Short communication: Effect of 3 phytoceutical products on elimination of bacteria in experimentally induced Streptococcus uberis clinical mastitis

Our objective was to assess the ability of 3 herbal products to eliminate experimentally induced Streptococcus uberis mastitis. These herbal products, also known as phytoceuticals, are used in organically managed dairy cattle to maintain or promote udder health. The products tested were an intramammary product, a topical product, and a product applied to the vulvar area. These products are not approved by the US Food and Drug Administration for treatment of mastitis but they are sold to enhance milk quality or for maintenance or improvement of udder health. Each of the products contains at least one component shown to have antibacterial activity. In this study, we successfully challenge-inoculated 25 lactating dairy cows maintained under organic conditions with an isolate of S. uberis. All challenged cows were positive for S. uberis by milk culture after challenge. When cows met predefined criteria indicating the presence of clinical mastitis, treatment with 1 of the 3 products was initiated based upon a predetermined random allocation. Culture of aseptically collected quarter milk samples was performed before, during, and following challenge with S. uberis. Eight, 8, and 9 cows received the intravulvar, intramammary, and topical treatments, respectively. Milk from all cows that were treated with phytoceuticals were culture-positive for S. uberis at every time point following treatment through 168 h following the last phytoceutical treatment. Based upon the presence of clinical signs and for humane reasons, 2 intravulvar-treated cows, 1 topical-treated, and 4 intramammary-treated cows received intramammary antibiotic therapy. We concluded that the phytoceuticals tested, as dosed and used in this trial, did not produce bacterial cures in S. uberis-induced mastitis.     

Technical note: Development of a challenge model for Streptococcus uberis mastitis in dairy heifers

A challenge model for experimentally inducing Streptococcus uberis mastitis in bred dairy heifers was developed. Qualifying heifers (n = 7) exhibited antibody titers of < 1:10,000 against Strep. uberis antigens and were free of intramammary infections (IMI). Two contralateral quarters of each heifer were assigned to receive an infusion of Strep. uberis (1,000 to 2,000 cfu); remaining quarters served as unchallenged controls. For a successful challenge and infection, 3 of 4 consecutive mammary secretion samples had to culture positive for Strep. uberis. Six of the 7 heifers were challenged successfully in both infused quarters with a mean dose of 1,080 cfu; once confirmed, infections were treated with a one-time infusion of nonlactating cow therapy. Before challenge, mammary secretion leukocyte counts averaged 8.4 × 10⁶/mL in all quarters. At 24 h after challenge, leukocyte count increased to 18.4 × 10⁶/mL in challenged quarters, peaking on d 5 at 24.3 × 10⁶/mL; unchallenged quarters remained at ≤ 10.4 × 10⁶/mL, but increased to 15.2 × 10⁶/mL on d 7 and then decreased. Before challenge, macrophages predominated (81%) in mammary secretions followed by lymphocytes (15.3%) and neutrophils (3.7%). By 24 h after challenge, neutrophils increased in challenged quarters and predominated for the duration of the trial (65.3 to 70%), whereas macrophages predominated in unchallenged control quarters (65.2 to 75.2%). The challenge model was successful in establishing Strep. uberis IMI in 85.7% of animals, and IMI were controlled (100% cure) by administering nonlactating cow therapy. All heifers calved free of IMI and antimicrobial residues, with milk production similar to that of herd mates and with somatic cell counts (SCC) < 200,000 cells/mL.     

The effect of J5 bacterins on clinical,behavioral, and antibody response following an Escherichia coli intramammary challenge in dairy cows at peak lactation

Vaccination against coliform mastitis has become part of mastitis control programs in the past 3 decades, as a means of reducing the severity of clinical mastitis. Our study objective was to evaluate the effect of 2 commercially available vaccines on clinical, behavioral, and antibody response following Escherichia coli intramammary challenge in cows near peak lactation. Cows (n = 12 per group) were vaccinated with vaccine 1 (V1) or vaccine 2 (V2) at dry-off, 21 d pre-calving, and 14 d post-calving. Twelve cows served as unvaccinated controls (CTL). Cows were challenged with E. coli in a rear quarter at approximately 100 d in milk. Milk samples were collected pre- and post-challenge to enumerate E. coli and determine somatic cell count. Serum was collected before each vaccination and at d 0, 1, 2, 3, 6, 30, and 60 relative to challenge, to study antibody response. Milk IgA and tumor necrosis factor-α concentrations were determined in whey. Vaginal temperature, cow activity, and milk yield and components were monitored post-challenge. Bacterial count, somatic cell score, milk yield and component decline, vaginal temperature, activity measures, and antibody and cytokine response were analyzed for treatment differences. The effects of parity, breed, and a repeated measure of time were also tested. Seven cows had to be removed from the study post-challenge for antibiotic treatment (CTL and V1, n = 3 each; V2, n = 1), 2 of which were euthanized (both CTL). Vaccinated cows exhibited fever (vaginal temperature ≥39.4°C) 3 h earlier than CTL cows, but we found no differences between treatments for bacterial count, somatic cell score, or milk yield reduction. Vaccinated cows spent more time lying per rest bout 2 d post-challenge, but total daily lying time was not different from CTL cows during the 7 d post-challenge. The vaccines differed in antibody response: V1 cows had greater serum IgG₁ and IgG₂ post-challenge. A parity effect was also evident: primiparous cows had lower bacterial counts, somatic cell score and a smaller milk yield decline than multiparous cows, but also had lower antibody production. Immunization with either J5 bacterin did not reduce clinical signs of mastitis in cows challenged at 100 d in milk, demonstrating that the effects of J5 vaccination had diminished at peak lactation.     

Efficacy of an Escherichia coli J5 mastitis vaccine in an experimental challenge trial.

An Escherichia coli (O111:B4) J5 bacterin was tested for efficacy in reducing IMI and severity of clinical coliform mastitis in an experimental challenge trial. Ten cows were immunized at drying off, 30 d after drying off, and at calving. Ten control cows were not immunized. Right front quarters of all cows were infused with a heterologous strain of E. coli approximately 30 d after calving. Vaccinated cows had lower bacterial counts in milk and lower rectal temperatures than unvaccinated controls following intramammary challenge. Milk production and DMI were more depressed in controls than in vaccinated cows. Milk SCC did not differ between experimental groups. Mean serum IgG titer to whole cell E. coli J5 was significantly greater in vaccinated than in unvaccinated cows at 30 d after drying off, day of challenge, and 7 d postchallenge. Milk IgG titer to E. coli J5 was higher at challenge in vaccinated than in control cows. Vaccination with the E. coli J5 bacterin did not prevent IMI but did reduce severity of clinical signs following intramammary experimental challenge with a heterologous E. coli strain.     

Evaluation of reticuloruminal temperature for the prediction of clinical mastitis in dairy cows challenged with Streptococcus uberis

Automated monitoring devices have become increasingly utilized in the dairy industry, especially for monitoring or predicting disease status. While multiple automated monitoring devices have been developed for the prediction of clinical mastitis (CM), limitations in performance or applicability remain. The aims of this study were to (1) detect variations in reticuloruminal temperature (RRT) relative to an experimental intramammary challenge with Streptococcus uberis and (2) evaluate alerts generated automatically based on variation in RRT to predict initial signs of CM in the challenged cows based on severity of clinical signs and the concentration of bacteria (cfu/mL) in the infected quarter separately. Clinically healthy Holstein cows without a history of CM in the 60 d before the experiment (n = 37, parity 1 to 5, ≥120 d in milk) were included if they were microbiologically negative and had a somatic cell count under 200,000 cells/mL based on screening of quarter milk samples 1 wk before challenge. Each cow received an intra-reticuloruminal automated monitoring device before the trial and was challenged with 2,000 cfu of Strep. uberis 0140J in 1 rear quarter. Based on interrupted time series analysis, intramammary challenge with Strep. uberis increased RRT by 0.54°C [95% confidence interval (CI): 0.41, 0.66] at 24 h after the challenge, which remained elevated until the end of the study. Alerts based on RRT correctly classified 78.3% (95% CI: 65.8, 87.9) of first occurrences of CM at least 24 h in advance, with a sensitivity of 70.0% (95% CI: 50.6, 85.3) and a specificity of 86.7% (95% CI: 69.3, 96.2). The accuracy of CM for a given severity score was 90.9% (95% CI: 70.8, 98.9) for mild cases, 85.2% (95% CI: 72.9, 93.4) for moderate cases, and 92.9% (95% CI: 66.1, 99.8) for severe cases. Test characteristics of the RRT alerts to predict initial signs of CM improved substantially after bacterial count in the challenged quarter reached 5.0 log₁₀ cfu/mL, reaching a sensitivity of 73.5% (95% CI: 55.6, 87.1) and a specificity of 87.5% (95% CI: 71.0, 96.5). Overall, the results of this study indicated that RRT was affected by the intramammary challenge with Strep. uberis and the RRT-generated alerts had similar accuracy as reported for other sensors and algorithms. Further research that includes natural infections with other pathogens as well as different variations in RRT to determine CM status is warranted.     

Efficacy of a novel internal dry period teat sealant containing 0.5% chlorhexidine against experimental challenge with Streptococcus uberis in dairy cattle

The incidence of clinical mastitis and infection status at calving was assessed in quarters treated with 1 of 2 internal teat sealants at the time of dry off. Two contralateral quarters per cow (n = 63 cows) were treated with a sealant that contained 0.5% chlorhexidine; the other quarters were treated with a commercial teat sealant. Ten cows were untreated (controls). On d 2, 4, and 16 after dry off, cows were challenged with Streptococcus uberis S210 strain. Cows were examined daily for 34 d after drying off and cases of clinical mastitis were recorded. Milk samples were collected for culture from any quarters that developed clinical mastitis during the first 34 d after drying-off and from all quarters on d −5 and 0 relative to treatment and at the first and twentieth milking after calving. The incidence of clinical mastitis during the examination period was lower in treated quarters (n = 7/252; 1.5%; lower incidence for those treated with chlorhexidine-containing teat sealant n = 3/126; 1.2%) than in untreated quarters (n = 13/40; 26.8%). The protection against intramammary infection after calving, adjusted for the effect of cow, was higher in quarters treated with the novel teat sealant (89/105; 15.2%; 95% CI = 9.6-23.4) than in those treated with the commercial teat sealant (71/104; 31.7%; 95% CI = 23.5-41.3) and untreated controls (6/28; 78.6%; 95% CI = 59.8-90.0), respectively. Quarters treated with teat sealants were less likely to have an intramammary infection after calving and had a lower incidence of clinical mastitis during the early dry period than did untreated controls in this challenge study.     

Immune response after an experimental intramammary challenge with killed Staphylococcus aureus in cows and heifers vaccinated and not vaccinated with Startvac,a polyvalent mastitis vaccine

An experimental trial was conducted to explore the effect of vaccination with a polyvalent vaccine against mastitis (Startvac) on the early immune response after experimental intramammary challenge with a heterologous killed Staphylococcus aureus strain. The effect of vaccination on milk production, clinical signs, quarter milk somatic cell count, milk polymorphonuclear neutrophilic leukocyte (PMN) concentration and viability, the concentration of antigen-specific antibodies [slime associated antigenic complex (SAAC) and J5] and their IgG1 and IgG2 subtypes in both serum and whey, and the antigen-specific IFN-γ, IL-4, and IL-17 production by blood lymphocytes after in vitro stimulation with S. aureus and Escherichia coli extracts were determined. A cohort of 8 clinically healthy end-term cows and heifers were conveniently selected, of which half was vaccinated with Startvac at 45 and 10 d before the expected calving date and half served as nonvaccinated control animals. At 15 d in milk, 2 contralateral quarters of each of the 8 animals were challenged with 2 × 10⁹ cfu/mL of the formaldehyde-killed S. aureusC195strain. The 2 other quarters were infused with phosphate-buffered saline and served as control quarters. The increase in both quarter milk somatic cell count and PMN concentration and the drop in milk production after S. aureus inoculation was less pronounced in the vaccinates than in the nonvaccinates, reflecting a less severe inflammatory response. No significant differences in PMN viability between vaccinates and nonvaccinates could be demonstrated. The serum SAAC- and J5-specific antibody concentration significantly increased across the dry period in the vaccinated animals only. The whey concentration of SAAC-specific antibodies was significantly higher in vaccinates than in nonvaccinates at both 15 and 17 d in milk, independent from the challenge status of the quarters. No significant differences in the whey J5-specific antibody concentration were observed. Vaccination with Startvac seems to primarily evoke a Th2 response for S. aureus characterized by a shift toward the IgG1 antibody subtype and accompanied by a less pronounced Th1 response. The type of response against E. coli was less clear, though a weak but significant shift toward the IgG2 antibody subtype after vaccination and high IFN-γ levels after in vitro stimulation suggest a Th1 response. The increased SAAC-specific antibody concentration in whey in vaccinates compared with nonvaccinates most probably triggers the opsonization of the inoculated S. aureus bacteria, resulting in a more efficient elimination of the bacteria from the mammary gland.     

Epidemiology of mastitis in pasture-grazed peripartum dairy heifers and its effects on productivity

An observational field study was conducted on 708 heifers in 30 spring-calving dairy herds in the Waikato region of New Zealand. The aim of the study was to describe patterns and effects of intramammary infection (IMI) and clinical mastitis (CM) in the peripartum period. Mammary secretion samples for bacteriological testing were taken from all quarters approximately 3 wk before the planned start of the calving period and within 5 d following calving, in addition to quarters diagnosed with CM within 14 d of calving. Precalving IMI was diagnosed in 18.5% of quarters, and of these, coagulase-negative staphylococci were the predominant isolate (13.5% of quarters). Streptococcus uberis prevalence increased 4-fold to 10.0% of quarters on the day of calving compared with the precalving period. Prevalence of all pathogens decreased rapidly following calving. Clinical mastitis cases were predominantly associated with Strep. uberis (64%). The daily hazard of diagnosis was higher in heifers than in cows (0.06 vs. 0.02/d on d 1 postcalving, respectively), but was not different by d 5 (0.005 vs. 0.002, respectively) of lactation. Intramammary infection with a major pathogen was associated with an increased risk of removal from the herd (15 vs. 10% for infected and noninfected heifers, respectively) and somatic cell count >200,000 cells/mL at subsequent herd tests (15 vs. 8%), but neither CM nor IMI were associated with reduced milk yield or milk solids production. Results suggest that bacterial species involved and the pattern of IMI prevalence in pasture-grazed peripartum heifers differ from those in other production systems. Further, mastitis control programs need to target major environmental pathogens causing precalving IMI, because new infections are likely before the onset of lactation, whereas existing detection and control measures are generally implemented after calving. Novel control programs that reduce new infections due to Strep. uberis immediately before calving are required to reduce the incidence of CM in pasture-grazed dairy heifers.     

Strain-specific pathogenicity of putative host-adapted and nonadapted strains of Streptococcus uberis in dairy cattle

Streptococcus uberis is an important cause of intramammary infection in dairy cattle. Strains of Strep. uberis appear to differ in their ability to cause disease based on previous epidemiological studies. We explored the pathogenicity of 2 strains of Strep. uberis, where one strain represented a putatively host-adapted type based on its ability to cause persistent infection and to spread from cow to cow in a lactating herd. This type was part of a clonal complex that is commonly associated with bovine mastitis. The other strain, which was isolated from a transient infection in a single animal in the same herd and did not belong to any known clonal complex, was selected as putatively nonadapted type. Cows (6 per strain) were experimentally challenged in a single hind quarter and the adjacent hind quarter was used as mock challenged control quarter. Both strains showed an equal ability to grow in the milk of challenge animals in vitro. All cows that were challenged with the putatively host-adapted strain developed clinical signs of mastitis, including fever and milk yield depression as well as elevated somatic cell count due to influx of polymorphonuclear leucocytes and lymphocytes. The cytokine response followed a specific order, with an increase in IL-1β, IL-6, and IL-8 levels at the time of first SCC elevation, followed by an increase in IL-10, IL-12p40, and tumor necrosis factor-α levels approximately 6 h later. In 4 of 6 animals, IL-17A was detected in milk between 57 and 168 h postchallenge. The increase in IL-17A levels coincided with inversion of the prechallenge CD4⁺-to-CD8⁺ T lymphocyte ratio, which was observed from 96 h postchallenge. This was followed by normalization of the CD4⁺-to-CD8⁺ ratio due to continued increase of the CD8⁺ concentration up to 312 h postchallenge. Spontaneous resolution of infection was observed in 5 animals and coincided with a measurable IL-17A response in 4 animals, suggesting that IL-17 may be involved in the resolution of intramammary infection. With the exception of minor elevation of IL-8 levels, no clinical, cytological, or immunological response was detected in quarters challenged with the nonadapted strain. The observed strain-specific pathogenicity was consistent across animals, implying that it is determined by pathogen factors rather than host factors.     

Evaluation of an alternative dosing regimen of a J-5 mastitis vaccine against intramammary Escherichia coli challenge in nonlactating late-gestation dairy cows

The objective of the study was to evaluate the efficacy of an alternative vaccination regimen of a J-5 bacterin against intramammary Escherichia coli challenge in nonlactating late-gestation dairy cows. The parameters analyzed to assess the effect of vaccination were milk yield, milk conductivity, somatic cell count, J-5-specific serum IgG titers, and clinical signs. Twenty multiparous Holstein cows from the Cornell teaching and research dairy herd were paired by days in milk and were randomly selected to receive either the alternative off-label regimen of commercial J-5 bacterin or act as nonvaccinated controls. Cows received a first dose of bacterin 15 d before dry off, a second dose with the same product at the day of dry off, and the third dose 2 wk after dry off. The cows in both groups were challenged 10 d before the expected calving date. Serum IgG (total, IgG1 and IgG2) levels were higher in vaccinates compared with control cows. Eighty-five percent of challenged quarters became infected between both groups of animals. Eight of the 10 vaccinated and 9 of the 10 control cows showed signs of clinical mastitis postfreshening. A non-severe clinical mastitis was observed 24 to 48 h postparturition, characterized by flakes or clots in milk and mild swelling or pain. Off-label vaccination did reduce the clinical severity of clinical mastitis in the vaccinated group of cows as evidenced by reduced California mastitis test score, fewer flakes and lower overall clinical mastitis score. No significant differences between vaccinated and control groups were detected for rectal temperature. In conclusion, the alternative off-label vaccination scheme used in our study and evaluated in a novel E. coli challenge model did not prevent new intramammary infections but reduced clinical severity of experimentally induced E. coli mastitis.     

Effects of local or systemic administration of meloxicam on mammary gland inflammatory responses to lipopolysaccharide-induced mastitis in dairy cows

Nonsteroidal anti-inflammatory drugs (NSAID) are commonly used in combination with antimicrobial mastitis treatments to reduce pain. Little is known about whether meloxicam, an NSAID designed for the preferential inhibition of cyclooxygenase-2 over cyclooxygenase-1, affects the mammary immune response. The objective of this study was to analyze the mammary immune response to intramammary (local) or intravenous (systemic) administration of meloxicam with or without immune activation by lipopolysaccharide (LPS). We challenged 108 quarters of 30 cows with or without a low or high dose of LPS from Escherichia coli (0.1 or 0.2 µg/quarter), with or without meloxicam via intramammary administration (50 mg/quarter) or intravenous injection (0.5 mg/kg of body weight; ~300 mg/cow). Intramammary administration of meloxicam alone did not trigger an acute inflammatory response, verified by unchanged somatic cell count (SCC) and lactate dehydrogenase (LDH), BSA, and IgG concentrations in milk, which are normally augmented during mastitis due to an opening of the blood–milk barrier. Similarly, intramammary meloxicam did not change the mRNA abundance of inflammatory factors in mammary gland tissue. As expected, quarters challenged with either dose of LPS showed increased leukocyte infiltration (SCC); increased LDH, BSA, IgG, Na, and Cl concentrations; and diminished K concentrations in milk. In contrast to our hypothesis, the addition of intramammary or intravenous meloxicam did not reduce these markers of mastitis in milk. Instead, intramammary meloxicam appeared to accelerate the SCC response to LPS, but only at the lower LPS dose. Moreover, the mRNA expression of inflammatory factors in mammary tissue was not modified by the intramammary application of meloxicam compared with the contralateral quarters that were challenged with LPS only. We demonstrated for the first time that intramammary meloxicam at a dose of 50 mg/quarter did not trigger an immune response in the mammary glands of dairy cows. At the doses we used, meloxicam (intramammary or systemic) did not lower inflammatory responses. The intramammary administration of meloxicam seemed to stimulate leukocyte recruitment into the milk in quarters challenged with a low dose of LPS. The integrity of the blood–milk barrier was not protected by meloxicam in LPS-stimulated quarters. This study provides the first indications that meloxicam does not limit the inflammatory response in the mammary gland, although it does not impair the mammary immune system.     

Efficacy of a lactoferrin-penicillin combination to treat {beta}-lactam-resistant Staphylococcus aureus mastitis

The efficacy of intramammary (IM) treatments containing penicillin G (PG) alone or a combination of PG and bovine lactoferrin (bLF) was evaluated using a model of experimentally induced chronic bovine mastitis caused by a clinical isolate of Staphylococcus aureus highly resistant to β-lactam antibiotics. First, we confirmed that this strain could cause mastitis and infection could not be cured with PG alone. In a second trial, chronic mastitis was induced in 19 late-lactating cows by injecting a low dose of Staph. aureus through the teat canal of all quarters. After 15 d, cows with stable infections in their 4 quarters had their mammary quarters randomly assigned, within cow, to 1 of 4 IM treatments as follows: 1) citrate buffer, 2) 100,000 IU of PG, (3) 1 g of bLF, or 4) 1 g of bLF + 100,000 IU of PG. Treatments were repeated twice a day for 5 d. A third trial was undertaken to investigate the effect of an extended therapy on chronic mastitis acquired in a previous lactation. One month before dry-off, 20 gravid cows regrouped by dates of calving were infected in their 4 quarters. Once infections were established, cows were dried off abruptly. After calving, aseptic milk samples were collected separately from all quarters for 4 wk to monitor infection. Mammary quarters from enrolled cows were then randomly assigned, within cow, to 1 of 2 treatments as follows: 1) 100,000 IU of PG or 2) 250 mg of bLF + 100,000 IU of PG. Treatments were administered IM twice a day for 7 d. For all trials, milk samples were taken to monitor bacterial concentration and somatic cell count. Bacteriological cure rate was determined using milk samples taken 3 and 4 wk after initiation of treatments. For the second trial, cure rate was null for control quarters, 11.1% for bLF, 9.1% for PG, and 45.5% for the bLF + PG combination. For cows infected in their previous lactation, cure rate was higher for the bLF + PG combination (33.3%) compared with PG alone (12.5%). In conclusion, bLF added to PG is an effective combination (i.e., 3- to 5-times higher cure rate) for the treatment of stable Staph. aureu infections highly resistant to β-lactam antibiotics.     

Evaluation of the use of dry cow antibiotics in low somatic cell count cows

The goal of dry cow therapy (DCT) is to reduce the prevalence of intramammary infections (IMI) by eliminating existing IMI at drying off and preventing new IMI from occurring during the dry period. Due to public health concerns, however, preventive use of antibiotics has become questionable. This study evaluated selective DCT in 1,657 cows with low somatic cell count (SCC) at the last milk recording before drying off in 97 Dutch dairy herds. Low SCC was defined as <150,000 cells/mL for primiparous and <250,000 cells/mL for multiparous cows. A split-udder design was used in which 2 quarters of each cow were treated with dry cow antibiotics and the other 2 quarters remained as untreated controls. The effect of DCT on clinical mastitis (CM), bacteriological status, SCC, and antibiotic use were determined at the quarter level using logistic regression and chi-squared tests. The incidence rate of CM was found to be 1.7 times (95% confidence interval = 1.4–2.1) higher in quarters dried off without antibiotics as compared with quarters dried off with antibiotics. Streptococcus uberis was the predominant organism causing CM in both groups. Somatic cell count at calving and 14 d in milk was significantly higher in quarters dried off without antibiotics (772,000 and 46,000 cells/mL, respectively) as compared with the quarters dried off with antibiotics (578,000 and 30,000 cells/mL, respectively). Quarters with an elevated SCC at drying off and quarters with a positive culture for major pathogens at drying off had a higher risk for an SCC above 200,000 cells/mL at 14 d in milk as compared with quarters with a low SCC at drying off and quarters with a negative culture for major pathogens at drying off. For quarters that were culture-positive for major pathogens at drying off, a trend for a higher risk on CM was also found. Selective DCT, not using DCT in cows that had a low SCC at the last milk recording before drying off, significantly increased the incidence rate of CM and SCC. The decrease in antibiotic use by drying off quarters without DCT was not compensated by an increase in antibiotic use for treating CM. Total antibiotic use related to mastitis was reduced by 85% in these quarters.     

Effects of adjuvants on safety and efficacy of an Escherichia coli J5 bacterin

The effects of using a water-soluble adjuvant or an emulsified oil-based adjuvant on the safety, antibody titer, and clinical responses of an Escherichia coli J5 bacterin were tested in an experimental infection trial. Fifty-one cows were assigned to 17 blocks of 3. Two cows within each block of 3 were vaccinated with a commercially prepared E. coli J5 bacterin containing either a water-soluble adjuvant or the same bacterin preparation emulsified in oil. One cow in each block was an unvaccinated control. Cows were immunized at drying off and 42 d later. The right or left front mammary quarter of each experimental cow was challenged by intramammary infusion of E. coli 727 between 14 and 35 DIM. Areas of inflammation at the primary injection site were greater 1, 2, and 3 d following primary vaccination for bacterin containing oil-in-water adjuvant compared with bacterin containing water-soluble adjuvant. Whey anti-E. coli J5 IgG titers were higher at calving for cows vaccinated with bacterin containing oil-in-water adjuvant than for cows either vaccinated with bacterin containing water-soluble adjuvant or unvaccinated controls. Serum x-E. coli J5 IgG titers were higher at calving for vaccinated cows than for unvaccinated controls. Peak bacterial counts in milk from challenged quarters were greater for unvaccinated controls than for cows vaccinated with bacterin containing water-in-oil adjuvant. Bacterial counts in milk from challenged quarters and clinical score both were greater in unvaccinated controls than cows vaccinated with bacterin containing water-in-oil adjuvant between 12 and 24 h postchallenge. Clinical responses were similar between unvaccinated controls and cows vaccinated with bacterin containing water-soluble adjuvant.     

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.     

Bovine intramammary Escherichia coli challenge infections in late gestation demonstrate a dominant antiinflammatory immunological response

Coliform mastitis that presents itself at parturition or in the early weeks of bovine lactation is often characterized by severe inflammation and impaired milk production and can lead to death of the animal. Chronic intramammary infections caused by persistent strains of Escherichia coli may result in high production losses. The aim of this study was to determine the inflammatory response to a teat-canal challenge of bovine mammary glands with a persistent strain of E. coli during late gestation (dry period) and into early lactation. Two weeks before parturition, animals were challenged in 2 quarters with 30 cfu of a persistent strain of E. coli; control quarters were vehicle-infused and not infused, respectively. Samples of dry cow secretions were taken from all quarters before challenge and at 6, 12, 18, 24, 48, 72, 96, and 120 h following challenge. Colostrum samples and milk samples were taken from all quarters at parturition and 6, 12, 18, 24, 48, 72, 96 and 120 h postpartum. Bacterial culture, combined with random amplified polymorphic DNA genetic strain-typing analysis, indicated recovery of the bacterial challenge strain until 48 to 96 h postchallenge, and again at parturition and up to 6 and 12h postpartum. One animal exhibited clinical mastitis and the bacterial challenge strain was evident to at least 12 d postpartum. During twice-daily milkings, production levels were lower in bacteria-challenged quarters compared with controls. Somatic cell counts decreased to normal levels at a slower rate in challenged quarters compared with control quarters. Cytokine analysis indicated a minimal proinflammatory cytokine response, including interleukin-1β and tumor necrosis factor-α in challenged-quarter dry cow samples up to 120 h postchallenge. Interleukin-10 levels were significantly increased by 12h postchallenge in secretions from challenged and control quarters. These preliminary results in 2 cows indicate that proinflammatory signaling after intramammary bacterial infection may be actively suppressed during late gestation. We hypothesize that this immune-inhibitory response allows intramammary infections to become persistent in the dry period and cause clinical signs immediately after parturition.     

The effect of intramammary antibiotic therapy at calving on udder health traits

The effect of intramammary antibiotic therapy at calving on mastitis infection prevalence, linear score milk somatic cell count, and milk NAGase activity, 30 d postpartum, and on milk production, 90 to 120 d postpartum, was tested. Cows (n = 175) were split into treatment and control groups at drying off. All cows received commercial dry cow therapy. At calving, treated cows received commercial lactating cow therapy in all quarters after the first two milkings; control cows were not treated. Composite milk samples were aseptically collected from all cows at drying off, calving, and 30 d postpartum. Udder health traits: linear score milk SCC, NAGase activity, and bacterial content in milk, were determined on all samples. The first three DHI milk weights were recorded for all cows. Treatment and control cows had similar prevalences of intramammary infections during the dry and 30-d postpartum periods. Least squares means of linear score milk SCC and NAGase activities were similar at drying off and calving. Cell count scores were similar between groups; NAGase activities were higher in control cows at 30 d postpartum. Control cows tended to produce more milk postpartum. Results demonstrated no advantage of intramammary therapy at calving in improving milk production or udder health.     

Intramammary challenge with Escherichia coli following immunization with a curli-producing Escherichia coli

Holstein and Jersey cattle were immunized with a curli-producing strain of Escherichia coli (pCRL65/A012) or a noncurli-producing strain (pUC18/HB101) to determine differences in resistance to establishment of experimental intramammary infection. Cows (n = 6 per group) were immunized at 14 d prior to drying off, 7 d of involution, and at calving with 3 x 10(10) E. coli in Freund's Incomplete Adjuvant. At 30 d of lactation, one mammary quarter of each cow was infused with a wild strain of E. coli (727). Escherichia coli 727 was isolated from a naturally occurring intramammary infection and produced curli. All challenged quarters became infected, and all cows developed acute clinical mastitis. Geometric mean duration of intramammary infections was 6 d for both immunization groups. All infections were spontaneously eliminated within 10 d. No differences occurred between immunization groups in blood selenium and glutathione peroxidase activity, plasma selenium, number of E. coli 727 isolated from secretion after challenge, rectal temperature and SCC response, clinical status of mammary quarters, or DMI. Reduction in milk production after challenge was greater for cows immunized with E. coli pCRL65/A012. Immunization of dairy cattle with a curli-producing strain of E. coli did not protect against experimental intramammary challenge during lactation.     

Quarter and cow risk factors associated with the occurrence of clinical mastitis in dairy cows in the United Kingdom

Quarter and cow risk factors associated with the development of clinical mastitis (CM) during lactation were investigated during a 12-mo longitudinal study on 8 commercial Holstein-Friesian dairy farms in the southwest of England. The individual risk factors studied on 1,677 cows included assessments of udder and leg hygiene, teat-end callosity, and hyperkeratosis; body condition score; and measurements of monthly milk quality and yield. Several outcome variables for CM were used for statistical analysis, which included use of generalized linear mixed models. Significant covariates associated with an increased risk of CM were increasing parity, decreasing month of lactation, cows with very dirty udders, and quarters with only very severe hyperkeratosis of the teat-end. Thin and moderate smooth teat-end callosity scores were not associated with an increased risk for CM. Cows that recorded a somatic cell count >199,000 cells/mL and a milk protein percentage <3.2 at the first milk recording after calving were significantly more likely to develop CM after the first 30 d of lactation. There was no association between cow body condition score and incidence of CM. Of the cases of CM available for culture, 171 (26.7%) were confirmed as being caused by Escherichia coli and 121 (18.9%) confirmed as being caused by Streptococcus uberis. Quarters with moderate and very severe hyperkeratosis of the teat-end were at significantly increased risk of clinical E. coli mastitis before the next visit. Quarters with very severe hyperkeratosis of the teat-end were significantly more likely to develop clinical Strep. uberis mastitis before the next visit. There were strong trends within the data to suggest an association between very dirty udders (an increased risk of clinical E. coli mastitis) and teat-ends with no callosity ring present (an increased risk of clinical Strep. uberis mastitis). These results highlight the importance of individual quarter- and cow-level risk factors in determining the risk of CM associated with environmental pathogens during lactation.