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Leptospirosis is a global zoonotic disease. Pathogenic Leptospira species, the causative agent of leptospirosis, colonize the renal tubules of chronically infected maintenance hosts such as dogs, rats and cattle. Maintenance hosts typically remain clinically asymptomatic and shed leptospires into the environment via urine. In contrast, accidental hosts such as humans can suffer severe acute forms of the disease. Infection results from direct contact with infected urine or indirectly, through contaminated water sources. In this study, a quantitative real-time PCR specific for lipL32 was designed to detect the urinary shedding of leptospires from dogs. The sensitivity and specificity of the assay was validated using both a panel of pathogenic Leptospira species and clinical microbial isolates, and samples of urine collected from experimentally infected rats and non-infected controls. The lower limit of detection was approximately 3 genome equivalents per reaction. The assay was applied to canine urine samples collected from local dog sanctuaries and the University Veterinary Hospital (UVH) at University College Dublin. Of 525 canine urine samples assayed, 37 were positive, indicating a prevalence of urinary shedding of leptospires of 7.05%. These results highlight the need to provide effective canine vaccination strategies and raise public health awareness.Dear Dr. Van Belkum, Thank you for considering our manuscript entitled "Detection and quantification of leptospires in urine of dogs; a maintenance host for the zoonotic disease leptospirosis" for publication in the European Journal of Clinical Microbiology and Infectious Diseases. I would also like to thank the reviewer for constructive critique. Please find below a response to each point raised by the reviewer; a revised manuscript has been prepared accordingly and is provided for your consideration for publication as a brief report.Reviewer #2:Comment: This manuscript has improved with the changes made, however I was expecting to see a concise communication, but the manuscript still appears as a full research paper.Response: The manuscript has been revised, as also requested by the editors, as a brief report.Comment: Some justification for the choice of specificity test organisms is provided in the cover letter; however, I still have concerns with regard to those tested. Are these organisms those that would normally be encountered as potential pathogens in dogs, especially from urine samples?Response: The reviewer is correct; these are microbes routinely encountered either as pathogens or contaminants in submitted canine urine samples and are routinely found in diagnostic laboratories. 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60 61 62 63 64 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 ...

Guidelines on prudent antimicrobial use in veterinary medicine have been developed to reduce inappropriate prescribing of antimicrobials. Such guidelines focus mainly on the clinical and pharmacological indications for prescribing. A questionnaire study of veterinary surgeons engaged in cattle practice was completed to determine if non-clinical issues influence the decision to prescribe antimicrobials, and to assess if pharmacological and non-pharmacological issues influence the choice of antimicrobial prescribed. Non-clinical issues, including issues related to professional stress, influenced the prescribing decision of the majority of respondents. However, the nature of the veterinarian-client relationship did not influence the prescribing behaviour of the majority of respondents. Pharmacological and non-pharmacological issues influenced the choice of antimicrobial prescribed. The veterinary surgeon's prior experience of a drug was considered 'often' or 'always' by 95.7 per cent of respondents when making this decision. The findings of this study have implications for the recognition and management of stress within the profession, and for the development of intervention strategies to reduce inappropriate antimicrobial prescribing.

Mycobacterium avium subspecies paratuberculosis (MAP) is the aetiological agent of Johne’s disease (JD), a chronic enteritis in ruminants that causes substantial economic loses to agriculture worldwide. Current diagnostic assays are hampered by low sensitivity and specificity that seriously complicate disease control; a new generation of diagnostic and prognostic assays are therefore urgently needed. Circulating microRNAs (miRNAs) have been shown to have significant potential as novel biomarkers for a range of human diseases, but their potential application in the veterinary sphere has been less well characterised. The aim of this study was therefore to apply RNA-sequencing approaches to serum from an experimental JD infection model as a route to identify novel diagnostic and prognostic miRNA biomarkers. Sera from experimental MAP-challenged calves (n = 6) and age-matched controls (n = 6) were used. We identified a subset of known miRNAs from bovine serum across all samples, with approximately 90 being at potentially functional abundance levels. The majority of known bovine miRNAs displayed multiple isomiRs that differed from the canonical sequences. Thirty novel miRNAs were identified after filtering and were found within sera from all animals tested. No significant differential miRNA expression was detected when comparing sera from MAP-challenged animals to their age-matched controls at six-month’s post-infection. However, comparing sera from pre-infection bleeds to six-month’s post-infection across all 12 animals did identify increased miR-205 (2-fold) and decreased miR-432 (2-fold) within both challenged and control groups, which suggests changes in circulating miRNA profiles due to ageing or development (P<0.00001). In conclusion our study has identified a range of novel miRNA in bovine serum, and shown the utility of small RNA sequencing approaches to explore the potential of miRNA as novel biomarkers for infectious disease in cattle.