Evaluation of antimicrobial resistance and risk factors for recovery of intrauterine Escherichia coli from cows with metritis on California commercial dairy farms.

The goals of this study were to evaluate factors affecting recovery and antimicrobial resistance (AMR) in intrauterine E. coli in post-partum dairy cows with and without metritis from commercial California dairy farms. Using a cross-sectional study design, a total of 307 cows were sampled from 25 farms throughout California, from which a total of 162 intrauterine E. coli isolates were recovered. During farm visits, cows within 21 days post-partum were categorized in one of three clinical presentation groups before enrollment: metritis (MET, n = 86), defined as a cow with watery, red or brown colored, and fetid vaginal discharge; cows with purulent discharge (PUS, n = 106), defined as a non-fetid purulent or mucopurulent vaginal discharge; and control cows, (CTL, n = 115) defined as cows with either no vaginal discharge or a clear, non-purulent mucus vaginal discharge. Cows diagnosed as MET had significantly higher odds for recovery of E. coli compared to cows diagnosed as CTL (OR = 2.16, 95% CI: 1.17-3.96), with no significant difference observed between PUS and CTL, and PUS and MET. An increase in days in milk (DIM) at the time of sampling was significantly associated with a decrease in the odds ratio for E. coli recovery from intrauterine swabs (OR = 0.94, 95% CI: 0.89-0.98). All intrauterine E. coli were resistant to ampicillin (AMP), with an AMR prevalence of 30.2% and 33.9% observed for chlortetracycline and oxytetracycline, respectively. Only 8.6% of isolates were resistant to ceftiofur (CEFT), one of the most common drugs used to treat cows on farms sampled. No significant difference in the prevalence of AMR was observed among clinical groups at the individual cow level. At the farm level, a significantly higher odds for isolating intrauterine E. coli resistant to chlortetracycline (OR: 2.6; 95% CI: 3.7-58.0) or oxytetracycline (OR: 1.9; 95% CI: 1.4-33.8) was observed at farms that used an intrauterine infusion of oxytetracycline as a treatment for metritis when compared to those farms that did not use this practice. Findings from this study indicate the need for further research supporting a broader understanding of farm practices driving AMR in cows with metritis, as well as data to increase the accuracy of breakpoints for AMR classification of intrauterine E. coli from cattle.

Correction: Risk factors affecting dairy cattle protective grouping behavior, commonly known as bunching, against Stomoxys calcitrans (L.) on California dairies.

[This corrects the article DOI: 10.1371/journal.pone.0224987.].

Risk factors affecting dairy cattle protective grouping behavior, commonly known as bunching, against Stomoxys calcitrans (L.) on California dairies.

Bunching is the term used to describe the protective aggregating behavior of cattle against the painful bites of stable flies (Stomoxys calcitrans), where cattle gather in a group with their heads to the center of the group and their tails to the outside to reduce stable fly attack. Both sexes of the stable fly feed on blood, and their painful bites negatively impact cattle health, productivity and welfare. A longitudinal study was conducted from April to July 2017 to estimate the stable fly activity on 20 California dairies (average herd size = 2466 ± 1050), to determine stable fly activity that induced bunching, and to evaluate the association between management and environmental factors, and cattle bunching. Stable fly activity was recorded weekly using trap counts and leg counts. Data was analyzed using linear mixed models with odds ratio. Cattle bunching at the dairy level was predicted by mean trap counts of ≥150 flies/trap/week, while bunching at the pen level was predicted by mean leg counts >1 fly/leg/cow or mean trap counts >50 flies/trap/week for traps closest to the pen. Additional risk factors predicting cattle bunching at the dairy level were study week (May/June vs July), presence of crops adjacent to dairy >2 sides, and feeding wet distillers grain. Additional risk factors predicting cattle bunching at the pen level were study week (May/June vs July), ambient temperature ≤30°C, pen design (freestall vs open dry lot or bedded pack), production status (lactating/dry vs close-up), presence of crops surrounding cattle pens, feeding rations containing molasses. Cattle bunching was reduced at the pen level by relative humidity >50%, and when the cattle pen was surrounded by other cattle pens or was bordered by a main road. At the dairy level, removal of manure along fence lines of cattle pens was protective against cattle bunching.

Effectiveness of zinc supplementation on diarrhea and average daily gain in pre-weaned dairy calves: A double-blind, block-randomized, placebo-controlled clinical trial.

The objective of this clinical trial was to evaluate the effectiveness of zinc supplementation on diarrhea and average daily weight gain (ADG) in pre-weaned dairy calves. A total of 1,482 healthy Holstein heifer and bull calves from a large California dairy were enrolled at 24 to 48 hours of age until hutch exit at approximately 90 days of age. Calves were block-randomized by time to one of three treatments: 1) placebo, 2) zinc methionine (ZM), or 3) zinc sulfate (ZS) administered in milk once daily for 14 days. Serum total protein at enrollment and body weight at birth, treatment end, and hutch exit were measured. Fecal consistency was assessed daily for 28 days post-enrollment. For a random sample of 127 calves, serum zinc concentrations before and after treatment and a fecal antigen ELISA at diarrhea start and resolution for Escherichia coli K99, rotavirus, coronavirus, and Cryptosporidium parvum were performed. Linear regression showed that ZM-treated bull calves had 22 g increased ADG compared to placebo-treated bulls (P = 0.042). ZM-treated heifers had 9 g decreased ADG compared to placebo-treated heifers (P = 0.037), after adjusting for average birth weight. Sex-stratified models showed that high birth weight heifers treated with ZM gained more than placebo-treated heifers of the same birth weight, which suggests a dose-response effect rather than a true sex-specific effect of ZM on ADG. Cox regression showed that ZM and ZS-treated calves had a 14.7% (P = 0.015) and 13.9% (P = 0.022) reduced hazard of diarrhea, respectively, compared to placebo-treated calves. Calves supplemented for at least the first five days of diarrhea with ZM and ZS had a 21.4% (P = 0.027) and 13.0% (P = 0.040) increased hazard of cure from diarrhea, respectively, compared to placebo-treated calves. Logistic regression showed that the odds of microbiological cure at diarrhea resolution for rotavirus, C. parvum, or any single fecal pathogen was not different between treatment groups. Zinc supplementation delayed diarrhea and expedited diarrhea recovery in pre-weaned calves. Additionally, zinc improved weight gain differentially in bulls compared to heifers, indicating a research need for sex-specific dosing.

Development of a clinical scoring system for bovine respiratory disease in weaned dairy calves.

Clinical scoring systems for bovine respiratory disease (BRD) in weaned dairy calves have been developed in the past with calves experimentally infected with specific respiratory pathogens. In this prevalent case control study, a BRD clinical scoring system for weaned calves was developed using field data from 689 dairy calves housed in group pens on 5 dairies in California. Of the 689 calves in the study, 89 were selected because they appeared sick based on the display of lethargy, depression, or separation from the group, whereas the remaining 600 were randomly selected. Clinical signs were recorded for all calves, and BRD case status was determined by thoracic auscultation and ultrasound examinations, which were interpreted in parallel. Of the 689 calves, 238 were identified as BRD cases. Five survey-adjusted generalized linear mixed models with a logit link function, calf as the unit of analysis, and dairy as a random intercept were assessed using 3-fold cross-validation. The best model chosen based on performance and parsimony contained the variables cough (2 points), abnormal respiration (1 point), low body condition (5 points), sunken eyes (4 points), and a 24-h ambient temperature range >15°C (1 point) with a 2-point cutoff for a BRD suspect score. An alternative model did not contain a score for the covariate 24-h ambient temperature range and had a 1-point cutoff. The best model was tested on 174 observations not used for model training and resulted in 77.0% screening sensitivity, 100% diagnostic sensitivity, and 61.9% specificity. Adding rectal temperature ≥39.2°C (102.5°F) as a second-tier test increased specificity to 76.7% and lowered the screening sensitivity to 64.8% and diagnostic sensitivity to 76.9%. The alternative model had a screening sensitivity of 84.2%, diagnostic sensitivity of 100%, and specificity of 45.7%. Adding rectal temperature ≥39.2°C (102.5°F) as a second-tier test for score-positive animals improved specificity of the alternative model to 62.6% while lowering its screening sensitivity to 70.5% and diagnostic sensitivity to 76.9%. Use of a 2-tier California BRD postweaning scoring system may provide producers and veterinarians with a new tool to monitor BRD in group-housed dairy calves. Furthermore, the scoring system may aid in judicious medical intervention for BRD cases and reduce unnecessary treatments of animals with antimicrobials.

Reliability of sampling strategies for measuring dairy cattle welfare on commercial farms.

Our objective was to evaluate how the proportion of high-producing lactating cows sampled on each farm and the selection method affect prevalence estimates for animal-based measures. We assessed the entire high-producing pen (days in milk <100; range = 81-241 cows) on 10 California farms using measures from the Welfare Quality Protocol for Cattle. Cows were restrained in head locks and visually evaluated for body condition, dirtiness, skin alterations (hair loss, lesions, or swelling), discharge (ocular, nasal, vulvar), diarrhea, and impaired respiration. Lameness was scored upon release. Prevalence was calculated as a percentage of assessed cows. The most common conditions were dirty hindquarters (33.5 ± 10.7%, mean ± standard deviation) and lesions or swelling on the carpal joint (34.4 ± 7.0%) and hock (26.4 ± 16.7%). Diarrhea (8.0 ± 5.8%), lameness (moderate = 7.3 ± 4.7%, severe = 2.2 ± 2.2%), and neck (5.8 ± 12.6%), flank (4.5 ± 5.0%), or hindquarter alterations (5.5 ± 3.9%) were less common. Very fat cows, vulvar discharge, and impaired respiration were rare (≤1%) and were excluded from further analysis. Four sampling strategies were used to generate 20 estimates for each animal-based measure. The strategies were (1) selecting every 10th, 5th, 4th, 3rd, 2nd, 2 of 3, or 3 of 4 cows at the feed bunk (7 estimates/measure); (2) randomly selecting 7 matching proportions of the pen; (3) randomly selecting cows using 3 sample size calculations from the Welfare Quality Protocol; and (4) selecting the first, middle, or final third of cows exiting the milking parlor. Estimates were compared with true values using regression analysis and were considered accurate if they met 3 criteria: the coefficient of determination was ≥0.9 and the slope and intercept did not differ significantly from 1 and 0, respectively. All estimates met the slope and intercept criteria, whereas the coefficient of determination increased when more cows were sampled. All estimates were accurate for neck alterations, ocular discharge (22.2 ± 27.4%), and carpal joint hair loss (14.1 ± 17.4%). Selecting a third of the milking order or using the Welfare Quality sample size calculations failed to accurately estimate all measures simultaneously. However, all estimates were accurate when selecting at least 2 of every 3 cows locked at the feed bunk. Using restraint position at the feed bunk did not differ systematically from computer-selecting the same proportion of cows randomly, and the former may be a simpler approach for welfare assessments.

Epidemiology of Salmonella sp. in California cull dairy cattle: prevalence of fecal shedding and diagnostic accuracy of pooled enriched broth culture of fecal samples.

BACKGROUND: The primary objective of this cross-sectional study was to estimate the crude, seasonal and cull-reason stratified prevalence of Salmonella fecal shedding in cull dairy cattle on seven California dairies. A secondary objective was to estimate and compare the relative sensitivity (Se) and specificity (Sp) for pools of 5 and 10 enriched broth cultures of fecal samples for Salmonella sp. detection. METHODS: Seven dairy farms located in the San Joaquin Valley of California were identified and enrolled in the study as a convenience sample. Cull cows were identified for fecal sampling once during each season between 2014 and 2015, specifically during spring, summer, fall, and winter, and 10 cows were randomly selected for fecal sampling at the day of their sale. In addition, study personnel completed a survey based on responses of the herd manager to questions related to the previous four month's herd management. Fecal samples were frozen until testing for Salmonella. After overnight enrichment in liquid broth, pools of enrichment broth (EBP) were created for 5 and 10 samples. All individual and pooled broths were cultured on selective media with putative Salmonella colonies confirmed by biochemical testing before being serogrouped and serotyped. RESULTS: A total of 249 cull cows were enrolled into the study and their fecal samples tested for Salmonella. The survey-weighted period prevalence of fecal shedding of all Salmonella sp. in the cull cow samples across all study herds and the entire study period was 3.42% (N = 249; SE 1.07). The within herd prevalence of Salmonella shed in feces did not differ over the four study seasons (P = 0.074). The Se of culture of EBP of five samples was 62.5% (SE = 17.12), which was not statistically different from the Se of culture of EBP of 10 (37.5%, SE = 17.12, P = 0.48). The Sp of culture of EBP of five samples was 95.24% (SE = 3.29) and for pools of 10 samples was 100.00% (SE = 0). There was no statistical difference between the culture relative specificities of EBP of 5 and 10 (P > 0.99). DISCUSSION: Our study showed a numerically higher prevalence of Salmonella shedding in the summer, although the results were not significant, most likely due to a lack of power from the small sample size. A higher prevalence in summer months may be related to heat stress. To detect Salmonella, investigators may expect a 62.5% sensitivity for culture of EBP of five, relative to individual fecal sample enrichment and culture. In contrast, culture of EBP of 10 samples resulted in a numerically lower Se. Culture of EBP of size 5 or 10 samples, given similar prevalence and limit of detection, can be expected to yield specificities of 95 and 100%, respectively.

Cost-effectiveness of diagnostic strategies using quantitative real-time PCR and bacterial culture to identify contagious mastitis cases in large dairy herds.

Diagnostic strategies to detect contagious mastitis caused by Mycoplasma bovis, Staphylococcus aureus, and Streptococcus agalactiae in dairy herds during an outbreak have been minimally studied with regard to cost and diagnostic sensitivity. The objective of this cross-sectional study was to compare the cost-effectiveness of diagnostic strategies for identification of infected cows in two California dairy herds during contagious mastitis outbreaks. M. bovis was investigated in a subset of a herd (n=1210 cows) with an estimated prevalence of 2.8% (95% CI=1.9, 3.7), whereas Staph. aureus and Strep. agalactiae were studied in a second herd (n=351 cows) with an estimated prevalence of 3.4% (95% CI=1.5, 5.3) and 16.8% (95% CI=12.9, 20.7), respectively. Diagnostic strategies involved a combination of testing stages that utilized bacterial culture, quantitative real-time PCR (qPCR), or both. Strategies were applied to individual or pooled samples of 5, 10, 50 or 100 samples. Culture was considered the gold standard for sensitivity estimation of each strategy. The reference strategy was the strategy with the lowest cost per culture-positive cow which for both M. bovis and Strep. agalactiae consisted of 2 stages, culture of samples in pools of 5 followed by culture of individual samples in positive pools with a sensitivity of 73.5% (95% CI: 55.6, 87.1) and 96.6% (95% CI: 27.7, 84.8), respectively. The reference strategy for Staph. aureus consisted of 3 stages, culture of individual samples in pools of 100 (stage 1), culture constituents of those positive from stage 1 in pools of 5 (stage 2), culture constituents of those positive from stage 2 individually (stage 3) which resulted in a sensitivity of 58.3% (95% CI: 88.3, 99.6). The most cost-effective alternative to the reference strategy was whole herd milk culture for all 3 pathogens. QPCR testing was a component of the second most cost-effective alternative for M. bovis and the third most cost-effective alternatives for the 3 pathogens. A stochastic model was used to assess the effect of prevalence or herd size on the cost-effectiveness of diagnostic strategies. In the current study, increasing the prevalence of mastitis did not alter the ranking of strategies by cost-effectiveness. However, larger herds could benefit from testing larger pools such as 50 or 100 samples to improve cost-effectiveness. Several diagnostic strategy options exist to identify contagious mastitis in herds, decisions should be based on cost and sensitivity of the strategies available.

A randomized controlled trial on preweaning morbidity, growth and mortality in Holstein heifers fed a lacteal-derived colostrum replacer or pooled maternal colostrum.

BACKGROUND: The objective of this randomized controlled trial was to determine the effect of feeding a commercial lacteal-derived colostrum replacer (CR) or pooled maternal colostrum (MC) on preweaning morbidity, growth and mortality in Holstein heifer calves. A total of 568 calves were randomly assigned to be fed either 3.8 L of pooled MC or two doses (200 g IgG) of a CR. Calves were monitored daily for preweaning morbidity until weaning at 60 d old. Birth and weaning weights were measured to estimate growth rates. RESULTS: Calves fed CR were significantly less likely to be affected with a diarrhea event (OR = 0.58; 95% CI, 0.38 to 0.88; P value = 0.011) and had a higher rate of daily weight gain (0.051 kg/day; 95% CI, 0.03 to 0.08; P value <0.001) compared to calves fed pooled MC. Use of lacteal-derived colostrum replacer was not significantly associated with respiratory disease (OR = 1.01; 95% CI 0.67 to 1.51; P value = 0.974 ), omphalitis (OR = 0.93; 95% CI 0.06 to 14.86; P value = 0.956), or mortality (HR = 0.71; 95% CI 0.27 to 1.92; P value = 0.505) in the study calves. CONCLUSIONS: The lacteal-derived CR fed at the study dose was a viable colostrum alternative in the event of poor quality pooled MC for the prevention of preweaning diarrhea and resulted in higher growth rates in comparison to calves fed pooled MC in the study herd.

Efficacy of feeding a lacteal-derived colostrum replacer or pooled maternal colostrum with a low IgG concentration for prevention of failure of passive transfer in dairy calves.

OBJECTIVE: To compare the efficacy of a lacteal-derived colostrum replacer (LDCR) for the prevention of failure of passive transfer of immunity (FPT) in calves with that of pooled maternal colostrum (MC). DESIGN: Randomized field trial. ANIMALS: 568 heifer calves from 1 California dairy. PROCEDURES: Calves were randomly allocated to 1 of 2 treatment groups and fed 2 doses (200 g of IgG) of an LDCR or 3.8 L of pooled MC. From each calf, blood samples were collected before and approximately 24 hours after treatment. Serum IgG and total protein (TP) concentrations were quantified with standard methods, and the apparent efficiency of IgG absorption was calculated. RESULTS: At 24 hours after treatment, mean serum TP and IgG concentrations were significantly lower for calves fed pooled MC (TP, 4.77 g/dL; IgG, 7.50 g/L), compared with those for calves fed the LDCR (TP, 5.50 g/dL; IgG, 15.15 g/L). Calves fed the LDCR were 95% less likely to develop FPT (OR, 0.05; 95% confidence interval, 0.03 to 0.08) than were calves fed pooled MC. However, the mean IgG concentration in the pooled MC fed during the study (21.1 g/L) was substantially lower than that (64.3 g/L) determined for representative samples of pooled MC from other southwestern US dairies during a national survey. CONCLUSIONS AND CLINICAL RELEVANCE: Results indicated that, on this particular dairy, calves fed an LDCR were at less risk of developing FPT than were calves fed pooled MC. The LDCR evaluated was a viable alternative for the prevention of FPT in calves.