Sally A. Argyle

Multidrug-resistant Escherichia coli from canine urinary tract infections tend to have commensal phylotypes, lower prevalence of virulence determinants and ampC-replicons

Multidrug-resistant Escherichia coli is an emerging clinical challenge in domestic species. Treatment options in many cases are limited. This study characterized MDR E. coli isolates from urinary tract infections in dogs, collected between 2002 and 2011. Isolates were evaluated in terms of β-lactamase production, phylogenetic group, ST type, replicon type and virulence marker profile. Comparisons were made with antibiotic susceptible isolates also collected from dogs with urinary tract infections. AmpC β-lactamase was produced in 67% of the MDR isolates (12/18). Of these, 8 could be specifically attributed to the CMY-2 gene. None of the isolates tested in either group expressed ESBLs. Phylo-group distribution was as expected in the susceptible isolates, with an over representation of the pathogenic B2 phylo-group (67%). In contrast, the phylogenetic background for the MDR group was mixed, with representation of commensal phylo-groups A and B1. The B2 phylo-group represented the smallest proportion (A, B1, B2 or D was 28%, 22%, 11% and 33%, respectively). Virulence marker profiles, evaluated using Identibac® microarray, discriminated between the two groups. Marker sequences for a core panel of virulence determinants were identified in most of the susceptible isolates, but not in most of the MDR isolates. These findings indicate that for MDR isolates, plasmid-mediated AmpC is an important resistance mechanism, and while still capable of causing clinical disease, there is evidence for a shift towards phylogenetic groups of reduced inferred virulence potential. There was no evidence of zoonotic potential in either the susceptible or MDR urinary tract isolates in this study.

Characterisation and differentiation potential of bone marrow derived canine mesenchymal stem cells.

Mesenchymal stem cells (MSCs) have potential for use in regenerative therapeutics, since they are capable of multi-lineage differentiation. In this study, primary canine MSCs (cMSCs) were isolated from bone marrow aspirates and characterised using marker expression and morphology. cMSCs expressed CD44 and STRO-1, but not CD34 or CD45. Morphologically, cMSCs were similar to previously described MSCs and were capable of chondrocyte differentiation towards articular type cartilage, characterised by increased collagen type II vs. collagen type I expression and expression of Sox-9. cMSCs demonstrated no significant alterations in marker profiles and failed to differentiate into cardiomyocytes in response to a cardiac differentiation protocol or when co-cultured with canine cardiac stem cells. The study indicated that cMSCs can be derived readily from bone marrow and are capable of differentiation into articular cartilage, but appear to have limited ability to differentiate into cardiomyocytes using current protocols.

Characterisation and cardiac directed differentiation of canine adult cardiac stem cells

This study describes the isolation and characterisation of adult canine cardiac stem cells, and explores their ability to differentiate into cardiac myocytes. Direct comparisons are also made with available human data. Atrial cardiac explants were taken from dogs post-mortem and cultured to isolate adult stem cells. Cells were able to survive successive passages in serum-free media, were able to form cardiospheres, and under controlled culture conditions were capable of clonal expansion, demonstrating their ability for self-renewal. Characterisation of these cells demonstrated the following marker profile: c-kit, GATA 4 and flk-1 positive; cardiac troponin T and NKx2.5 low. Cardiac lineage directed differentiation was performed based on the published literature. Gene expression studies demonstrated that cardiac directed differentiation was partially achieved, with up-regulation of cardiac troponin T and NKx2.5, and down-regulation of c-kit and endothelial lineage markers. However the cells did not express the ryanodine receptor or β(1)-adrenergic receptors and did not contract spontaneously.

The long-acting COX-2 inhibitor mavacoxib (Trocoxil™) has anti-proliferative and pro-apoptotic effects on canine cancer cell lines and cancer stem cells in vitro

BackgroundThe NSAID mavacoxib (Trocoxil™) is a recently described selective COX-2 inhibitor used for the management of inflammatory disease in dogs. It has a long plasma half-life, requiring less frequent dosing and supporting increased owner compliance in treating their dogs. Although the use of NSAIDs has been described in cancer treatment in dogs, there are no studies to date that have examined the utility of mavacoxib specifically.ResultsIn this study we compared the in vitro activity of a short-acting non-selective COX inhibitor (carprofen) with mavacoxib, on cancer cell and cancer stem cell survival. We demonstrate that mavacoxib has a direct cell killing effect on cancer cells, increases apoptosis in cancer cells in a manner that may be independent of caspase activity, and has an inhibitory effect on cell migration. Importantly, we demonstrate that cancer stem cells derived from osteosarcoma cell lines are sensitive to the cytotoxic effect of mavacoxib.ConclusionsBoth NSAIDs can inhibit cancer cell proliferation and induce apoptosis in vitro. Importantly, cancer stem cells derived from an osteosarcoma cell line are sensitive to the cytotoxic effect of mavacoxib. Our results suggest that mavacoxib has anti-tumour effects and that this in vitro anti-cancer activity warrants further study.

Apparent bypass of negative selection in CD8 + tumours in CD2- myc transgenic mice

A role for antigen stimulation in lymphoid neoplasia has been postulated and is supported by indirect evidence that suggests that the interaction of antigen with both T cells and B cells may constitute an epigenetic event that can contribute to tumour induction or tumour progression. Using myc-bearing transgenic mice that develop mainly clonal T-cell lymphomas we have investigated the possibility that endogenous antigen-mediated clonal deletion might be overridden in tumorigenesis. CD2-myc transgenic mice were backcrossed on to a CBA/Ca background to ensure Mtv-mediated deletion of V beta 11-expressing T cells in the resultant offspring. Lymphomas arising from these mice were subsequently screened for V beta 11 expression. There was a clear correlation between the age at which mice developed neoplasia and the tumour phenotype. Mice with CD4- CD8+ tumours succumbed to thymic lymphoma at a significantly younger age than mice developing CD4+ CD8+ tumours. A small number of tumours consisted of the forbidden V beta 11 phenotype, showing that cells vulnerable to transformation could escape negative selection. The majority of the V beta 11-positive tumours were CD4- CD8+ and were only observed in mice showing clinical evidence of tumour development at a relatively young age. The phenotype of these cells and the age at which tumours arose suggests that T cells escaping tolerance may be susceptible to transformation.

Type III Secretion-Dependent Sensitivity of Escherichia coli O157 to Specific Ketolides

ABSTRACT A subset of Gram-negative bacterial pathogens uses a type III secretion system (T3SS) to open up a conduit into eukaryotic cells in order to inject effector proteins. These modulate pathways to enhance bacterial colonization. In this study, we screened established bioactive compounds for any that could repress T3SS expression in enterohemorrhagic Escherichia coli (EHEC) O157. The ketolides telithromycin and, subsequently, solithromycin both demonstrated repressive effects on expression of the bacterial T3SS at sub-MICs, leading to significant reductions in bacterial binding and actin-rich pedestal formation on epithelial cells. Preincubation of epithelial cells with solithromycin resulted in significantly less attachment of E. coli O157. Moreover, bacteria expressing the T3SS were more susceptible to solithromycin, and there was significant preferential killing of E. coli O157 bacteria when they were added to epithelial cells that had been preexposed to the ketolide. This killing was dependent on expression of the T3SS. Taken together, this research indicates that the ketolide that has accumulated in epithelial cells may traffic back into the bacteria via the T3SS. Considering that neither ketolide induces the SOS response, nontoxic members of this class of antibiotics, such as solithromycin, should be considered for future testing and trials evaluating their use for treatment of EHEC infections. These antibiotics may also have broader significance for treating infections caused by other pathogenic bacteria, including intracellular bacteria, that express a T3SS.

Nanocrystalline silver dressing and subatmospheric pressure therapy following neoadjuvant radiation therapy and surgical excision of a feline injection site sarcoma

Clinical summary This is the first clinical report of use of a combination of nanocrystalline silver and subatmospheric pressure therapy to treat a resistant wound infection, following tumour removal and radiation therapy, in a difficult-to-manage surgical site in a cat. Practical relevance The therapy was well tolerated and the authors suggest it is a valid treatment protocol for management of non-healing or infected wounds in the cat.

Phylogenomic approaches to determine the zoonotic potential of Shiga toxin-producing Escherichia coli (STEC) isolated from Zambian dairy cattle

This study assessed the prevalence and zoonotic potential of Shiga toxin-producing Escherichia coli (STEC) sampled from 104 dairy units in the central region of Zambia and compared these with isolates from patients presenting with diarrhoea in the same region. A subset of 297 E. coli strains were sequenced allowing in silico analyses of phylo- and sero-groups. The majority of the bovine strains clustered in the B1 ‘commensal’ phylogroup (67%) and included a diverse array of serogroups. 11% (41/371) of the isolates from Zambian dairy cattle contained Shiga toxin genes (stx) while none (0/73) of the human isolates were positive. While the toxicity of a subset of these isolates was demonstrated, none of the randomly selected STEC belonged to key serogroups associated with human disease and none encoded a type 3 secretion system synonymous with typical enterohaemorrhagic strains. Positive selection for E. coli O157:H7 across the farms identified only one positive isolate again indicating this serotype is rare in these animals. In summary, while Stx-encoding E. coli strains are common in this dairy population, the majority of these strains are unlikely to cause disease in humans. However, the threat remains of the emergence of strains virulent to humans from this reservoir.

Convergence of plasmid architectures drives emergence of multi-drug resistance in a clonally diverse Escherichia coli population from a veterinary clinical care setting

Highlights • Multi-drug resistant E. coli associated with urinary tract infections in dogs have a commensal strain background.• Beta-lactam resistance is associated with blaCMY-2 located exclusively on a highly clonal IncI1 plasmid.• IncI1 plasmids carried no other identifiable resistance genes.• Isolates in some cases carried up to 5 plasmids, responsible for carriage of the additional resistances.

Antibiotic prophylaxis in veterinary cancer chemotherapy: A review and recommendations

Bacterial infection following cancer chemotherapy-induced neutropenia is a serious cause of morbidity and mortality in human and veterinary patients. Antimicrobial prophylaxis is controversial in the human oncology field, as any decreased incidence in bacterial infections is countered by patient adverse effects and increased antimicrobial resistance. Comprehensive guidelines exist to aid human oncologists in prescribing antimicrobial prophylaxis but similar recommendations are not available in veterinary literature. As the veterinarians role in antimicrobial stewardship is increasingly emphasized, it is vital that veterinary oncologists implement appropriate antimicrobial use. By considering the available human and veterinary literature we present an overview of current clinical practices and are able to suggest recommendations for prophylactic antimicrobial use in veterinary cancer chemotherapy patients.