Antimicrobial resistance and prevalence of virulence factor genes in fecal Escherichia coli of Holstein calves fed milk with and without antimicrobials

Diarrhea in calves has a significant effect on the dairy industry. A common management practice for preventing or decreasing the effects of such disease in preweaned calves is by the use of antimicrobials in milk or milk replacer. In this study, Escherichia coli antimicrobial resistance in fecal samples collected from calves 2 to 8 d of age that had or had not received antimicrobials in the milk and that had or had not signs of diarrhea by inspection of fecal consistency were investigated. Specifically, resistance of E. coli isolates to individual antimicrobials, multiresistance patterns, and presence of virulence factors were analyzed. Escherichia coli isolates were tested for susceptibility to 12 antimicrobials by use of a Kirby-Bauer disk diffusion assay. The study was conducted at 3 farms, 1 administering growth-promoting antimicrobials (GPA) in the milk and 2 not using GPA in the milk (NGPA). All isolates were susceptible to ciprofloxacin and cefepime. From the total isolates tested, 84% (n = 251) were resistant to at least 2 antimicrobials and 81% (n = 251) were resistant to 3 or more antimicrobials. When antimicrobial resistance was compared between GPA and NGPA, it was observed that the GPA group had higher odds of antimicrobial resistance for most of the individual antimicrobials tested. No significant correlation of virulence factors in GPA or NGPA and diarrheic or non-diarrheic (control) fecal samples was found. Of the 32 virulence factors evaluated, 21 were detected in the study population; the incidence of only 1 virulence factor was statistically significant in each of the diarrheic status (diarrheic or non-diarrheic) and treatment status (NGPA or GPA) groups. Phylogenetic analysis based on the nucleotide sequence of the DNA gyrase gene (gyrB) from 31 fecal E. coli isolates revealed 3 main clades.     

Exposure to antimicrobials through the milk diet or systemic therapy is associated with a transient increase in antimicrobial resistance in fecal Escherichia coli of dairy calves

The objective of this prospective cohort study was to describe the relationship between exposure to antimicrobials, through both the milk diet and systemic therapy, and to describe antimicrobial resistance of fecal Escherichia coli in dairy calves pre- and postweaning. A convenience sample of 15 Minnesota dairy farms was chosen, representing 3 equal cohorts of milk diet fed to preweaned calves: medicated milk replacer (MMR), nonmedicated milk replacer (NMR), or pasteurized nonsaleable milk (PNM). Five newborn calves were enrolled on each farm, with fecal samples collected from each calf at 1, 3, 5, and 16 wk of age. After isolation, 3 colonies of E. coli were randomly selected from each sample to determine antimicrobial susceptibility by minimum inhibitory concentration (Sensititer, Thermo Scientific, Waltham, MA) to 8 antimicrobials in 8 classes. The isolate was given an antimicrobial resistance score (ARS) according to the number of antimicrobial classes to which it was resistant. Any isolate resistant to 3 or more antimicrobials was defined as being multidrug resistant (MDR). Relationships between ARS and MDR (dependent variables) and possible explanatory variables were analyzed using mixed multivariable linear and logistic regression models, respectively, with critical P-values adjusted for multiple contrasts. Seventy percent of isolates were resistant to sulfadimethoxine. For wk 1 and 3, the mean ARS values were greatest for fecal E. coli from calves fed MMR or PNM compared with NMR, with no difference in ARS values between the MMR and PNM groups at either time point. At wk 5, the mean ARS value was greatest for fecal E. coli from calves fed MMR (3.56 ± 0.45; mean ± SE), intermediate for calves fed PNM (2.64 ± 0.45), and lowest for calves fed NMR (1.54 ± 0.45). However, by wk 16, the mean ARS values were ≤1.0 and did not differ among milk diets. Evaluation of the proportion of isolates with MDR mirrored the results of the ARS analysis (MDR more prevalent in MMR and PNM groups preweaning; no difference among milk diets at 16 wk). There was a tendency for an increase in ARS at wk 5 (1.28 ± 0.70), and the odds for MDR in fecal E. coli were estimated to be 5.2 (95% confidence interval = 0.67, 35.7) and 101.1 (95% confidence interval = 1.15, >999.9) higher at wk 3 and 5 if the calf was treated with a systemic antimicrobial within the 14-d period before sampling. These findings suggest that exposure to antimicrobials through the milk diet or systemic therapy may result in a transient increase in resistance in fecal E. coli, but once the antimicrobial pressure is removed, susceptible E. coli are able to flourish again, resulting in an overall decrease in resistance.     

Feeding of waste milk to Holstein calves affects antimicrobial resistance of Escherichia coli and Pasteurella multocida isolated from fecal and nasal swabs

The use of milk containing antimicrobial residues in calf feeding programs has been shown to select for resistant fecal Escherichia coli in dairy calves. However, information is scarce about the effects of feeding calves waste milk (WM) on the prevalence of multidrug-resistant bacteria. The objective of this study was to determine the antimicrobial resistance patterns of fecal E. coli and nasal Pasteurella multocida isolates from calves fed either milk replacer (MR) or WM in 8 commercial dairy farms (4 farms per feeding program). Fecal and nasal swabs were collected from 20 ± 5 dairy calves at 42 ± 3.2 d of age, and from 10 of these at approximately 1 yr of age in each study farm to isolate the targeted bacteria. Furthermore, resistance of E. coli isolates from calf-environment and from 5 calves at birth and their dams was also evaluated in each study farm. Resistances were tested against the following antimicrobial agents: amoxicillin-clavulanic acid, ceftiofur, colistin, doxycycline (DO), enrofloxacin (ENR), erythromycin, florfenicol, imipenem, and streptomycin. A greater number of fecal E. coli resistant to ENR, florfenicol, and streptomycin and more multidrug-resistant E. coli phenotypes were isolated in feces of calves fed WM than in those fed MR. However, the prevalence of fecal-resistant E. coli was also influenced by calf age, as it increased from birth to 6 wk of age for ENR and DO and decreased from 6 wk to 1 yr of age for DO regardless of the feeding program. From nasal samples, an increase in the prevalence of colistin-resistant P. multocida was observed in calves fed WM compared with those fed MR. The resistance patterns of E. coli isolates from calves and their dams tended to differ, whereas similar resistance profiles among E. coli isolates from farm environment and calves were observed. The findings of this study suggest that feeding calves WM fosters the presence of resistant bacteria in the lower gut and respiratory tracts of dairy calves.     

Effect of on-farm use of antimicrobial drugs on resistance in fecal Escherichia coli of preweaned dairy calves

Respiratory disease and diarrhea are the 2 most common diseases that result in the use of antimicrobial drugs in preweaned calves. Because the use of drugs in food animals, including dairy calves, has the potential for generating cross-resistance to drugs used in human medicine, it is vital to propose farm practices that foster the judicious use of antimicrobials while assuring animal health and productivity. The objective of this study was to use dairy farm calf treatment records to identify antimicrobial drug treatments in calves and to evaluate their effects on the prevalence of antimicrobial-resistant Escherichia coli from rectal swabs of preweaned dairy calves. Eight farms from central New York participated in the study, 3 farms using individual pen housing management and 5 farms using group pen housing management. Eligible study farms could not add antimicrobial drugs to the milk fed to preweaned calves and were required to have farm records documenting antimicrobial drug treatment of calves from birth to weaning. Three fecal E. coli isolates per calf were tested for susceptibility to 12 antimicrobial drugs using a Kirby-Bauer disk diffusion assay. A total of 473 calves were sampled, from which 1,423 commensal E. coli isolates were tested. Of the 9 antimicrobial drugs used on study farms, only enrofloxacin was significantly associated with reduced antimicrobial susceptibility of E. coli isolates, although treatment with ceftiofur was associated with reduced susceptibility to ceftriaxone. The median numbers of days from treatment with ceftiofur and enrofloxacin to rectal swab sampling of calves were 16 d (range: 1–39) and 12 d (range: 6–44), respectively. At the isolate level, treatment with enrofloxacin resulted in odds ratios of 2 [95% confidence interval (CI): 1–4] and 3 (95% CI: 2–6), respectively, for isolation of nonsusceptible E. coli to nalidixic acid and ciprofloxacin compared with calves not treated with enrofloxacin. Treatment with ceftiofur resulted in an odds ratio of 3 (95% CI: 0.9–12) for isolation of nonsusceptible E. coli to ceftriaxone compared with calves not treated with ceftiofur. Treatment with enrofloxacin resulted in selection of isolates that presented phenotypic resistance to both ciprofloxacin and ceftriaxone. Treatment with ceftiofur resulted in a higher prevalence of isolates resistant to ≥3 antimicrobial drugs (97%) compared with no treatment with ceftiofur (73%). These findings reinforce the necessity for continued implementation of practices at the dairy farm that support the sustainable and judicious use of antimicrobial drugs in dairy calves.     

Effect of preweaned dairy calf housing system on antimicrobial resistance in commensal Escherichia coli

Group housing of preweaned dairy calves is a growing practice in the United States. The objective of this practice is to increase the average daily gain of calves in a healthy and humane environment while reducing labor requirements. However, feeding protocols, commingling of calves, and occurrence of disease in different calf-housing systems may affect the prevalence of antimicrobial drug-resistant bacteria. This study evaluated the effect of a group pen-housing system and individual pen-housing system on antimicrobial resistance trends in fecal Escherichia coli of preweaned dairy calves and on the prevalence of environmental Salmonella. Twelve farms from central New York participated in the study: 6 farms using an individual pen-housing system (IP), and 6 farms using a group pen-housing system (GP). A maximum of 3 fecal E. coli isolates per calf was tested for susceptibility to 12 antimicrobial drugs using a Kirby-Bauer disk diffusion assay. Calves in GP had a significantly higher proportion of E. coli resistant to ciprofloxacin and nalidixic acid, whereas calves in IP had a significantly higher proportion of E. coli resistant to ampicillin, ceftiofur, gentamycin, streptomycin, and tetracycline. Calf-housing system had an effect on resistance to individual antimicrobial drugs in E. coli, but no clear-cut advantage to either system was noted with regard to overall resistance frequency. No outstanding difference in the richness and diversity of resistant phenotypes was observed between the 2 calf-housing systems.     

Multidrug residues and antimicrobial resistance patterns in waste milk from dairy farms in Central California

Waste milk (WM) is a common source of feed for preweaned calves in US dairy farms. However, limited information is available about characteristics of this product, including concentration of drug residues and potential hazards from antibiotic-resistant bacteria present in the milk. The aims of this cross-sectional study were to (1) identify and measure the concentration of antimicrobial residues in raw WM samples on dairy farms in the Central Valley of California, (2) survey farm management practices for factors associated with the occurrence of specific antimicrobial residues in raw WM, (3) characterize the antimicrobial resistance patterns of E. coli cultured from raw WM samples, and (4) evaluate the potential association between WM quality parameter and risk of identifying drug residues in milk. A single raw bulk tank WM sample was collected from dairy farms located in California's Central Valley (n = 25). A questionnaire was used to collect information about farm management practices. Waste milk samples were analyzed for a multidrug residue panel using liquid chromatography–tandem mass spectrometry. Bacteria were cultured and antimicrobial resistance was tested using standard techniques; milk quality parameters (fat, protein, lactose, solids-not-fat, somatic cell count, coliform count, and standard plate count) were also measured. Of the 25 samples collected, 15 (60%) contained detectable concentrations of at least 1 antimicrobial. Of the drug residue–positive samples, 44% (11/25) and 16% (4/25) had detectable concentrations of β-lactams and tetracycline, respectively. The most prevalent drug residues were ceftiofur (n = 7, 28%), oxytetracycline (n = 4, 16%), and cephapirin (n = 3, 12%). No significant associations were identified between farm characteristics or management practices and presence of drug residues in WM. In this study, 20% of farms did not pasteurize WM before feeding to calves. Two of the 10 Escherichia coli isolated from WM samples were multidrug resistant. Streptococcus spp. (n = 21, 84%) was the most common genus cultured from WM samples, followed by Staphylococcus spp. (n = 20, 80%) and E.coli (n = 10, 40%). Mycoplasma spp. was cultured from 2 WM samples (n = 2, 8%). The presence of drug residues in WM at concentrations that increase selection of resistant bacteria indicates the need for additional studies targeting on-farm milk treatments to degrade drug residues before feeding to calves. The presence of multidrug-resistant E. coli in WM urges the need for on-farm practices that reduce calf exposure to resistant bacteria, such as pasteurization.     

Risk assessment modelling of fecal shedding caused by extended-spectrum cephalosporin-resistant Escherichia coli transmitted through waste milk fed to dairy pre-weaned calves

Waste milk feeding is a common practice in dairy operations. Regardless of the benefits of this practice to the dairy farmers, concerns from the potential dissemination of antimicrobial-resistant bacteria through the gut and subsequent shedding by calves into the environment are increasing. In this study, we employed Monte Carlo simulation to assess the risk of shedding extended-spectrum cephalosporin-resistant Escherichia coli (ESC-R E. coli) caused by waste milk feeding in pre-weaned calves using an exponential dose-response model fit to data for E. coli O157:H7 in cattle. Data from pertinent studies were included in our model to predict the risk of shedding. The median (5th and 95th percentiles) for the daily risk of shedding ESC-R E. coli by calves fed only contaminated waste milk was predicted to be 2.9 × 10^?3 (2.1 × 10^?3, 3.7 × 10^?3), representing a median daily risk of 29 out of 10,000 calves shedding ESC-R E. coli due to exclusive feeding of waste milk containing ESC-R E. coli. This median value was reduced by 94% when accounting for the proportion of waste milk that does not contain ESC-R E. coli. The overall risk of shedding ESC-R E. coli through the pre-weaning period for farms that feed waste milk to calves was 5.7 × 10^?3 (2.4 × 10^?3, 1.1 × 10^?2), representing 57 out of 10,000 calves. When accounting for the proportion of farms that do not feed waste milk, the pre-weaning period risk was reduced by 23%. By varying the prevalence of ESC-R E. coli in waste milk using values of 3, 1.5, and 1%, the daily risk of shedding decreased by factors of 50, 65, and 82%, respectively, which supports the reduction of contamination or discontinuation of feeding waste milk containing ESC-R E. coli as major mitigation measures to reduce the risk of shedding caused by ingestion of resistant bacteria. It is anticipated that the effects of antimicrobial residues in waste milk, which was not considered herein due to lack of data, would further increase risks. Although waste milk feeding to calves may be economically beneficial to the dairy farmers, there exists the risk of dissemination of ESC-resistant bacteria into the environment.     

Short communication: Selection of extended-spectrum β-lactamase-producing Escherichia coli in dairy calves associated with antibiotic dry cow therapy—A cohort study

Antimicrobial residues in milk have been discussed as a possible selector for Enterobacteriaceae that produce extended-spectrum β-lactamases (ESBL) in dairy herds. Such residues are found in waste milk after antibiotic treatment of mastitis, but antibiotic dry cow therapy might also lead to antibiotic residues in colostrum and in milk during early lactation. While it is known that feeding of waste milk selects ESBL bacteria in calves, this was not investigated for colostrum yet, which is supposed to contain much lower antibiotic concentrations than waste milk. In this observational prospective case study on 2 farms, we hypothesized that blanket dry cow treatment with β-lactams would have more selective (here: increasing) effects on ESBL concentrations than selective (here: individually chosen) antibiotic dry cow therapy. Thus, we compared concentrations of ESBL-producing Enterobacteriaceae in feces of calves (n = 50) at 2 dairy farms with different management of antibiotic dry cow therapy. Considerably higher concentrations of ESBL-producing Escherichia coli were observed in blanket antibiotic dry cow therapy on d 3 of the calf's life (7.6 vs. 5.3 log cfu/g of calf feces). Both farms used narrow-spectrum penicillin combined with aminoglycosides for drying off, and the majority of ESBL isolates (93%) were co-resistant to aminoglycosides. No waste milk was fed to calves and no calf was treated with β-lactam antibiotics or aminoglycosides during the first 3 d of life, thus differences were most likely associated with different frequency of antibiotic dry cow therapy on farms (19 of 25 mother cows on farm A, 9 of 25 on farm B). Even though the presumable selection effect of antibiotics used for drying off decreased within the next 3 wk, this result further emphasizes the need for the reduction and prudent use of antibiotic dry cow therapy on farms.     

Baicalin inhibits Escherichia coli isolates in bovine mastitic milk and reduces antimicrobial resistance

In this study, we aimed to evaluate the inhibitory effect of baicalin on Escherichia coli in vitro and the effects of baicalin treatment on antimicrobial resistance of the E. coli isolates. Through isolation, purification, and identification, a total of 56 E. coli strains were isolated from 341 mastitic milk samples. The study of inhibition effect of baicalin on the E. coli strains in vitro was focused on permeability and morphology of the isolates using an alkaline phosphatase kit and scanning electron microscopy. Furthermore, the resistance spectrum of the isolates to the common antimicrobial agents was tested at sub-minimum inhibitory concentrations of baicalin by the agar dilution method. Extended-spectrum β-lactamase and plasmid-mediated quinolone resistance genes were amplified by PCR before and after incubation with baicalin. The results revealed that baicalin has certain inhibitory effects on the isolates in vitro. The alkaline phosphatase enzyme activity was significantly increased from 1.246 to 2.377 U/100 mL, and the surface of E. coli was concave and shriveled. Analysis of the resistance spectrum and PCR amplification showed that, after administration with baicalin, the sensitivity of most strains to the selected antimicrobial agents was enhanced. Strikingly, the drug-resistant genes from 71.43% (40/56) of these isolates were found to have drug-resistant genes to different extents. Altogether, the current study confirmed both the inhibitory effect on Escherichia coli in vitro and the reduction of antimicrobial resistance by baicalin. This is the first comprehensive study to report on baicalin, a traditional Chinese medicine that acts on E. coli isolated from the mastitic milk samples.     

Presence of Bacillus species in pasteurised milk and their phenotypic and genotypic antimicrobial resistance profile

High-temperature short-time (HTST) pasteurisation is not sufficient for complete elimination of Bacillus sp. in milk. Hence, it is important to assess their prevalence, phenotypic and genetic antimicrobial resistance profiles in pasteurised milk. In this study, we investigated Bacillus prevalence and their phenotypic resistance to 25 antimicrobials and prevalence of five plasmid antimicrobial resistance genes. All isolates were resistant to β-lactams; most isolates were susceptible to chloramphenicol (n = 52), ciprofloxacin, meropenem, sulphonamides, tetracycline and vancomycin (n = 56). Intriguingly, despite phenotypic sulphonamide and tetracycline sensitivity, Sul2 and tetA genes were detected in different Bacillus species, signifying a potential risk of horizontal gene transfer of these mobile resistance genes through pasteurised milk.     

The influence of mulberry leaf flavonoids and Candida tropicalis on antioxidant function and gastrointestinal development of preweaning calves challenged with Escherichia coli O141:K99

This study investigated the effects of mulberry (Morus alba L.) leaf flavonoids and Candida tropicalis on the growth, gastrointestinal development, enzyme activity of gastrointestinal digesta, and blood antioxidant function of dairy calves challenged with Escherichia coli O141:K99. A total of 15 Holstein preweaning bull calves (age = 28 d; body weight = 51 ± 3.9 kg, mean ± standard deviation) were randomly divided into 3 homogeneous groups with no supplementation (control, CON), Candida tropicalis added at 1 g/calf per day (CT), and mulberry leaf flavonoids added at 3 g/calf per day (MLF). The feed efficiency was greater in calves fed MLF than in CON either before or after the E. coli challenge test. The fecal score increased after E. coli challenge, but it was greater in the CON group than in the CT treatment and in the MLF group at d 1, 2, and 3 after the E. coli challenge. Calves fed the CT diet showed decreased blood β-hydroxybutyrate concentrations compared with those on the CON and MLF diets before the E. coli challenge. Dietary supplementation with CT resulted in lower blood superoxide dismutase activity than the CON after E. coli challenge. The calves fed CT and MLF showed much lower blood glutathione peroxidase activity than the CON group after being challenged with E. coli. The tunica mucosa of the duodenum was lower in CT and MLF than in CON. The lactase activity of the jejunal digesta was greater in MLF than in CON and CT. In summary, MLF and CT supplementation might effectively improve antioxidant function and reduce the incidence of oxidative stress after challenge with E. coli in 28-d-old preweaning calves, and this effect may be partially explained by the relative decreases in blood superoxide dismutase and glutathione peroxidase as well as fecal score.     

Short communication: Extended-spectrum cephalosporin-resistant Escherichia coli in colostrum from New Brunswick,Canada, dairy cows harbor blaCMY-2 and blaTEM resistance genes

Dairy calves are colonized shortly after birth by multidrug resistant (MDR) bacteria, including Escherichia coli. The role of dairy colostrum fed to calves as a potential source of MDR bacteria resistance genes has not been investigated. This study determined the recovery rate of extended-spectrum cephalosporin-resistant (ESC-R) E. coli in colostrum from cows. The ESC-R E. coli isolates were further investigated to determine their phenotypic antimicrobial resistance pattern and the genes conferring ESC-R. Fresh colostrum was collected from 452 cows from 8 dairy herds in New Brunswick, Canada. The ESC-R E. coli was isolated from the colostrum by using the VACC agar, a selective media for extended-spectrum β-lactamase producing Enterobacteriaceae. Minimum inhibitory concentration was determined for all the suspected ESC-R E. coli isolates using a commercial gram-negative broth microdilution method. Two multiplex PCR were conducted on all the suspected ESC-R E. coli isolates to determine the presence of the bla_CTX-M (groups 1, 2, 9, and 8/25) bla_CMY-2, bla_SHV, and bla_TEM resistance genes. The ESC-R E. coli were detected in 20 (4.43%) of the colostrum samples. At least 1 ESC-R E. coli isolate was detected in 6 (75%) of the dairy herds. All ESC-R E. coli had MDR profiles based on minimum inhibitory concentration testing. No bla_CTX-M groups genes were detected; however, the bla_CMY-2 gene was detected in 9 or 20 (45%) and bla_TEM was detected in 7 of 20 (35%) of the ESC-R E. coli. No ESC-R E. coli had both bla_CMY-2 and bla_TEM resistance genes. This is the first report of bla_CMY-2 and bla_TEM genes found in E. coli isolates cultured from dairy colostrum to our knowledge.     

Feeding an acetate-based oral electrolyte reduces the ex vivo Escherichia coli growth potential in the abomasum of calves fed oral electrolytes alone or 30 minutes following a milk feeding compared with feeding a bicarbonate-based oral electrolyte

Oral electrolyte solutions (OES) are a common, on-farm therapy to reestablish hydration and electrolyte balances in scouring and stressed calves. The objectives were to determine the effects of OES alkalinizing agent and the presence of a milk replacer feeding before OES administration on the abomasal environment in healthy Holstein calves. Abomasum cannulation was performed on 16 Holstein bull calves at 5 d of age. One calf was removed from the study before the calves were randomly assigned to treatments at 9 d of age. Treatments were arranged as a 2-by-2 factorial, with the following factors: oral electrolyte alkalinizing agent [acetate (A) or bicarbonate (B)] and liquid meal type milk replacer (MR) + OES (MR-A, MR-B), or OES only (OES-A, OES-B)]. The OES differed only by alkalinizing agent. On d 9, calves assigned to MR-A (n = 4) or MR-B (n = 4) received their morning MR aliquot 0.5 h before feeding 2 L of OES; the OES-A (n = 3) and OES-B (n = 4) treatment groups were fed 2 L of OES only. Peripheral blood samples and postprandial abomasal fluid samples were collected to assess abomasal pH, abomasal emptying rate (AER), and ex vivo abomasal Escherichia coli growth potential. Postprandial pH was greater in calves fed MR or B-based OES. Abomasal emptying rate was slower in calves receiving MR + OES, regardless of the alkalinizing agent. Ex vivo E. coli colony-forming unit counts were greater in calves fed either MR + OES or bicarbonate-based OES. Supplementing bicarbonate OES in addition to MR alters abomasal dynamics and may promote E. coli growth in postprandial abomasal fluid, partially due to sustained elevations in gastric pH and delayed gastric emptying rates. The OES containing sodium acetate limited ex vivo E. coli growth potential in abomasal fluid, thereby potentially reducing the risk of additional enteric bacterial complications associated with OES therapy.     

Targeting therapy to minimize antimicrobial use in preweaned calves: Effects on health, growth, and treatment costs

Prophylactic and therapeutic antimicrobial use in food animals is questioned because of the potential for development of resistant bacteria and future inability to use some antimicrobials for human or animal disease. The objectives of this study were to determine the effect of raising preweaned dairy calves without antimicrobials in the milk and minimizing therapeutic antimicrobial treatment on morbidity, mortality, weight gain, and treatment costs. Newborn calves (n = 358) were allocated to 1 of 4 groups, housed outdoors in individual hutches, and monitored for 28 d. Calves in the conventional therapy (CT) group were treated as per dairy protocol with sulfamethoxazole/trimethoprim, spectinomycin, penicillin, and bismuth-pectin for diarrhea. The targeted therapy (TT) group included bismuth-pectin for diarrhea and antimicrobial treatment only in cases of fever or depressed attitude. Within CT and TT groups, calves were equally assigned to receive neomycin and tetracycline in their milk for the first 2 wk of life (AB-milk) or no antimicrobials (NoAB-milk). Daily health evaluations included fecal consistency, respiratory disease, attitude, and hydration status as well as milk and grain consumption. A negative binomial model evaluated the total number of days with diarrhea days in each group. General linear models were used to assess average daily weight gain and grain consumption. Conventionally treated calves had 70% more days with diarrhea than TT calves, and AB-milk calves had 31% more days with diarrhea compared with NoAB-milk calves. The TT calves tended to have a higher average daily gain by 28 d and consumed more grain compared with CT calves. If antimicrobials were used only for diarrhea cases with fever, inappetence, or depression and no in-milk antimicrobials were used, a $10 per calf savings could be realized. Targeting antimicrobial therapy of calf diarrhea cases is prudent not only to save the drugs for future use but also to prevent the potential for antibiotic-associated diarrhea and reduce calf-rearing costs.     

Effect of passive antibodies derived from rotavirus-like particles on neonatal calf diarrhea caused by rotavirus in an oral challenge model

Our objective was to evaluate the efficacy of bovine rotavirus antigen-specific passive antibody for reducing the duration of diarrhea induced by oral challenge with bovine rotavirus in a neonatal calf model. The bovine rotavirus-specific passive antibodies were produced before the study by hyperimmunization of pregnant cows during the dry period with an adjuvanted vaccine containing recombinantly-expressed rotavirus virus-like particles. Eighty-three calves were cleanly collected at birth and randomly assigned to 1 of 3 groups as follows: (1) control group that was colostrum deprived and fed milk replacer for first feeding, (2) group that was colostrum deprived and fed milk replacer mixed with antirotavirus antibodies for first feeding, or (3) group that was fed colostrum replacer mixed with antirotavirus antibodies and a product approved by the US Department of Agriculture containing antibodies against Escherichia coli K99 and bovine coronavirus for first feeding. One of the 3 treatments was administered within 6 h of birth to each calf, followed by oral challenge with bovine rotavirus 3 h later. Calves were observed through 7 d of age and scored according to a standardized scale for clinical signs of diarrhea, change in appetite, depression, and dehydration. Twice daily, measurements of rectal temperature and collection of feces were performed. Fecal samples were assessed for infectious agents commonly associated with diarrhea, and bovine rotavirus shedding was quantified. There were 24 of 28 (86%) calves in the control group that received no antibodies that had signs of severe diarrhea, whereas 57% of the calves that received antirotavirus in milk replacer experienced severe diarrhea, and 7% of calves that received colostrum replacer mixed with antigen-specific bovine rotavirus antibodies showed signs of severe diarrhea. Calves that received colostrum replacer mixed with antigen-specific bovine rotavirus antibodies had a mean duration of 0.9 d of diarrhea compared with 2.7 d in the control group. Calves in the group that was colostrum deprived and fed milk replacer with antirotavirus antibodies had a mean duration of diarrhea of 1.7 d. Rotavirus peak fecal shedding was 3.5 d in the group with milk replacer only, 5.5 d in the milk replacer with antibody group, and 6.5 d in calves in the colostrum replacer group. When bovine rotavirus antigen-specific antibody was fed in milk replacer to colostrum-deprived calves or in conjunction with colostrum replacer that also contained supplemental antibodies against Escherichia coli K99 and bovine coronavirus, those calves were observed to have reduced the onset, duration, and severity of diarrhea when compared with milk replacer placebo.     

Effect of waste milk pasteurization on fecal shedding of Salmonella in preweaned calves

The objective of the current research was to determine if pasteurization of nonsaleable waste milk influences fecal Salmonella concentrations and prevalence, or antimicrobial susceptibility and serotype of the cultured isolates. Holstein dairy calves (n = 211) were housed on a single commercial dairy in the southwestern United States and randomly allotted to be fed either pasteurized (PWM; n = 128 calves) or nonpasteurized waste milk (NPWM; n = 83 calves). Fecal samples were collected via rectal palpation or from freshly voided, undisturbed fecal pats, weekly during the first 4 wk of the animal's life and then again at weaning. Eight total collections were made and 1,117 fecal samples cultured for Salmonella. One isolate from each culture-positive fecal sample was preserved for antimicrobial susceptibility screening and serotyping. Sixty-nine percent of the fecal samples were culture positive for Salmonella with no difference due to treatment (67.7 and 69% Salmonella positive for PWM and NPWM treatments, respectively). Few fecal samples (178/1,117; 15.9%) contained Salmonella concentrations above the limit of detection (～1 cfu/g of feces) with concentrations ranging from 1.0 to 6.46 cfu (log₁₀)/g of feces. Concentration was not affected by treatment. Seventeen different serotypes were identified, the majority of which were Montevideo and Anatum. A greater percentage of Typhimurium (87 vs. 13%), Muenchen (88 vs. 12%), and Derby (91 vs. 9%) were recovered from calves fed PWM compared with NPWM-fed calves. Conversely, Newport (12.5 vs. 86%), Bredeney (22.2 vs. 77.8%), and Muenster (12.5 vs. 87.5%) were lower in PWM compared with NPWM treatments. The majority (66.7%) of isolates were susceptible to all of the antibiotics examined. Results from this one commercial dairy suggest that milkborne Salmonella is not an important vector of transmission in dairy neonates, nor does pasteurization of waste milk influence fecal shedding of this pathogen. Caution should be used, however, when extrapolating results to other farms as Salmonella contamination of milk on farm is well documented. The potential benefits of pasteurization in disease prevention outweigh the potential risks of feeding a nonpasteurized product and warrants incorporation into any calf-rearing program using nonsaleable waste milk for feeding young dairy neonates.     

The effect of neomycin inclusion in milk replacer on the health,growth, and performance of male Holstein calves during preweaning

The prophylactic use of oral antimicrobials, such as neomycin, in milk replacer (MR) or whole milk is a common practice in calf-rearing that is thought to aid in preventing disease. Heavy reliance on antimicrobials is of concern not only because of the development of antimicrobial resistance, but also because of the potentially negative effects on health. The objective of this study was to investigate the effects of neomycin on calf health and growth performance. One hundred and sixty calves (approximately 3–10 d of age), distributed across 2 experimental periods, were stratified by body weight (BW) and serum total protein, and assigned to 1 of 3 treatments: control (CON; nonmedicated MR, n = 60), short-term antibiotic (ST; neomycin mixed in MR from d 1–14, n = 50), or long-term antibiotic (LT; neomycin in ?MR from d 1–28, n = 50). Arrival BW (47.69 ± 0.87 kg) and serum total protein (5.67 ± 0.09 g/dL) were not different between treatment groups. Neomycin in ST and LT was dosed in MR at a rate of 20 mg/kg of BW and was adjusted weekly according to BW. Calf BW was measured weekly for 49 d, and health indicators (fecal score, attitude score, respiratory score, and rectal temperature), MR intake, starter intake, and the use of additional electrolytes and antimicrobials were recorded daily. Calves in the CON group experienced a higher proportion of days with diarrhea (20.32 ± 0.02%) compared with ST (14.70 ± 0.02%) or LT (13.80 ± 0.02%) calves, as well as longer bouts of diarrhea (7.45 ± 0.38 d, 5.69 ± 0.46 d, and 5.62 ± 0.45 d for CON, ST, and LT calves, respectively). Calves in the CON group also experienced higher fecal scores (score of 0.64 ± 0.04) than ST (score of 0.53 ± 0.04) or LT (score of 0.49 ± 0.04) calves, especially at d 7. However, no differences were observed in other health-related measures. The time to reach first diarrhea and first respiratory illness was not different between treatments, nor was the time to recover from respiratory illness. The time to intervention with additional electrolytes or antimicrobials was not different between treatment groups. Furthermore, growth performance, feed intake, and feed conversion ratio were not different. No differences were found when comparing ST and LT, except in the defined daily dose of total antimicrobials received. Calves in the LT group received a higher overall dose than ST calves, and both ST and LT calves received a higher dose than CON calves, which received no prophylactic antimicrobials. Given that there were no differences in performance variables and no additional health benefits aside from reduced fecal scores in calves fed neomycin, current practices involving the use of antimicrobials on dairy and veal operations need to be considered more prudently.     

Influence of farming methods on microbiological contamination and prevalence of resistance to antimicrobial drugs in isolates from beef

The presence of Escherichia coli, Staphylococcus aureus, Listeria monocytogenes and Salmonella spp. was determined in 75 samples of conventional beef and in 75 samples of organic beef. All samples came from cattle slaughtered and processed in the same slaughterhouse and quartering room. A total of 180 E. coli, 180 S. aureus and 98 L. monocytogenes strains were analyzed by an agar disk diffusion assay for their resistance to 11 antimicrobials, for the case of E. coli and S. aureus, or 9 antimicrobials, for the case of L. monocytogenes. Salmonella spp. were not isolated from any of the beef samples. No significant differences in prevalence were obtained for any of the bacterial species tested between organic and conventional beef. E. coli isolated from organic beef exhibited significant differences in antimicrobial resistance against 5 of the 11 antimicrobials tested as compared to isolates recovered from conventional beef. In the case of S. aureus, these differences were only found for 3 of the 11 antimicrobials tested and for L. monocytogenes, no differences were obtained between isolates obtained from organic or conventional beef. Although no significant differences were obtained in microbiological contamination, E. coli and S. aureus isolates from organically farmed beef samples showed significantly lower rates of antimicrobial resistance in E. coli and S. aureus isolates.     

Effects of mannan-oligosaccharide and Bacillus subtilis supplementation to preweaning Holstein dairy heifers on body weight gain,diarrhea, and shedding of fecal pathogens

The objective of this clinical trial was to evaluate the effectiveness of probiotic, prebiotic, and synbiotic supplementation on average daily weight gain (ADG), duration of diarrhea, age at incidence of diarrhea, fecal shedding of Cryptosporidium oocysts, enteric pathogens, and the odds of pneumonia in preweaning dairy heifer calves on a commercial dairy. Feeding prebiotics and probiotics may improve health and production of calves. Hence, healthy Holstein heifer calves (n = 1,801) from a large California dairy were enrolled at 4 to 12 h of age and remained in this study until weaning at 60 d of age. Calves were block randomized to 1 of 4 treatments: (1) control, (2) yeast culture enriched with mannan-oligosaccharide (prebiotic), (3) Bacillus subtilis (probiotic), and (4) combination of both products (synbiotic), which were fed in milk twice daily from enrollment until weaning. Serum total protein at enrollment and body weight at 7, 42, and 56 d of age were measured. Fecal consistency was assessed daily for the entire preweaning period. A subgroup of 200 calves had fecal samples collected at 7, 14, 21, and 42 d for microbial culture and enumeration of Cryptosporidium oocysts by direct fluorescent antibody staining. Synbiotic-treated calves had 19 g increased ADG compared with control calves for overall ADG, from 7 to 56 d. From 42 to 56 d, prebiotic-treated calves had 85 g greater ADG and synbiotic-treated calves had 78 g greater ADG than control calves. There was no difference in duration of the first diarrhea episode, hazard of diarrhea, or odds of pneumonia per calf with treatment. Probiotic-treated calves had 100 times lower fecal shedding of Cryptosporidium oocysts at 14 d and prebiotic-treated calves had fewer Escherichia coli and pathogenic E. coli at 42 d compared with control calves. Although there were no effects on duration of diarrhea or pneumonia incidence, greater ADG in the late preweaning period may reflect treatment effects on enteric pathogens during the rearing process. The decreased shedding of Cryptosporidium should reduce infectious pressure, environmental contamination, and public health risks from Cryptosporidium. Our findings suggest ADG and potential health benefits for calves fed prebiotics, probiotics, and synbiotics and can help the dairy industry make informed decisions on the use of these products in dairy production.