An observational cohort study on antimicrobial usage on dairy farms in Quebec, Canada.

Quantification of antimicrobial usage (AMU) is crucial to measure the effect of intervention programs, to determine associations between usage and resistance, to compare populations, and for benchmarking purposes. The primary objective of the study was to describe quantitatively the AMU on Quebec dairy farms over 1 yr: (1) the total AMU, (2) the AMU per administration route (intramammary, injectable, oral, intrauterine), and (3) the AMU per antimicrobial class and according to the categorizations of Health Canada and the World Health Organization. The secondary objective was to assess the effect of several characteristics (herd size, level of milk production, and incidence rate of common infectious diseases) on AMU rate. The AMU data were obtained for 101 dairy farms randomly selected in 3 important Quebec dairy regions by collecting and recording all empty drug packaging and invoices for medicated feed (spring 2017 to spring 2018). The AMU rate was reported in number of Canadian defined course doses for cattle per 100 cow-years. The average herd size was 67 cows per farm, and 2/101 farms were certified organic. Overall, an estimated mean of 537 Canadian defined course doses for cattle/100 cow-years was observed. The intramammary route during lactation was the most frequently observed, followed, in decreasing order of usage, by oral route in the feed, intramammary route at drying-off, and injectable route. Oral (other than in animal feed) and intrauterine formulations were infrequently collected from the garbage cans. The 5 most frequently observed antimicrobial classes were, by decreasing order of usage, ionophores, penicillins, aminocoumarins, aminoglycosides, and polymyxins. Highest priority critically important antimicrobials as defined by the World Health Organization were mainly collected from intramammary formulations during lactation followed by injectable and drying-off intramammary formulations. The herd size was positively associated with the total AMU rate but not with the usage rate of highest priority critically important antimicrobials. Incidence of diseases along with preventive use of antimicrobials (drying-off and medicated feed with antimicrobials) explained 48% of the variance in total AMU rate.

Use of a bacterial antimicrobial resistance gene microarray for the identification of resistant Staphylococcus aureus.

As diagnostic and surveillance activities are vital to determine measures needed to control antimicrobial resistance (AMR), new and rapid laboratory methods are necessary to facilitate this important effort. DNA microarray technology allows the detection of a large number of genes in a single reaction. This technology is simple, specific and high-throughput. We have developed a bacterial antimicrobial resistance gene DNA microarray that will allow rapid antimicrobial resistance gene screening for all Gram-positive and Gram-negative bacteria. A prototype microarray was designed using a 70-mer based oligonucleotide set targeting AMR genes of Gram-negative and Gram-positive bacteria. In the present version, the microarray consists of 182 oligonucleotides corresponding to 166 different acquired AMR gene targets, covering most of the resistance genes found in both Gram-negative and -positive bacteria. A test study was performed on a collection of Staphylococcus aureus isolates from milk samples from dairy farms in Québec, Canada. The reproducibility of the hybridizations was determined, and the microarray results were compared with those obtained by phenotypic resistance tests (either MIC or Kirby-Bauer). The microarray genotyping demonstrated a correlation between penicillin, tetracycline and erythromycin resistance phenotypes with the corresponding acquired resistance genes. The hybridizations showed that the 38 antimicrobial resistant S. aureus isolates possessed at least one AMR gene.

Methicillin-resistant Staphylococcus aureus in horses at a veterinary teaching hospital: frequency, characterization, and association with clinical disease.

Methicillin-resistant Staphylococcus aureus (MRSA) is an emerging equine pathogen. To attempt to control nosocomial and zoonotic transmission, an MRSA screening program was established for all horses admitted to the Ontario Veterinary College Veterinary Teaching Hospital, whereby nasal screening swabs were collected at admission, weekly during hospitalization, and at discharge. MRSA was isolated from 120 (5.3%) of 2,283 horses: 61 (50.8%) at the time of admission, 53 (44.2%) during hospitalization, and 6 from which the origin was unclear because an admission swab had not been collected. Clinical infections attributable to MRSA were present or developed in 14 (11.7%) of 120 horses. The overall rate of community-associated colonization was 27 per 1,000 admissions. Horses colonized at admission were more likely to develop clinical MRSA infection than those not colonized at admission (OR 38.9, 95% CI 9.49 160, P < 0.0001). The overall nosocomial MRSA colonization incidence rate was 23 per 1,000 admissions. The incidence rate of nosocomial MRSA infection was at the rate of 1.8 per 1,000 admissions, with an incidence density of 0.88 per 1,000 patient days. Administration of ceftiofur or aminoglycosides during hospitalization was the only risk factor associated with nosocomial MRSA colonization. MRSA screening of horses admitted to a veterinary hospital was useful for identification of community-associated and nosocomial colonization and infection, and for monitoring of infection control practices.

Methicillin-resistant Staphylococcus aureus in horses and horse personnel, 2000-2002.

Methicillin-resistant Staphylococcus aureus (MRSA) infection was identified in 2 horses treated at a veterinary hospital in 2000, prompting a study of colonization rates of horses and associated persons. Seventy-nine horses and 27 persons colonized or infected with MRSA were identified from October 2000 to November 2002; most isolations occurred in a 3-month period in 2002. Twenty-seven (34%) of the equine isolates were from the veterinary hospital, while 41 (51%) were from 1 thoroughbred farm in Ontario. Seventeen (63%) of 27 human isolates were from the veterinary hospital, and 8 (30%) were from the thoroughbred farm. Thirteen (16%) horses and 1 (4%) person were clinically infected. Ninety-six percent of equine and 93% of human isolates were subtypes of Canadian epidemic MRSA-5, spa type 7 and possessed SCCmecIV. All tested isolates from clinical infections were negative for the Panton-Valentine leukocidin genes. Equine MRSA infection may be an important emerging zoonotic and veterinary disease.

Mesenchymal stem cells combined with biphasic calcium phosphate ceramics promote bone regeneration.

The reconstruction and repair of large bone defects, resulting from trauma, cancer or metabolic disorders, is a major clinical challenge in orthopaedics. Clinically available biological and synthetic grafts have clear limitations that necessitate the development of new graft materials and/or strategies. Human mesenchymal stem cells (MSCs), obtained from the adult bone marrow, are multipotent cells capable of differentiating into various mesenchymal tissues. Of particular interest is the ability of these cells to differentiate into osteoblasts, or bone-forming cells. At Osiris, we have extensively characterized MSCs and have demonstrated MSCs can induce bone repair when implanted in vivo in combination with a biphasic calcium phosphate, specifically hydroxyapatite/tricalcium phosphate. This article reviews previous and current studies utilizing mesenchymal stem cells and biphasic calcium phosphates in bone repair.

Emergence of Salmonella typhimurium definitive type 104 (DT104) as an important cause of salmonellosis in horses in Ontario.

Salmonella Typhimurium definitive type 104 (DT104) has emerged as a common cause of salmonellosis in humans and cattle, yet previous reports involving horses are sparse. This study reports the emergence of DT104 as an important pathogen in horses in Ontario. The first clinical case of DT104 infection at the Ontario Veterinary College was identified in 1997. Seventeen cases of DT104-associated salmonellosis were identified between 1997 and 2000. In 2000, 12 of 13 cases of salmonellosis were due to DT104. Salmonellosis in horses due to DT104 is of concern, since the organism is multiresistant to antibiotics and poses increased zoonotic risk. Phage type distribution of Salmonella isolates should be monitored to determine whether DT104 will remain a prevalent equine pathogen.

Right body side performance decrement in congenitally dyslexic children and left body side performance decrement in congenitally hyperactive children.

OBJECTIVE: Simple and complex visuomotor performance of the right and left sides of the body was investigated in 37 children with left hemisphere lesions, 35 children with right hemisphere lesions, 53 developmentally dyslexic children, 29 developmentally hyperactive children, and 35 "normal" children who had endured a very mild head injury with no sequelae. BACKGROUND: Lateralized soft signs, EEG topography, metabolic brain imaging, and neuropsychological test profiles suggest a predominance of left hemisphere dysfunction in dyslexia and right hemisphere dysfunction in hyperactivity. METHOD: Nine measures of lateralized performance were drawn from the Purdue pegboard, Letter cancellation, Rey complex figure, Wechsler Intelligence Scale for Children (WISC) Mazes, and WISC Picture completion tests. RESULTS: The children with left hemisphere lesions manifested significantly weaker performance on test components involving the right body side, relative to the normal controls, on the Purdue pegboard, Rey complex figure (delayed recall condition), and WISC Picture completion tests, and the dyslexic children on the former two. The children with right hemisphere lesions manifested significantly weaker performance on test components involving the left body side, relative to the normal controls, on the WISC Mazes test, as did the hyperactive children. CONCLUSIONS: We propose that (1) contralateral performance decrement results from a unilateral cortical lesion in children, and (2) developmental dyslexia may comprise a slight predominance of left hemisphere dysfunction and developmental hyperactivity of right hemisphere dysfunction.

Inhibition of adherence of Actinobacillus pleuropneumoniae to porcine respiratory tract cells by monoclonal antibodies directed against LPS and partial characterization of the LPS receptors.

Actinobacillus pleuropneumoniae is the causative agent of porcine fibrinohemorrhagic necrotizing pleuropneumonia. We have previously identified the lipopolysaccharides (LPS) as the major adhesin of A. pleuropneumoniae involved in adherence to porcine respiratory tract cells. In the present study, adherence of A. pleuropneumoniae to porcine tracheal frozen sections was inhibited by homologous monovalent Fab fragments produced from monoclonal antibodies 5.1 G8F10 and 102-G02 directed, respectively, against the A. pleuropneumoniae serotype 1 or serotype 2 O-antigens. These results confirm the important role played by LPS in adherence of A. pleuropneumoniae and suggest that these adhesins might represent good vaccine candidates. We also investigated the presence of A. pleuropneumoniae receptors in tracheal cell preparations from piglets of four different breeds. Using Far-Western binding assays, we identified proteins recognized by whole cells of A. pleuropneumoniae reference strains for serotype 1 and 2, and local isolates belonging to the same serotypes, and also recognized by extracted LPS from both reference strains. We confirmed the proteinaceous nature of these LPS-binding molecules by their staining with Coomassie brilliant blue, sensitivity to proteinase K digestion, resistance to sodium m-periodate oxidation, and their inability to stain with glycoprotein-specific reagents. Four low-molecular-mass bands (14-17 kDa) seemed to correspond to histones. We also identified proteins at Mr 38,500 that could represent putative receptors for A. pleuropneumoniae LPS in swine respiratory tract cells.

Evaluation of the hemoglobin-binding activity of Actinobacillus pleuropneumoniae using fluorescein-labeled pig hemoglobin and flow cytometry.

In the present study, the hemoglobin (Hb)-binding activity of Actinobacillus pleuropneumoniae was examined using fluorescein-labeled pig Hb and flow cytometry. Comparison of the Hb-binding activity of A. pleuropneumoniae serotype 1 strain 4074 grown under iron-restricted conditions with cells grown under iron-sufficient conditions indicated that iron-restriction in A. pleuropneumoniae promotes the expression of Hb receptors, and that Hb-binding activity is, at least in part, iron-repressible. Hb-binding activity was also observed in representative strains of A. pleuropneumoniae belonging to serotypes 1 and 2. In addition, A. pleuropneumoniae serotype 1 LPS or capsule isogenic mutants were tested in flow cytometry in order to understand the influence of surface polysaccharides on Hb-binding activity. Experiments with an acapsulated mutant indicated that surface molecules with Hb-binding activity are more exposed at the cell surface in the absence of capsular polysaccharides. However, the Hb-binding activity of LPS mutants analyzed in this study was unchanged compared to the parent strain. The outer membrane proteins profile of A. pleuropneumoniae serotype 1 grown under iron-restricted or iron-sufficient conditions was also evaluated by polyacrylamide gel electrophoresis. Iron-regulated outer membrane proteins were observed under iron-restricted growth conditions which suggests that one or more of these outer membrane proteins may play a role in the Hb-binding activity detected by flow cytometry.

Topically applied diacylglycerols increase pigmentation in guinea pig skin.

Exposure of human and murine melanocytes in vitro to the diacylglycerol (DAG) 1-oleoyl-2-acetyl-sn-glycerol (OAG) markedly increases melanin production within 24 h. To determine whether OAG can increase melanin production in vivo, increasing concentrations of OAG (10-60 mg/ml) in propylene glycol were applied daily for 5 d to shaved guinea pigs. Dose-dependent increased pigmentation was visible first on days 17-22 and persisted for 10-14 weeks. Peak epidermal melanin content in OAG-treated sites was more than twice that of untreated or vehicle-treated sites, as assessed by computerized image analysis of Fontana-Masson stained biopsy cross sections. In another experiment to assess the mechanism of DAG-mediated pigmentation, guinea pigs received twice daily separate applications of OAG, dipalmitoylglycerol (diC16), dioctanoylglycerol (diC8), each 50 mg/ml, 20 microliters/application, and propylene glycol vehicle alone for 5 d. Increased pigmentation was visible after 10 d in the OAG and diC8 sites but not in diC16 or vehicle sites. These results correlate with the reported ability of these compounds to activate protein kinase C in vitro. In a final experiment, guinea pigs received OAG 25 mg/ml three times daily to one test site, and once daily ultraviolet B (70 mJ/cm2, equivalent to 0.6 minimal erythemal dose) radiation to another for 10 d. The OAG and ultraviolet B test sites developed comparable pigmentation by both clinical and histologic criteria. Our data demonstrate that topically applied DAGs can produce a long-lasting increase in epidermal pigmentation, presumably through protein kinase C activation, which clinically and histologically closely resembles ultraviolet-induced tanning.

Keratinocytes and fibroblasts in a human skin equivalent model enhance melanocyte survival and melanin synthesis after ultraviolet irradiation.

To investigate paracrine effects of fibroblasts and keratinocytes on melanocyte behavior after ultraviolet (UV) irradiation, we compared an in vitro skin equivalent model with melanocyte cultures. Human melanocytes were maintained alone in monolayer cultures or on dermal equivalents with or without keratinocytes and were irradiated daily with solar-simulated light. After seven daily UV irradiations, monolayer melanocytes displayed dose-dependent increases in cellular damage. In contrast, melanocytes on dermal equivalents survived strikingly better. Moreover, UV-irradiated skin equivalent melanocytes became highly dendritic as compared with sham-irradiated cells, closely mimicking their morphology in UV-irradiated skin. In addition, in skin equivalents melanocytes migrated from the center to the periphery of the keratinocyte layer after UV irradiation. Melanin production per culture, as measured by 14C-dihydroxyphenylalanine incorporation, was consistently higher in skin equivalent melanocytes than in monolayer melanocytes from the same donor, and it was highest in melanocytes from skin equivalents containing both keratinocytes and fibroblasts. Our data strongly suggest that fibroblasts and keratinocytes modulate melanocyte function in skin. The skin equivalent is a valuable model for investigating paracrine effects on melanocytes after UV irradiation.

The trk family of receptors mediates nerve growth factor and neurotrophin-3 effects in melanocytes.

We have recently shown that (a) human melanocytes express the p75 nerve growth factor (NGF) receptor in vitro; (b) that melanocyte dendricity and migration, among other behaviors, are regulated at least in part by NGF; and (c) that cultured human epidermal keratinocytes produce NGF. We now report that melanocyte stimulation with phorbol 12-tetra decanoate 13-acetate (TPA), previously reported to induce p75 NGF receptor, also induces trk in melanocytes, and TPA effect is further potentiated by the presence of keratinocytes in culture. Moreover, trk in melanocytes becomes phosphorylated within minutes after NGF stimulation. As well, cultures of dermal fibroblasts express neurotrophin-3 (NT-3) mRNA; NT-3 mRNA levels in cultured fibroblasts are modulated by mitogenic stimulation, UV irradiation, and exposure to melanocyte-conditioned medium. Moreover, melanocytes constitutively express low levels of trk-C, and its expression is downregulated after TPA stimulation. NT-3 supplementation to cultured melanocytes maintained in Medium 199 alone prevents cell death. These combined data suggest that melanocyte behavior in human skin may be influenced by neurotrophic factors, possibly of keratinocyte and fibroblast origin, which act through high affinity receptors.

[Acceptability of the Hemoccult test in general medical practice. Results of a pilot study]. / Acceptabilité du test Hémoccult en consultation de médecine générale. Résultats d'une étude pilote.

In view of preparing a controlled trial to assess the efficacy of screening for colorectal cancer by fecal occult blood testing in reducing cancer mortality, a pilot study was performed to evaluate the acceptability rate of the Hemoccult test in non selected subjects consulting in a general practice. 566 subjects aged 45 to 74 years from two small towns, Neuville-aux-Bois (Loiret) and Vicherey (Vosges) were included in the study. The screening test was proposed by GPs to 89.2% of their patients; of these, 5.6% refused the test and 9.4% did not return it. Of the tests carried out, 80.8% were performed spontaneously, and 19.2% after a recall letter. Acceptability depended neither on age or on sex. The patients' confidence in his GP was the most important acceptability factor (60%), followed by explanations the GP had provided, and ease of application. The results suggest that after receiving the correct information, a GP will succeed in prescribing the Hemoccult test to most high-risk subjects and that acceptability then proves excellent. Experience drawn from the pilot study has been very useful in conceiving the on-going controlled trial in Burgundy.