Factors associated with the risk of antibiotic residues and intramammary pathogen presence in milk from heifers administered prepartum intramammary antibiotic therapy.

Heifers (n=136) from 5 herds were treated with a commercially available beta-lactam intramammary (IMM) antibiotic preparation containing cephapirin sodium at 10-21 d prior to anticipated parturition to evaluate the risk of antibiotic residues occurring in milk postpartum and to determine factors associated with antibiotic residues and IMM pathogen presence in milk postpartum. Mammary secretions collected from quarters before antibiotic administration and during weeks 1, 2 and 3 postpartum were analyzed for mastitis pathogens. Composite milk was collected at milkings 3, 6 and 10 postpartum and analyzed for beta-lactam residues using a microbial inhibition antibiotic residue screening test. Antibiotic residues were confirmed with beta-lactamase treatment and re-tested for residues. Residues were detected in 28.0, 8.82 and 3.68% of milk samples obtained at the third, sixth, and tenth milking postpartum, respectively. Increases in interval between prepartum antibiotic therapy and parturition and an increase in the postpartum interval to sampling were associated with a decrease in risk of antibiotic residues. The presence of antibiotic residues in milk at the third milking was associated with a reduced risk for IMM pathogen prevalence in the first 21 d postpartum. Lower somatic cell counts, an increase in mean milk yield over 200 days in milk and a reduction in IMM pathogen prevalence were associated with the presence of an antibiotic in milk postpartum. Screening milk for antibiotic residues in milk postpartum following prepartum antibiotic therapy in heifers is recommended to reduce the risk for antibiotic residue contamination of milk.

Effects of prepartum intramammary antibiotic therapy on udder health, milk production, and reproductive performance in dairy heifers.

Preparturient heifers (n = 561) from 9 herds in 6 US states and 1 Canadian province were enrolled in a study to test the hypothesis that prepartum intramammary therapy would cure existing intramammary infections (IMI) and lead to increased milk production, reduced linear somatic cell count (LSCC), and improved reproductive performance. Mammary secretions were collected 10 to 21 d before expected calving from each quarter. Heifers were then assigned by identification number to receive intramammary therapy consisting of infusion of one tube per mammary quarter of a lactating cow commercial antibiotic preparation containing cephapirin or to a nontreated control group. Overall, 34.1% of mammary quarters were infected with a mastitis pathogen before parturition and 63.4% of heifers had at least one mammary quarter infected. The coagulase-negative staphylococci (CNS) caused the majority (74.8%) of prepartum IMI. Coagulase-positive staphylococci, environmental streptococci, and coliforms accounted for 24.5% of prepartum infections. Treatment had a significant effect on the cure rate of infected mammary quarters. Mammary quarters that were infected prepartum and treated with antibiotics had a 59.5% efficacy of cure rate and the percentage reduction in heifers with IMI was 51.9. Control quarters had a spontaneous cure rate of 31.7%. Treatment did not significantly affect milk production or LSCC in the first 200 d of lactation; however, there was a significant treatment by herd interaction for milk production. Quarters cured of either CNS or major pathogens had a lower LSCC in the first 200 d of lactation. No significant effect on services per conception or days open between treatment and control groups was observed. This trial demonstrated that prepartum intramammary antibiotic therapy did reduce the number of heifer IMI postpartum. Milk production, LSCC, and reproductive performance during the first 200 d of the first lactation were not significantly affected by treatment. Given these results, use of prepartum intramammary antibiotic therapy in heifers as a universal strategy to increase milk production in first-lactation dairy cows may not be warranted.

Efficacy of a 0.1% iodine teat dip against Staphylococcus aureus and Streptococcus agalactiae during experimental challenge.

An experimental challenge trial was performed according to the guidelines recommended by the National Mastitis Council (NMC). A 0.1% iodine teat dip (Quartermate with I-Tech) was examined. This product gave an 87.9% reduction of new intramammary infections with Staphylococcus aureus and a 66.5% reduction for Streptococcus agalactiae compared with a negative control. Teat end and teat skin characteristics remained excellent throughout the trial.

Efficacy of two iodine teat dips during experimental challenge with Staphylococcus aureus and Streptococcus agalactiae.

An experimental challenge trial was performed against Staphylococcus aureus and Streptococcus agalactiae following the procedures recommended by the National Mastitis Council. The efficacy of two teat dips, product 1 (Bovadine with I-Tech II) and product 2 (Bovadine with I-Tech, used as a positive control), was determined. Both teat dips contain 1% iodine and 10% glycerin. Product 1 established an 89.7% reduction in infections against Staph. aureus and 73.1% reduction in infections against Strep. agalactiae. Product 2 demonstrated an 86.2% reduction in infections against Staph. aureus and 78.4% reduction in infections against Strep. agalactiae. Teat skin and teat ends were evaluated before and after the trial. No significant change in teat condition was observed for either product.

Germicidal activities of representatives of five different teat dip classes against three bovine mycoplasma species using a modified excised teat model.

Six representative teat dips from five different teat dip classes were tested for germicidal activity against challenge exposure to Mycoplasma bovis, Mycoplasma californicum, and Mycoplasma bovigenitalium using a modified excised teat model. All teat dip formulations tested were efficacious against all of the Mycoplasma species, providing bacterial logarithmic reductions above 4. The germicides performed best against M. bovigenitalium with an average log reduction (LR) of 6.29. Average LR were 5.41 and 5.70 against M. bovis and M. californicum, respectively. The iodine and chlorhexidine products performed best against M. bovis and M. californicum, respectively, with complete kill of all organisms. The chlorhexidine and the barrier chlorine product also had complete kill of M. bovigenitalium organisms.

Prevalence of mastitis in dairy heifers and effectiveness of antibiotic therapy.

Dairy heifers were treated 0 to 90 d, 90 to 180 d, or 180 to 270 d prepartum with one of five different antibiotic products to determine the best time and with which product they should be treated prior to calving. Two hundred thirty-three heifers were included in the study. At the initial sampling, 56.5% of quarters were infected with some type of organism and 15.4% of quarters were infected with Staphylococcus aureus. Treatments included a cephapirin dry cow product, a penicillin-novobiocin dry cow product, a penicillin-streptomycin dry cow product, an experimental dry cow product containing tilmicosin, and a cephalonium dry cow product not available in the United States. Cure rates for the five antibiotic products indicated that all were equally effective against Staph. aureus and all were significantly more effective than the spontaneous cure rate observed in untreated control quarters. No differences in efficacy were observed due to the different treatment times prepartum. However, fewer new Staph. aureus infections occurred after treatment in the group treated at 180 to 270 d prepartum, indicating that treatment in the third trimester will reduce the chances of new intramammary infections occurring after treatment and persisting to calving.

A comparison of two commercially available Escherichia coli J5 vaccines against E. coli intramammary challenge.

The efficacy of two commercially available Escherichia coli J5 bacterins was investigated. Jersey cows were randomly assigned to one of three treatment groups: 1) unvaccinated controls, 2) vaccinated with J.VAC (Merial Limited, Athens, GA), and 3) vaccinated with J5 bacterin. All cows were vaccinated at drying off and at 2 wk before anticipated calving. Cows that were vaccinated with the J5 bacterin also received a third immunization at calving. One quarter of each cow was challenged with approximately 64 cfu of E. coli at 14 to 30 d postcalving. Immunization by either vaccine did not influence the severity of coliform mastitis; however, the mean number of colony-forming units of E. coli recovered from challenged quarters was significantly lower for immunized cows than for control cows at 144 h postchallenge. Serum and mammary secretion immunoglobulin (Ig)G, IgG1, and IgG2 titers against E. coli J5 whole-cell antigens were enhanced in vaccinated cows. Serum and mammary secretion IgM were not different among treatment groups. Somatic cell counts in milk from challenged quarters, rectal temperatures, and the clinical status of cows following intramammary challenge were not different among treatment groups.

Deoxyribonucleic acid fingerprinting of Staphylococcus aureus from heifer mammary secretions and from horn flies.

Staphylococcus aureus isolated from heifer mammary secretions, streak canals, and horn flies (Haematobia irritans) were evaluated by randomly amplified polymorphic DNA fingerprinting. The relationship between DNA fingerprint patterns of S. aureus isolated from horn flies and S. aureus isolated from heifer mammary glands was examined. Amplified DNA fragments were visualized by agarose gel electrophoresis and were analyzed by densitometry. Analysis of DNA fingerprint patterns of 56 S. aureus isolates that were obtained from heifer mammary secretions or streak canals resulted in three distinct subtypes of S. aureus. Of these, 31 isolates (55%) belonged to subtype 1, 22 isolates (39%) belonged to subtype 2, and 3 (5%) belonged to subtype 3. Eight of 10 S. aureus isolates from horn flies belonged to subtype 1, and 2 isolates belonged to subtype 2. Thus, all of the S. aureus isolates from horn flies had DNA fingerprint patterns identical to the majority (95%) of S. aureus isolates from heifer mammary secretions or streak canals. In addition, 10 S. aureus isolates from multiparous cows from the same herd were examined by randomly amplified polymorphic DNA. All S. aureus isolates from multiparous cows belonged to subtype 3. Results of this study suggest that horn flies may play an important role in the transmission of S. aureus to nulligravid and primigravid heifers. Furthermore, this study demonstrates the usefulness of randomly amplified polymorphic DNA fingerprinting to distinguish between different subtypes of S. aureus and to draw epidemiological inferences from the information it provides.

Efficacy of parenterally or intramammarily administered tilmicosin or ceftiofur against Staphylococcus aureus mastitis during lactation.

Two antibiotic preparations, tilmicosin and ceftiofur, were tested intramammarily and parenterally against Staphylococcus aureus mastitis in lactating cows. Neither product was effective as a lactating cow treatment at the doses and durations of treatment tested. Injection or infusion of tilmicosin and infusion of ceftiofur resulted in reductions of bacteria present in milk; however, only one quarter treated with infusion of tilmicosin was cured, and no cures were observed for the other treatments. Somatic cell counts were transiently reduced by infusion of ceftiofur and by infusion and injection of tilmicosin; however, they returned to pretreatment values by 28 d posttreatment.

Comparison of tilmicosin and cephapirin as therapeutics for Staphylococcus aureus mastitis at dry-off.

Forty-four cows (26 Jerseys and 18 Holsteins) that had at least 1 mammary quarter that was naturally (n = 12) or experimentally (n = 84) infected with Staphylococcus aureus were allotted to three treatment groups of approximately equal number at the end of lactation. Cows were dried off by abrupt cessation of milking, and dry cow therapy was administered as an intramammary infusion of cephapirin benzathine at 10 ml per quarter, an intramammary infusion of tilmicosin (solution containing 300 mg/ml) at 5 ml per quarter, or a subcutaneous injection of tilmicosin at 5 mg/kg of body weight on the day of drying off and another injection 4 d later. Mammary secretions were monitored during the dry period and postpartum for antimicrobial residues, intramammary infection (IMI) status, and somatic cell counts. Results demonstrated the following percentage cures for IMI caused by Staph. aureus at 28 d postcalving based on individual mammary quarters: cephapirin benzathine, 78.1%; tilmicosin infused, 74.2%; and tilmicosin injected, 9.1%. During the first 4 wk after drying off, the mean concentration of tilmicosin in mammary secretions from cows infused with the antibiotic remained approximately 10-fold higher than that in secretions from cows injected with the antibiotic (3.43 vs. 0.32 ppm), and, by the time of calving, concentrations for cows treated with both methods were below the dilution limit of the assay (< 0.1 ppm). Results demonstrated that intramammary infusion of tilmicosin was equally as effective as cephapirin benzathine in curing IMI caused by Staph. aureus at drying off; however, the subcutaneous injection of tilmicosin at the dose used was not effective as a dry cow therapeutic against Staph. aureus.

Influence of route of vaccine administration against experimental intramammary infection caused by Escherichia coli.

The route of immunization of a commercially available Escherichia coli J5 bacterin was investigated. Jersey cows were randomly assigned to one of three treatment groups: 1) unvaccinated (control), 2) vaccinated subcutaneously in the neck, and 3) vaccinated in the area of the supramammary lymph node. Cows were vaccinated at drying off and at 2 wk prior to anticipated calving. Two quarters of each cow were challenged with approximately 60 cfu of E. coli at 14 d postcalving. Route of immunization in the neck or the area of the supramammary lymph node did not influence severity of coliform mastitis. However, the mean number of colony-forming units of E. coli recovered from challenged quarters was significantly lower for vaccinated cows than for control cows at 24 h postchallenge. A quicker milk yield recovery following intramammary challenge was also observed for vaccinated cows. Serum immunoglobulin (Ig) G, IgG1, and IgG2 and whey IgG1 and IgG2 antibody titers against E. coli J5 whole-cell antigens were significantly enhanced in vaccinated cows. Somatic cell counts in milk from challenged quarters and rectal temperatures following intramammary challenge were not different for cows across treatment groups. Immunization did not prevent intramammary infection.

Role of horn flies (Haematobia irritans) in Staphylococcus aureus-induced mastitis in dairy heifers.

OBJECTIVE: To determine whether Staphylococcus aureus can colonize in horn flies and whether colonization is sufficiently persistent for transmission of the organism to cows by flies. ANIMALS: 2 Jersey heifers exposed to infected horn flies. PROCEDURE: Staphylococcus aureus was allowed to colonize in horn flies, and duration of colonization was determined. Flies with colonized S aureus were allowed to feed on teats of uninfected heifers to determine whether intramammary infection could be transmitted from fly to heifer. Scab material from naturally infected heifers was submitted for bacteriologic culture to determine whether S aureus was present and whether scabs could serve as a possible source of S aureus for flies. RESULTS: Staphylococcus aureus colonized in horn flies and remained for up to 96 hours after exposure. Exposure of teats of uninfected heifers to horn flies colonized with S aureus resulted in intramammary infection in 3 of 4 exposed teats. Culture of scab material from teats of naturally infected heifers revealed high concentration of S aureus (> 107 colony-forming units/mg), and flies without previously colonized S aureus were allowed to feed on scabs; S aureus colonized in them just as readily as it did in flies that had fed on experimentally infected blood. CONCLUSIONS: Horn flies are capable of transmitting S aureus-induced intramammary infection to heifers, and scabs on teats are a potential source of S aureus. Fly control on dairy cows in herds with known S aureus problems is recommended as a method to help prevent these infections.

Comparison of success of antibiotic therapy during lactation and results of antimicrobial susceptibility tests for bovine mastitis.

Antimicrobial susceptibility testing was conducted on a variety of mastitis pathogens. The infected quarters were subsequently treated during lactation with a commercially available product containing penicillin and novobiocin that was designed for lactating cows. Cows were treated as per the recommendations of the product manufacturer, and cures were determined by the absence of bacteria in both sets of duplicate quarter milk samples that were collected at 28 d posttreatment. Comparisons were made between the susceptibility of the bacteria and the therapeutic success or failure. All isolates tested were considered to be susceptible to the penicillin and novobiocin combination. Bacteriologic cure rates for newly acquired Staphylococcus aureus intramammary infection (IMI) (< 2 wk in duration) at 28 d posttreatment were 70%. Cure rates for chronic Staph. aureus IMI (> 4 wk duration) were much lower (35%), reaffirming previous reports of the intractable nature of chronic Staph. aureus IMI. Cure rates for subclinical IMI caused by other organisms were 90% for Streptococcus agalactiae, 91% for Streptococcus uberis, 90% for Streptococcus dysgalactiae, 77% for other Streptococcus spp., and 71% for Staphylococcus spp. other than Staph. aureus. In vitro testing was considered to be a predictor of therapy outcome for IMI caused by Staphylococcus spp., newly acquired Staph. aureus, Strep. uberis, Strep. dysgalactiae, and Strep. agalactiae, but was not considered to be a useful predictor of efficacy for chronic IMI caused by Staph. aureus.

Therapeutic and prophylactic effect of prepartum antibiotic infusion in heifers.

Prepartum bacteriologic examination of secretions from 42 dairy heifers 12-14 weeks prepartum revealed a total of 24 Staphylococcus aureus infected quarters, 53 Staphylococcus species infected quarters, and 20 Streptococcus species infected quarters. Prepartum intramammary therapy of primigravid dairy heifers with two commercially available dry cow antibiotics (penicillin-novobiocin or cephapirin) resulted in cure rates of 94%, 97%, and 100% for S. aureus, Staphylococcus species, and Streptococcus species intramammary infections (IMI), respectively. No protective effect was observed for dry cow treatment of uninfected quarters of heifers for any of the antibiotic preparations. No antibiotic was detectable in heifer secretions collected at parturition indicating that antibiotic concentrations may have fallen below protective levels prior to parturition.

Changes in bovine mammary-secretion composition during early involution following intramammary infusion of recombinant bovine cytokines.

This study was designed to determine whether intramammary infusion of recombinant bovine interleukin-1 beta or interleukin-2 had any adverse influence on bovine mammary function during the early non-lactating period. Mammary glands of eight Jersey cows were infused with either 10 micrograms of interleukin-1 beta or 1.0 mg of interleukin-2 following abrupt cessation of milking. Mammary secretions from each gland were collected from cows frequently during early involution and evaluated for changes in secretion composition. Percentage of milk fat and solids-not-fat during the experimental period was variable for all treatments. Percentage of protein increased throughout the study for all treatments, but mammary glands treated with interleukin-1 beta had higher protein percentages. All interleukin-treated mammary glands had lower concentrations of lactose than controls. Mammary secretions from interleukin-1 beta treated glands had higher concentrations of lactoferrin at 7 and 14 days of involution. Similarly, mammary secretions from interleukin-2 treated glands had higher concentrations of serum albumin on day 3 of involution and throughout the remainder of the study. Concentration of citrate and the citrate: lactoferrin molar ratio generally decreased throughout the study for all treatment groups. Concentration of alpha-lactalbumin in mammary secretions decreased throughout the study and was not different between treatment groups. Though significant changes in mammary secretion composition due to the presence of cytokines were observed in this study, resulting in an apparent acceleration of involution, there was no indication that these changes adversely affected normal bovine mammary function during the early non-lactating period.

Mastitis in dairy heifers: initial studies on prevalence and control.

Initial studies to determine the prevalence of mastitis in heifers of breeding age and in pregnant dairy heifers demonstrated that IMI were present in 97% of heifers and 75% of quarters. The most common isolates were Staphylococcus aureus, Staphylococcus hyicus, and Staphylococcus chromogenes; SCC ranged from 12.4 to 17.3 x 10(6)/ml. Approximately 29% of heifers and 15% of quarters exhibited clinical mastitis at breeding age, as evidenced by clots or flakes in mammary secretions. Histologic examination of mammary tissues demonstrated significant reductions in alveolar epithelial and luminal areas and increases in connective tissue stroma and leukocytosis, illustrating limited development and marked inflammation of infected tissues. A one-time infusion of antibiotic for nonlactating cows into infected quarters > or = 45 d prepartum reduced incidence of IMI by 59% at calving compared with the pretreatment level; the cure rate for Staph. aureus IMI was > 90%. Prophylactic treatment of uninfected quarters > or = 45 d prepartum reduced new Streptococcus sp. IMI by 93%. The mean SCC was 50% lower at calving for treated heifers, and milk yield over the first 2 mo of lactation was 10% greater than that of untreated controls. Heifers from herds using fly control had a lower prevalence of IMI than herds without fly control. Prevalences of IMI and SCC in dairy heifers were higher than previously realized, but mastitis at calving was controlled by use of therapeutic products for nonlactating cows during pregnancy.

Prepartum antibiotic therapy with a cephapirin dry-cow product against naturally occurring intramammary infections in heifers.

Intramammary infusion of a dry-cow antibiotic preparation containing 300 mg of cephapirin benzathine into 18 Jersey heifers, 10-12 weeks prepartum, resulted in cure rates of existing intramammary infection (IMI) of 96% (24/25), 100% (4/4), and 90% (28/31) for Staphylococcus aureus, Streptococcus species, and Staphylococcus species, respectively. Cure rates of IMI that had been treated with a lactating-cow therapy containing 200 mg cephapirin benzathine at parturition were 62.5% (15/24), 100% (22/22), and 100% (3/3) for Staphylococcus aureus, Streptococcus species, and Staphylococcus species, respectively. Initial somatic cell counts (SCC) of secretions from infected quarters were greater than from uninfected quarters. At 2 months postpartum, the SCC of milk from treated and cured quarters were reduced in comparison with quarters that remained infected. Cephapirin benzathine was present at detectable concentrations in 94, 80, 68, and 61% of treated quarters at 1, 2, 3, and 4 weeks after infusion of the cephapirin dry-cow product, respectively. At parturition, 24% of treated quarters were positive for antibiotic, however, no quarters remained positive for antibiotic at 5 days postpartum. An additional 40 heifers from a commercial herd were sampled and infused in all quarters with the cephapirin dry-cow product at 16-20 weeks prepartum. Cure rates for the commercial herd were 94% (29/31), 94% (16/17), 100% (44/44), and 100% (3/3), respectively, for quarters infected by S. aureus, Streptococcus species, Staphylococcus species, and coliforms.

The postantibiotic effect of selected antibiotics on Staphylococcus aureus Newbould 305 from bovine intramammary infection.

The postantibiotic effect (PAE) was determined for selected antibiotics against Staphylococcus aureus Newbould 305 originating in vivo from mastitic milk and compared with the PAE for the same S. aureus strain cultured in vitro. The PAE was measured at 2 and 4 times the MIC for 1 and 2 h of exposure. The PAE of penicillin, pirlimycin, and tilmicosin were reduced against S. aureus 305 originating in vivo compared with S. aureus 305 grown in vitro. The PAE of cephapirin was increased against S. aureus originating in vivo. Minimal effect on PAE was noted for novobiocin. The PAE for rifampicin extended beyond the limits of the test parameters (> 180 min) for all antibiotic concentrations, media, and exposure times tested, except at 2 times the MIC and at 1 h exposure when the PAE was reduced to 60 min for S. aureus 305 originating in vivo. The PAE of an antibiotic may be an important consideration in determining therapy intervals and antibiotic concentrations for treatment of bovine mastitis.

Effects of interleukin-1 and interleukin-2 on mammary gland leukocyte populations and histology during the early nonlactating period.

Eight Jersey cows free of intramammary infection (IMI) with major mastitis pathogens were intramammarily infused with interleukin (IL)-1, IL-2, or phosphate-buffered saline (PBS) after the last milking prior to drying off. Rectal temperatures were monitored hourly through 6 hours after infusion and at 24-hour intervals thereafter. Quarter secretions were collected 7 days prior to drying off, at drying off, and on days 1, 2, 3, 4, 7, and 14 of the nonlactating period. Cows were slaughtered and mammary tissues were collected at 7 (3 cows) and 14 (5 cows) days after treatment. The infusion of 10 micrograms of IL-1 produced an elevation in rectal temperature that was significantly higher than that of cows receiving IL-2 or PBS at 4, 5, and 6 hours after treatment, whereas cows receiving 1.0 mg of IL-2 exhibited no change in rectal temperature. Somatic cell counts (SCC) in mammary glands infused with IL-1 or IL-2 remained significantly elevated over PBS-infused control glands through day 4 after treatment. Neutrophils predominated in secretions of IL-1-treated glands, and macrophages and lymphocytes predominated in IL-2-treated glands. The mammary involution process was accelerated by IL-2, and tissues exhibited reductions in alveolar epithelial and luminal areas and increases in connective tissue stroma and degree of leukocytosis compared with IL-1 and control glands. Concentrations of plasma cells producing IgG1, IgG2, IgA, and IgM also were greatest following IL-2 infusion. Results suggest that IL-2 is superior to IL-1 in stimulating an increase in mammary mononuclear cell populations and in accelerating involution during the early nonlactating period.