Preinfection in vitro chemotaxis, phagocytosis, oxidative burst, and expression of CD11/CD18 receptors and their predictive capacity on the outcome of mastitis induced in dairy cows with Escherichia coli.

Four to 6 wk after parturition, 12 cows in second, fourth, or fifth lactation were experimentally infected in one gland with Escherichia coli. The capacity of chemotaxis, phagocytosis, oxidative burst, and expression of CD11/CD18 receptors to predict the severity of IMI was measured. Bacterial counts in the infected quarter, expressed as area under the curve, and residual milk production in the uninfected quarters were compared to determine severity of the infection. Although these two outcome parameters were highly negatively correlated, regression models with preinfection tests for leukocyte function fitted best with bacterial counts as an outcome parameter. Of the preinfection tests for leukocyte function, chemotaxis best predicted the outcome of the IMI that had been experimentally induced by E. coli. The number of circulating peripheral leukocytes just prior to inoculation was used to predict 52 and 45% of the severity of IMI for bacterial counts and residual milk production, respectively. As a categorical variable, parity predicted 75 and 56% of the severity of IMI expressed as bacterial counts and residual milk production, respectively. Because of the strong effect of parity on the outcome of the experimentally induced mastitis, analysis was performed to discriminate between second parity cows and older cows. Significant differences were found for the number of circulating peripheral leukocytes and for the expression of CD11b/CDl8 and CD11c/CD18 receptors between younger and older cows

Велес – повелитель мертвих

In providing advice on herd health, veterinarians need to be aware of farmers' goals and priorities. To determine the level of awareness, 29 veterinarians from 15 practices completed questionnaires during visits to dairy farms within the scope of veterinary herd health management (VHHM) programmes. The farmers (n = 30) were asked to complete a questionnaire and their discussions with the veterinarian were recorded using a voice recorder. Herd performance goals were set by the farmer and veterinarian in 24% of cases. Veterinarians who did not set goals indicated that they and the farmer 'intuitively knew' what each wanted to achieve, and that the setting of performance goals was considered 'too formal'. Veterinarians often could not identify a farmer's main goal, and typically found milk production and nutrition significantly more important (P <0.01, and P <0.02, respectively), and fertility significantly less important (P <0.01) than the farmers. During on-farm conversations, veterinarians did not actively seek to identify farmers' goals or problems, suggest a co-operative strategy or summarise any advice given. The findings of this survey suggest that veterinarians need to focus more on goal setting, since awareness of goals and priorities is important for both communication and compliance with advice given. The needs of farmers with respect to herd health should also be more actively sought by veterinarians as the findings indicate that most farmers do not readily volunteer such information. (C) 2013 Published by Elsevier Ltd

Kinetochore inactivation by expression of a repressive mRNA.

Differentiation programs such as meiosis depend on extensive gene regulation to mediate cellular morphogenesis. Meiosis requires transient removal of the outer kinetochore, the complex that connects microtubules to chromosomes. How the meiotic gene expression program temporally restricts kinetochore function is unknown. We discovered that in budding yeast, kinetochore inactivation occurs by reducing the abundance of a limiting subunit, Ndc80. Furthermore, we uncovered an integrated mechanism that acts at the transcriptional and translational level to repress NDC80 expression. Central to this mechanism is the developmentally controlled transcription of an alternate NDC80 mRNA isoform, which itself cannot produce protein due to regulatory upstream ORFs in its extended 5' leader. Instead, transcription of this isoform represses the canonical NDC80 mRNA expression in cis, thereby inhibiting Ndc80 protein synthesis. This model of gene regulation raises the intriguing notion that transcription of an mRNA, despite carrying a canonical coding sequence, can directly cause gene repression

Improved genome-wide localization by ChIP-chip using double-round T7 RNA polymerase-based amplification

Chromatin immunoprecipitation combined with DNA microarrays (ChIP-chip) is a powerful technique to detect in vivo protein–DNA interactions. Due to low yields, ChIP assays of transcription factors generally require amplification of immunoprecipitated genomic DNA. Here, we present an adapted linear amplification method that involves two rounds of T7 RNA polymerase amplification (double-T7). Using this we could successfully amplify as little as 0.4 ng of ChIP DNA to sufficient amounts for microarray analysis. In addition, we compared the double-T7 method to the ligation-mediated polymerase chain reaction (LM-PCR) method in a ChIP-chip of the yeast transcription factor Gsm1p. The double-T7 protocol showed lower noise levels and stronger binding signals compared to LM-PCR. Both LM-PCR and double-T7 identified strongly bound genomic regions, but the double-T7 method increased sensitivity and specificity to allow detection of weaker binding sites

The effect of milk production level on host resistance of dairy cows, as assessed by the severity of experimental Escherichia coli mastitis

The culturability of 10 strains of Campylobacter jejuni and Campylobacter coli was studied after the bacteria were exposed to acid conditions for various periods of time. Campylobacter cells could not survive 2 h under acid conditions (formic acid at pH 4). The 10 Campylobacter strains could not be recovered, even when enrichment media were used. Viable cells, however, could be detected by a double-staining (5-cyano-2,3-ditolyl tetrazolium chloride [CTC]-4',6'-diamidino-2-phenylindole [DAPI]) technique, demonstrating that the treated bacteria changed into a viable but nonculturable (VBNC) form; the number of VBNC forms decreased over time. Moreover, some VBNC forms of Campylobacter could be successfully resuscitated in specific-free-pathogen fertilized eggs via two routes, amniotic and yolk sac injecting

An investigation of the dynamics of intramammary infections acquired during the dry period on European dairy farms

The dry period is acknowledged as playing a key role in mastitis epidemiology and yet surprisingly few studies have explored dry period infection dynamics in detail. The aim of this study was to investigate the dynamics of intramammary infection across a cohort of dairy herds in Europe. Five hundred and twenty-two cows were recruited from 12 farms in 6 European countries. All cows received antibiotic dry cow therapy but teat sealants were not used. All quarters of all cows were sampled for bacteriology at drying off and in the week immediately postcalving. Two ipsilateral quarters were also sampled for bacteriology in each cow 2 and 6 wk after drying off. Cows were body condition scored and teats assessed for cleanliness at all sampling time points and for the presence of a keratin plug during the dry period. Other cow-level parameters such as historic somatic cell counts and milk yields before drying off were collated from farm records. Univariable and multivariable analyses were undertaken to investigate the etiology, prevalence, and dynamics of infection during the dry period and associated influential factors. In summary, environmental mastitis pathogens predominated. Although gram-positive major pathogens were typically well controlled and did not increase in prevalence across the dry period, gram-negative pathogens generally increased in prevalence. There was an increase in the number of quarters that yielded no growth across the dry period, although this was driven by minor rather than major mastitis pathogen control. Other than the presence of a gram-positive or gram-negative pathogen 6 wk after drying off, the measured parameters were not influential when considering their effect on the presence of pathogens postcalving. Analysis also suggested that the early and mid dry period may be more important with respect to the timing of acquisition of infection than previously thought. We observed substantial variation in the etiology and prevalence of different pathogens on different farms with, in all cases, at least one of the 12 herds experiencing the opposite of the others with respect to increases and decreases in pathogen prevalence. Overall, this study confirms the importance of the dry period in mastitis epidemiology but highlights the importance of assessing and understanding infection dynamics on individual units. The lack of influence of the cow and quarter factors measured in this study suggests that herd and management factors may be more influential

Addition of meloxicam to the treatment of clinical mastitis improves subsequent reproductive performance

AbstractA blinded, negative controlled, randomized intervention study was undertaken to test the hypothesis that addition of meloxicam, a nonsteroidal anti-inflammatory drug, to antimicrobial treatment of mild to moderate clinical mastitis would improve fertility and reduce the risk of removal from the herd. Cows (n=509) from 61 herds in 8 regions (sites) in 6 European countries were enrolled. Following herd-owner diagnosis of mild to moderate clinical mastitis within the first 120d of lactation in a single gland, the rectal temperature, milk appearance, and California Mastitis Test score were assessed. Cows were randomly assigned within each site to be treated either with meloxicam or a placebo (control). All cows were additionally treated with 1 to 4 intramammary infusions of cephalexin and kanamycin at 24-h intervals. Prior to treatment and at 14 and 21d posttreatment, milk samples were collected for bacteriology and somatic cell count. Cows were bred by artificial insemination and pregnancy status was subsequently defined. General estimating equations were used to determine the effect of treatment (meloxicam versus control) on bacteriological cure, somatic cell count, the probability of being inseminated by 21d after the voluntary waiting period, the probability of conception to first artificial insemination, the number of artificial insemination/conception, the probability of pregnancy by 120 or 200d postcalving, and the risk of removal by 300d after treatment. Cox’s proportional hazards models were used to test the effect of treatment on the calving to first insemination and calving to conception intervals. Groups did not differ in terms of age, clot score, California Mastitis Test score, rectal temperature, number of antimicrobial treatments given or bacteria present at the time of enrollment, but cows treated with meloxicam had greater days in milk at enrollment. Cows treated with meloxicam had a higher bacteriological cure proportion than those treated with the placebo [0.66 (standard error=0.04) versus 0.50 (standard error=0.06), respectively], although the proportion of glands from which no bacteria were isolated posttreatment did not differ between groups. No difference was observed in the somatic cell count between groups pre- or posttreatment. The proportion of cows that underwent artificial insemination by 21d after the voluntary waiting period was unaffected by treatment. Treatment with meloxicam was associated with a higher proportion of cows conceiving to their first artificial insemination (0.31 versus 0.21), and a higher proportion of meloxicam-treated cows were pregnant by 120d after calving (0.40 versus 0.31). The number of artificial inseminations required to achieve conception was lower in the meloxicam compared with control cows (2.43 versus 2.92). No difference was observed between groups in the proportion of cows pregnant by 200d after calving or in the proportion of cows that were culled, died, or sold by 300d after calving (17% versus 21% for meloxicam versus control, respectively). It was concluded that use of meloxicam, in conjunction with antimicrobial therapy, for mild to moderate cases of clinical mastitis, resulted in a higher probability of bacteriological cure, an increased probability of conception to first artificial insemination, fewer artificial inseminations, and a greater proportion of cows pregnant by 120d in milk