Dual role of splenic mononuclear and polymorphonuclear cells in the outcome of ciprofloxacin treatment of Salmonella enterica infections.

OBJECTIVES: To determine the immune cell populations associated with Salmonella enterica serovar Typhimurium before and after ciprofloxacin treatment using a murine model of systemic infection. The effect of depletion of immune cells associating with Salmonella on treatment outcome was also determined. METHODS: We infected mice with a Salmonella enterica serovar Typhimurium strain expressing GFP and used multicolour flow cytometry to identify splenic immune cell populations associating with GFP-positive Salmonella before and after treatment with ciprofloxacin. This was followed by depletion of different immune cell populations using antibodies and liposomes. RESULTS: Our results identified CD11b+CD11chi/lo (dendritic cells/macrophages) and Ly6G+CD11b+ (neutrophils) leucocytes as the main host cell populations that are associated with Salmonella after ciprofloxacin treatment. We therefore proceeded to test the effects of depletion of such populations during treatment. We show that depletion of Ly6G+CD11b+ populations resulted in an increase in the number of viable bacterial cells in the spleen at the end of ciprofloxacin treatment. Conversely, treatment with clodronate liposomes during antimicrobial treatment, which depleted the CD11b+CD11chi/lo populations, resulted in lower numbers of viable bacteria in the tissues. CONCLUSIONS: Our study identified host cells where Salmonella bacteria persist during ciprofloxacin treatment and revealed a dual and opposing effect of removal of Ly6G+CD11b+ and CD11b+CD11chi/lo host cells on the efficacy of antimicrobial treatment. This suggests a dichotomy in the role of these populations in clearance/persistence of Salmonella during antimicrobial treatment

Reduced Lentivirus Susceptibility in Sheep with TMEM154 Mutations

Visna/Maedi, or ovine progressive pneumonia (OPP) as it is known in the United States, is an incurable slow-acting disease of sheep caused by persistent lentivirus infection. This disease affects multiple tissues, including those of the respiratory and central nervous systems. Our aim was to identify ovine genetic risk factors for lentivirus infection. Sixty-nine matched pairs of infected cases and uninfected controls were identified among 736 naturally exposed sheep older than five years of age. These pairs were used in a genome-wide association study with 50,614 markers. A single SNP was identified in the ovine transmembrane protein (TMEM154) that exceeded genome-wide significance (unadjusted p-value 3×10−9). Sanger sequencing of the ovine TMEM154 coding region identified six missense and two frameshift deletion mutations in the predicted signal peptide and extracellular domain. Two TMEM154 haplotypes encoding glutamate (E) at position 35 were associated with infection while a third haplotype with lysine (K) at position 35 was not. Haplotypes encoding full-length E35 isoforms were analyzed together as genetic risk factors in a multi-breed, matched case-control design, with 61 pairs of 4-year-old ewes. The odds of infection for ewes with one copy of a full-length TMEM154 E35 allele were 28 times greater than the odds for those without (p-value<0.0001, 95% CI 5–1,100). In a combined analysis of nine cohorts with 2,705 sheep from Nebraska, Idaho, and Iowa, the relative risk of infection was 2.85 times greater for sheep with a full-length TMEM154 E35 allele (p-value<0.0001, 95% CI 2.36–3.43). Although rare, some sheep were homozygous for TMEM154 deletion mutations and remained uninfected despite a lifetime of significant exposure. Together, these findings indicate that TMEM154 may play a central role in ovine lentivirus infection and removing sheep with the most susceptible genotypes may help eradicate OPP and protect flocks from reinfection

Emerging viruses of zoonotic and veterinary importance.

To enable discussion of all aspects of emerging virus infections, an Emerging Viruses meeting was held at the University of Nottingham, UK, on 27–29 July 2015. Given the success of this meeting, a second meeting was organised, now called ‘Emerging Viruses of Zoonotic and Veterinary Importance’, at Churchill College, Cambridge, UK, on 24–26 July 2017 to encourage discussion of emerging virus infections from a One Health perspective.Newnham College, Cambridg

Monoclonal antibodies raised to the human FOXP3 protein can be used effectively for detecting Foxp3(+) T cells in other mammalian species.

A population of primarily CD4(+)CD25(+) regulatory T cells (Tregs), that have a critical role in maintaining the balance between tolerance and immunity, have been identified through their ability to provide protection against autoimmune disease. There is considerable interest in further exploring the role that Tregs play in autoimmune disease, cancer, and in regulating the immune response to pathogens. Currently the best single marker for labelling Tregs is the forkhead transcription factor FOXP3. Consistent with its essential functional role, sequence alignment showed that the FOXP3 protein is highly conserved across mammalian species. Lymphoid tissues were analysed for nuclear Foxp3 protein expression by immunohistochemistry to evaluate the utility of monoclonal antibodies raised to the human FOXP3 protein for labelling Foxp3(+) Tregs in other mammalian species. The T-cell specificity of those anti-FOXP3 antibodies that gave the most effective staining on each species was confirmed by double labelling with FOXP3 and CD3. Antibodies 236A/E7 and 206D/B1 showed least reactivity with other species, while 259D/C7 commonly exhibited non-specific nuclear staining of non-human lymphoid tissues. Antibodies 86D/D6, 150D/E4 and 157B/F4 are recommended as those which are most effective for labelling Foxp3(+) Tregs in studies utilising animal models

Cloning and sequencing of ovine Flt3 ligand

A cDNA (879 bp) containing the complete open reading frame of ovine Fms-related tyrosine kinase 3 ligand (Flt3-L) was amplified by reverse transcriptase polymerase chain reaction (RT-PCR), cloned and sequenced. The deduced amino acid sequence has 97.6% similarity with bovine Flt3-L isoform 1 and shares the long cytoplasmic domain observed in bovine Flt3-L but not in human Flt3-L

Clinical and histopathological characterization of a large animal (ovine) model of collagen-induced arthritis

Collagen induced arthritis (CIA) is the most studied and used rheumatoid arthritis (RA) model in animals, as it shares many pathological and immunological features of the human disease. The aim of this study was to characterize clinical and immunological aspects of the ovine CIA model, and develop lameness and histopathological scoring systems, in order to validate this model for use in therapeutic trials. Sheep were sensitized to bovine type II collagen (BCII), arthritis was induced by injection of bovine collagen type II into the hock joint and the response was followed for two weeks. Clinical signs of lameness and swelling were evident in all sheep and gross thickening of the synovium surrounding the tibiotarsal joint and erosion on the cartilage surface in the arthritic joints. Leucocyte cell counts were increased in synovial fluid and there was synovial hyperplasia, thickening of the intimal layer, inflammation and marked angiogenesis in the synovial tissue. There was a large influx of monocytes and lymphocytes into the synovial tissue, and increased expression of TNF-α and IL-1β in arthritic intima, angiogenesis and upregulation of VCAM-1. CIA in sheep appears to be an excellent large animal model of RA and has the potential for testing biological therapeutics for the treatment of rheumatoid arthritis. © 2014 Elsevier B.V. All rights reserved.Collagen induced arthritis (CIA) is the most studied and used rheumatoid arthritis (RA) model in animals, as it shares many pathological and immunological features of the human disease. The aim of this study was to characterize clinical and immunological aspects of the ovine CIA model, and develop lameness and histopathological scoring systems, in order to validate this model for use in therapeutic trials. Sheep were sensitized to bovine type II collagen (BCII), arthritis was induced by injection of bovine collagen type II into the hock joint and the response was followed for two weeks. Clinical signs of lameness and swelling were evident in all sheep and gross thickening of the synovium surrounding the tibiotarsal joint and erosion on the cartilage surface in the arthritic joints. Leucocyte cell counts were increased in synovial fluid and there was synovial hyperplasia, thickening of the intimal layer, inflammation and marked angiogenesis in the synovial tissue. There was a large influx of monocytes and lymphocytes into the synovial tissue, and increased expression of TNF-α and IL-1β in arthritic intima, angiogenesis and upregulation of VCAM-1. CIA in sheep appears to be an excellent large animal model of RA and has the potential for testing biological therapeutics for the treatment of rheumatoid arthritis