined clinical threshold. Here, we developed and validated a hydrolysis probe quantitative PCR (qPCR) assay which universally detects and quantifies T. orientalis and identifies the clinically associated Ikeda and Chitose genotypes (UIC assay). Comparison of the UIC assay results with previously validated universal and genotype-specific cPCR results demonstrated that qPCR detects and differentiates T. orientalis with high sensitivity and specificiy. Comparison of quantitative results based on percent parasitemia, determined via blood film analysis and packed cell volume (PCV) revealed significant positive and negative correlations, respectively. One-way analysis of variance (ANOVA) indicated that blood s les from animals with clinical signs of disease contained statistically higher concentrations of T. orientalis DNA than animals with subclinical infections. We propose clinical thresholds to assist in classifying high-, moderate-, and low-level infections and describe how parasite load and the presence of the Ikeda and Chitose genotypes relate to disease.
Publisher: MDPI AG
Date: 30-09-2022
DOI: 10.3390/PATHOGENS11101135
Abstract: Theileria orientalis is an emerging apicomplexan pathogen of cattle occurring in areas populated by the principal vector tick, Haemaphysalis longicornis. Unlike transforming Theileria spp. that induce cancer-like proliferation of lymphocytes via their schizont stage, T. orientalis destroys host erythrocytes during its piroplasm phase resulting in anaemia. The underlying pathogenic processes of T. orientalis infection are poorly understood consequently, there are no vaccines for prevention of T. orientalis infection and chemotherapeutic options are limited. To identify antigens expressed during the piroplasm phase of T. orientalis, including those which may be useful targets for future therapeutic development, we examined the proteome across three common genotypes of the parasite (Ikeda, Chitose and Buffeli) using preparations of piroplasms purified from bovine blood. A combination of Triton X-114 extraction, one-dimensional electrophoresis and LC-MS/MS identified a total of 1113 proteins across all genotypes, with less than 3% of these representing host-derived proteins. Just over three quarters of T. orientalis proteins (78%) identified were from the aqueous phase of the TX-114 extraction representing cytosolic proteins, with the remaining 22% from the detergent phase, representing membrane-associated proteins. All enzymes involved in glycolysis were expressed, suggesting that this is the major metabolic pathway used during the T. orientalis piroplasm phase. Proteins involved in binding and breakdown of haemoglobin were also identified, suggesting that T. orientalis uses haemoglobin as a source of amino acids. A number of proteins involved in host cell interaction were also identified which may be suitable targets for the development of chemotherapeutics or vaccines.
Start Date: 2020
End Date: 09-2024
Amount: $496,349.00
Funder: Australian Research Council
View Funded ActivityStart Date: 2023
End Date: 12-2025
Amount: $434,107.00
Funder: Australian Research Council
View Funded Activity