Effect of β-hydroxybutyric acid, parity, and body condition score on phenotype and proliferative capacity of colostral mononuclear leukocytes of high-yielding dairy cows

In neonatal calves, the ingestion of colostrum is imperative for preventing infectious diseases. Investigations into the transfer of passive immunity of cattle have primarily focused on the importance of colostral immunoglobulins, with a recent increase in focus on understanding the role of colostral leukocytes. The main objective of the present study was to measure the influence of parity, body condition score, serum nonesterified fatty acids, and serum beta-hydroxybutyrate concentrations of periparturient cows on phenotype and mitogen-and antigen-induced proliferative capacity of bovine colostral leukocytes. Holstein-Friesian cows (n = 141) were intramuscularly vaccinated at 60 and 30 d before the expected parturition date with a tetanus toxoid vaccine. Of these 141 animals, 28 primiparous and 72 multiparous cows were sampled immediately. after parturition. Colostrum mononuclear cell populations were identified by flow cytometry using bovine cluster of differentiation markers, and the proliferative capacity of these cells was determined using a H-3-thymidine proliferation assay. Under-conditioned cows had a significantly higher percentage of colostral macrophages than normal-conditioned animals, whereas over-conditioned cows had significantly more colostral B-lymphocytes. Serum beta-hydroxybutyrate was significantly associated with higher numbers of colostral T-lymphocytes and macrophages. Heifers had significantly higher mitogen- and antigen-induced proliferation of their colostral leukocytes than third parity or older cows. In conclusion, body condition score, parity, and serum beta-hydroxybutyrate concentration of periparturient high-yielding dairy cows were shown to influence the number of colostral macrophages or the mitogen-and antigen-induced proliferation of colostral leukocytes, possibly influencing the cellular immunity of the newborn calf

Local host response following an intramammary challenge with Staphylococcus fleurettii and different strains of Staphylococcus chromogenes in dairy heifers

Coagulase-negative staphylococci (CNS) are a common cause of subclinical mastitis in dairy cattle. The CNS inhabit various ecological habitats, ranging between the environment and the host. In order to obtain a better insight into the host response, an experimental infection was carried out in eight healthy heifers in mid-lactation with three different CNS strains: a Staphylococcus fleurettii strain originating from sawdust bedding, an intramammary Staphylococcus chromogenes strain originating from a persistent intramammary infection (S. chromogenes IM) and a S. chromogenes strain isolated from a heifer's teat apex (S. chromogenes TA). Each heifer was inoculated in the mammary gland with 1.0 x 10(6) colony forming units of each bacterial strain (one strain per udder quarter), whereas the remaining quarter was infused with phosphate-buffered saline. Overall, the CNS evoked a mild local host response. The somatic cell count increased in all S. fleurettii-inoculated quarters, although the strain was eliminated within 12 h. The two S. chromogenes strains were shed in larger numbers for a longer period. Bacterial and somatic cell counts, as well as neutrophil responses, were higher after inoculation with S. chromogenes IM than with S. chromogenes TA. In conclusion, these results suggest that S. chromogenes might be better adapted to the mammary gland than S. fleurettii. Furthermore, not all S. chromogenes strains induce the same local host response

Characteristics of Escherichia coli Isolated from bovine mastitis exposed to subminimum inhibitory concentrations of cefalotin or ceftazidime

Escherichia coli is a major udder pathogen causing clinical mastitis in dairy cattle and its heat stable endotoxin in powdered infant formula milk is a potential risk factor in neonatal infections. Cephalosporins are frequently used for treatment of mastitis caused by mastitis; however, use of these antimicrobials may induce antimicrobial resistance in E. colt. The objective of this study was to explore the in vitro effect of subminimum inhibitory concentrations (sub-MIC) of cefalotin (CF) and ceftazidime (CAZ) on the morphology, antimicrobial resistance, and endotoxin releasing characteristics of 3 E. colt isolates recovered from bovine clinical mastitis. The parent E. colt isolates, which were susceptible to CF and CAZ, were exposed to CF or CAZ separately at sub-MIC levels to produce 9 generations of induced isolates. Colonies of the CAZ-induced isolates from all 3 parent E. coli were smaller on blood agar and the bacteria became filamentous, whereas the CF-induced isolates did not demonstrate prominent morphological changes. After induction by CF or CAZ, many induced isolates showed resistance to cefoxitin, CAZ, CF, kanamycin, ampicillin, and amoxicillin/clavulanic acid while their parent isolates were susceptible to these antimicrobials. Notably, 5 CAZ-induced isolates from the same parent isolate were found to produce extended-spectrum beta-lactamase (ESBL) though none of the tested ESBL related genes could be detected. All CAZ-induced isolates released more endotoxin with a higher release rate, whereas endotoxin release of CF-induced E. colt isolates was not different from parent isolates. The exposure of cephalosporins at sub-MIC levels induced resistant Escherichia colt. We inferred that cephalosporins, especially CAZ, should be used prudently for treatment of clinical E. colt mastitis

Etiology and antimicrobial susceptibility of udder pathogens from cases of subclinical mastitis in dairy cows in Sweden

<p>Abstract</p> <p>Background</p> <p>A nationwide survey on the microbial etiology of cases of subclinical mastitis in dairy cows was carried out on dairy farms in Sweden. The aim was to investigate the microbial panorama and the occurrence of antimicrobial resistance. Moreover, differences between newly infected cows and chronically infected cows were investigated.</p> <p>Methods</p> <p>In total, 583 quarter milk samples were collected from 583 dairy cows at 226 dairy farms from February 2008 to February 2009. The quarter milk samples were bacteriological investigated and scored using the California Mastitis Test. Staphylococci were tested for betalactamase production and presence of resistance was evaluated in all specific udder pathogens. Differences between newly infected cows and chronically infected cows were statistically investigated using logistic regression analysis.</p> <p>Results</p> <p>The most common isolates of 590 bacteriological diagnoses were <it>Staphylococcus (S) aureus </it>(19%) and coagulase-negative staphylococci (CNS; 16%) followed by <it>Streptococcus (Str) dysgalactiae </it>(9%), <it>Str. uberis </it>(8%), <it>Escherichia (E.) coli </it>(2.9%), and <it>Streptococcus </it>spp. (1.9%). Samples with no growth or contamination constituted 22% and 18% of the diagnoses, respectively. The distribution of the most commonly isolated bacteria considering only bacteriological positive samples were: <it>S. aureus </it>- 31%, CNS - 27%, <it>Str. dysgalactiae </it>- 15%, <it>Str. uberis </it>- 14%, <it>E. coli </it>- 4.8%, and <it>Streptococcus </it>spp. - 3.1%. There was an increased risk of finding <it>S. aureus, Str. uberis </it>or <it>Str. dysgalactiae </it>in milk samples from chronically infected cows compared to findings in milk samples from newly infected cows. Four percent of the <it>S. aureus </it>isolates and 35% of the CNS isolates were resistant to penicillin G. Overall, resistance to other antimicrobials than penicillin G was uncommon.</p> <p>Conclusions</p> <p><it>Staphylococcus aureus </it>and CNS were the most frequently isolated pathogens and resistance to antimicrobials was rare.</p

Bacteriological etiology and treatment of mastitis in Finnish dairy herds

Background: The Finnish dairy herd recording system maintains production and health records of cows and herds. Veterinarians and farmers register veterinary treatments in the system. Milk samples for microbiological analysis are routinely taken from mastitic cows. The laboratory of the largest dairy company in Finland, Valio Ltd., analyzes most samples using real-time PCR. This study addressed pathogen-specific microbiological data and treatment and culling records, in combination with cow and herd characteristics, from the Finnish dairy herd recording system during 2010-2012. Results: The data derived from 240,067 quarter milk samples from 93,529 dairy cows with mastitis; 238,235 cows from the same herds served as the control group. No target pathogen DNA was detected in 12% of the samples. In 49% of the positive samples, only one target species and in 19%, two species with one dominant species were present. The most common species in the samples with a single species only were coagulase-negative staphylococci (CNS) (43%), followed by Staphylococcus aureus (21%), Streptococcus uberis (9%), Streptococcus dysgalactiae (8%), Corynebacterium bovis (7%), and Escherichia coli (5%). On average, 36% of the study cows and 6% of the control cows had recorded mastitis treatments during lactation. The corresponding proportions were 16 and 6% at drying-off. For more than 75% of the treatments during lactation, diagnosis was acute clinical mastitis. In the milk samples from cows with a recorded mastitis treatment during lactation, CNS and S. aureus were most common, followed by streptococci. Altogether, 48% of the cows were culled during the study. Mastitis was reported as the most common reason to cull; 49% of study cows and 18% of control cows were culled because of mastitis. Culling was most likely if S. aureus was detected in the milk sample submitted during the culling year. Conclusions: The PCR test has proven to be an applicable method also for large-scale use in bacterial diagnostics. In the present study, microbiological diagnosis was unequivocal in the great majority of samples where a single species or two species with one dominating were detected. Coagulase-negative staphylococci and S. aureus were the most common species. S. aureus was also the most common pathogen among the culled cows, which emphasizes the importance of preventive measures.Peer reviewe

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

Utility of Survival Motor Neuron ELISA for Spinal Muscular Atrophy Clinical and Preclinical Analyses

Genetic defects leading to the reduction of the survival motor neuron protein (SMN) are a causal factor for Spinal Muscular Atrophy (SMA). While there are a number of therapies under evaluation as potential treatments for SMA, there is a critical lack of a biomarker method for assessing efficacy of therapeutic interventions, particularly those targeting upregulation of SMN protein levels. Towards this end we have engaged in developing an immunoassay capable of accurately measuring SMN protein levels in blood, specifically in peripheral blood mononuclear cells (PBMCs), as a tool for validating SMN protein as a biomarker in SMA.A sandwich enzyme-linked immunosorbent assay (ELISA) was developed and validated for measuring SMN protein in human PBMCs and other cell lysates. Protocols for detection and extraction of SMN from transgenic SMA mouse tissues were also developed.The assay sensitivity for human SMN is 50 pg/mL. Initial analysis reveals that PBMCs yield enough SMN to analyze from blood volumes of less than 1 mL, and SMA Type I patients' PBMCs show ∼90% reduction of SMN protein compared to normal adults. The ELISA can reliably quantify SMN protein in human and mouse PBMCs and muscle, as well as brain, and spinal cord from a mouse model of severe SMA.This SMN ELISA assay enables the reliable, quantitative and rapid measurement of SMN in healthy human and SMA patient PBMCs, muscle and fibroblasts. SMN was also detected in several tissues in a mouse model of SMA, as well as in wildtype mouse tissues. This SMN ELISA has general translational applicability to both preclinical and clinical research efforts