NE112: Mastitis Resistance to Enhance Dairy Food Safety

(Multistate Research Project)

Status: Inactive/Terminating

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SAES-422 Reports

Annual/Termination Reports:

[11/10/2000] [02/05/2003] [11/24/2003]

Date of Annual Report: 11/10/2000

Report Information

Annual Meeting Dates: 11/10/2001 - 11/10/2001
Period the Report Covers: 01/01/2000 - 12/01/2000

Participants

Sheila Andrew;
Walt Hurley, Dawn Morin, Univ. Of Illinois;
Bob Harmon, Kentucky;
Ynte Schukken, Cornell;
Jerry Roberson, Virginia;
Jan Sargeant, Kansas State;
Phil Sears, Ron Erskine, Michigan;
Steve Oliver, Tennessee;
Al Guidry, USDA, Maryland;
Larry Fox, Washington STate;
Leo Timms, Iowa State;
Bill Owens;
John Bramley, John Barlow, Vermont;
Dean Kerr;

Brief Summary of Minutes

Accomplishments

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Impact Statements

Date of Annual Report: 02/05/2003

Report Information

Annual Meeting Dates: 11/15/2002 - 11/15/2002
Period the Report Covers: 01/01/2002 - 12/01/2002

Participants

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Accomplishments

COMPLETED RESEARCH Objective 1. Characterize host responses or factors that affect resistance of the mammary gland to mastitis. Illinois. Exposing cows to short days (8 h light:16 h darkness) during the non-lactating period enhanced neutrophil chemotaxis and lymphocyte blastogenesis responses at calving and was associated with a 50% decrease in new intramammary infections during the non-lactating period and a 50% decrease in clinical mastitis incidence in the first 10 days of lactation, compared with long days (16 h light:8 h darkness). However, cows exposed to short days appeared to have a slightly increased local inflammatory response to experimental Streptococcus uberis mastitis. Iowa. A) A collaborative (Guelph, NY, IA, KS) study evaluating cow and quarter factors associated with new dry period was completed and encompassed 300 cows and 1178 quarters. 47% of teats were still open 1 wk post dry off, 24% never closed (herd range from 7-63%), and cracked teat ends and milk production > 21 KG at dry off were associated with higher mastitis risks. B) Three trials evaluating a salve and 2 novel high emollient (>70%) winter teat dips showed no statistical difference in teat end health compared to a commercial non-barrier dip and BMP. C) A six herd winter field study was conducted to evaluate weather related teat end changes and barrier dip usage. There were some significantly higher herd teat scores with the barrier dips, related to improper milking hygiene (failure to remove dip at premilking), and only a few individual cows with severe trauma when exposed to wind chills > 0 F postmilking. D) Overall sensitivity and specificity of CMT during the 1st week postpartum was 68.8 and 71.5%, respectively; with the highest % on day 4 (82.4 and 80.6%) (Guelph, IA, NY, KS). Louisiana. Efficacy of two iodine teat dips and two sodium chloride teat dips was evaluated using the NMC protocol B of guidelines. A barrier teat sealant was evaluated for use as a dry cow product. Michigan. Our preliminary gene expression experiments indicate that steroids of bovine parturition induce an extended life span in neutrophils that become trapped in blood, with concurrent down-regulation of cellular metabolic processes that could otherwise lead to severe systemic inflammation. As such, the parturient blood environment induces an adaptive response in bovine neutrophils that may be critically important to overall animal health. New York. The immunological response to E.coli mastitis was evaluated in both a field study and a challenge study. The results indicate a significant increase in IgG2 antibody titer after challenge both in the natural challenge and the experimental challenge. Ohio. Immunization site reactions were negatively correlated with clinical signs in cows immunized with E. coli J5 and challenged with a heterologous strain of E. coli. Tennessee. A) Interleukin-8 receptor polymorphisms are present at defined frequencies and may be linked to mastitis development. B) Experimental Strep. uberis IMI induced local production of TNF-alpha, IL-1, and IL-8, which may play a role in the pathogenesis of Strep. uberis mastitis. Other mediators may be involved in initial leukocyte recruitment to the mammary gland since increases in milk somatic cells occurred earlier than adhesion molecule expression or cytokine production. C) A cloned calf from a cow susceptible to mastitis exhibited significantly greater percentages of gd-T cells when compared to age-matched controls. Further investigation using cloned animals could result in identification of potential genes, immune components, and/or other factors associated with and responsible for mastitis resistance. Objective 2. Characterization of virulence factors of mammary gland pathogens. Louisiana. Efficacy of prepartum therapy of dairy heifers with a lactating cow product was evaluated as a part of the NE1009 group study. New York. S. uberis infections in dairy cows are increasing. In this project we aim to better understand the differences in pathogenesis of S. uberis strains isolated from cows and from the environment. First a genetic classification system will be developed, and second further studies into the pathogenesis of S. uberis infections will be undertaken. Ohio. Culture of E. coli in medium to induce FecA expression resulted in decreased susceptibility of bacteria to phagocytosis. Tennessee. Strep. uberis avoids intracellular bactericidal mechanisms by exploiting preferential intracellular trafficking pathways in bovine mammary epithelial cells. This strategy would allow Strep. uberis to persist intracellularly and may help explain establishment and persistence of chronic intramammary infections observed in lactating and nonlactating dairy cows. Washington. In a study of the shedding pattern of Mycoplasma sp. in cows with mastitis, mycoplasma was not recovered in 29% of composite samples, while 54% of samples had > 1 million colonies per ml in milk. In 43% of the quarter milk samples mycoplasma was not recovered while in 39% of the samples there were > 1 million colonies per ml. Freezing and prolonged sample storage had a negative impact on organism recovery. Objective 3. Development and evaluation of techniques for modulation of host responses to mastitis pathogens. Illinois. Antimicrobial susceptibility test results, using the disk diffusion method, were not correlated with clinical or bacteriological outcome of clinical mastitis episodes treated with intramammary cephapirin and/or intravenous oxytetracycline. A DNA amplification method was developed for detection of Mycoplasma species in bulk milk and can detect 1,000 cfu/ml at a similar cost to traditional culture. Intramuscular administration of ceftiofur HCl (2.2 mg/kg q 24 hours for 3 treatments) to cows with experimentally induced coliform mastitis was associated with a more rapid return to normal of milk albumin, lactose, sodium, and chloride concentrations, compared with no treatment. Iowa. A) 27 new teat dip sealant prototypes for dry cows were evaluated for persistency, with 3 prototypes showing 40% increased persistency compared to commercially available products. B) Mean % and mean total number of cytokeratin positive cells/ml milk (flow cytometry) before and 12 hrs after endotoxin challenge were 4.18% and 7711, and 40.63% and 2,954,554, respectively. Treatment with steroidal or NSAID did not mitigate these changes. C) Treatment with isoflupredone acetate (20 mg IV 1X) following an endotoxin challenge model and clinical signs resulted in statistically similar milk production as healthy untreated controls. D) A study evaluating prepartum treatment of heifers (single tube of lactating cow antibiotic 8-14 days prepartum) was initiated on a 1200 cow commercial dairy with 110 animals completed through the study. Kansas. A study was completed that determined the reliability of the test procedure that has been documented and used to assess teat canal closure during the dry period. Although the test procedure appeared to be fairly reliable when a single technician used the procedure, reliablity decreased when multiple technicians were employed. These data should be considered for studies employing teat end closure techniques. Louisiana. Various methods of treating teat injuries in lactating cows were described. Michigan. A) Multiple J5 dosing (6+ vaccinations at monthly intervals throughout periparturition and lactation) is needed to elicit significant IgG2 antibody responses in serum and milk. The IgG2 antibodies in hyperimmune serum also interacted with multiple coliform mastitis pathogens in ELISA assays, and recognized a predominant ~ 38 kDa protein in cell membrane proteins not recognized by IgG1. Thus, we may have identified a coliform outer membrane protein that could improve coliform mastitis vaccines. B) We continue to evaluate specific antigens of S. aureus cell wall and capsules for specific activity towards staphyloccal organisms. Several subsets of antigens have been evaluated as a prelude to antibiotic treatment. D) We have evaluated a nisin preparation as an alternative to conventional antibiotic therapy for the treatment of mastitis. Experimental infection models have demonstrated efficacy and safety of intramammary infusions of this product on milk quality and cow health. New York. Chronic coliform mastitis was studied in three large dairy herds. Incidence of cases was evaluated and isolates were obtained. These isolates will be studied in more detailed in the years to come. Ohio. Efficacy of an oil and water adjuvant was greater than water soluble adjuvant in a polyvalent E. coli bacterin. Washington. A study was done to determine the effectiveness of removal of udder and teat hair on new IMI and bacterial counts in milk. Hair removal did not have a significant affect on milk bacterial counts. Thus the results of this study would not support the requirement that udder hair surrounding the teats be removed to reduce IMI and the number of bacteria in milk. However, hair removal might be effective in improving milk quality when less stringent pre- and/or post-milking preparation procedures are used. Objective 4. Characterize dairy food safety issues relevant to bovine mastitis. Connecticut. A) This study evaluated the effect of Ig, milk fat, milk protein, mastitis pathogens and SCC in colostrum and transition milk from Holstein heifers on the specificity rates of several commercially available B-Lactam antibiotic residue-screening tests. Specificity rates ranged from 0.16 to 0.88 for colostrum and 0.60 to 1.0 for transition milk. Increased milk protein and IgG1 concentrations in milk were associated with an increase in the probability of false positive outcomes for the Charm Cowside, CITE Snap and Penzyme tests. Milk should not be tested for antibiotic residues prior to the sixth milking after parturition to avoid high rates of false positive outcomes. B) Fifty multiparous Holstein and Jersey dairy are currently being studied to determine if the combined effects of NEB and ketosis may be important factors in the increased risk of mastitis during early lactation. C) University of Connecticut is 1 of 10 NE112 evaluating the effects of prepartum intramammary antibiotic therapy on the incidence of mastitis and milk production for first lactation heifers throughout a full lactation. Forty heifers, confirmed pregnant and identified as being in good health are currently enrolled in the study. D) A new formulation of a chlorohexidine teat disinfectant is being evaluated as a split herd design to determine if the new formulation is as effective or more effective than a 1% iodine teat disinfectant. New York. In a comparative study, the genome of S. agalactia isolated from cows and from humans was compared. We evaluated a number of classification methods including antimicrobial resistance genes, PCR based methods and sequencing. An important overlap was shown to be present between human and bovine isolates. Further studies are needed to look at the potential importance of this finding. Tennessee. Rhamnose may be useful in differential isolation media for Shiga toxin-producing E. coli (STEC)O26. Presence of shiga toxin 1 gene sequences may aid in rapid identification of E. coli O26. -- About 98% (148 of 151) of E. coli O157 strains (largely serotype H7) were stx+ and almost all elaborated Shiga toxins. Acid-resistance (~96% of STEC) and tellurite-resistance (~86% of STEC) were common characteristics.

Publications

Connecticut Andrew, S. M. 2002. Got Biosecurity? What‘s New in Residue Avoidance. Pages 36-44 In Proc. Natl. Mastitis Counc. 2002. Syracuse NY. Andrew, S. M. 2001. Effect of Composition of Colostrum and Transition Milk from Holstein Heifers on Specificity Rates of Antibiotic Residue Tests. J. Dairy Sci. 84:100-106. Illinois Constable, P.D., and D.E. Morin. 2002. Use of antimicrobial susceptibility testing of bacterial pathogens isolated from the milk of dairy cows with clinical mastitis to predict response to treatment with cephapirin and oxytetracycline. J. Am. Vet. Med. Assoc. 221:103-108. Iowa Dingwell, R.T. , K.E. Leslie, Y.H. Schukken, J.M. Sargent, L.L. Timms, and J. E. Shirley. 2002. Association of cow and quarter level factors at drying off and new intramammary infections during the dry period. Kansas Dairy Day Proceedings. Dingwell, R.T. , K.E. Leslie, Y.H. Schukken, J.M. Sargent, L.L. Timms, and J. Conklin. 2002. Evaluation of the California mastitis test to determine udder health status of early lactation dairy cows. Kansas Dairy Day Proceedings. Hemling, T.C., G.A.Mein, F.Neijenhuis, W.F.Morgan, D.J.Reinemann, J.E.Hillerton, J.R.Baines, I.Ohnstad, M.D.Rasmussen, L.Timms, J.S.Britt, R.Farnsworth & N. Cook (2002) Evaluation of bovine teat condition in commercial dairy herds: 6. Teat condition - prevention and cure through teat dips. Proceedings of the 2nd Panamerican Congress on Milk Quality and Mastitis Control, Ribeiro Preto, Brazil, November 2002, S02-04. Hillerton, J.E., G.A.Mein, F.Neijenhuis, W.F.Morgan, D.J.Reinemann, J.R.Baines, I.Ohnstad, M.D.Rasmussen, L.Timms, J.S.Britt, R.Farnsworth, N.Cook & T. Hemling. 2002. Evaluation of bovine teat condition in commercial dairy herds: 5. Environmental factors. Proceedings of the 2nd Panamerican Congress on Milk Quality and Mastitis Control, Ribeiro Preto, Brazil, November 2002, S01-02, 6pp. Timms, L.L. 2002. Efficacy of barrier teat dips in preventing dry period mastitis. DeLaval Hygiene Technology Center Inaugural International Symposium Proc.:67. Timms, L.L. 2002. Maintaining healthy teats: problems and solutions. Four State Dairy Management Seminar. MWPS-4SD12:17. Timms, Leo. 2002. Winter teat lesions / changes: causes, corrections, and cures. Minnesota Veterinary Medical Assoc. Ann. Conf. Proc.: 6. Kansas Leininger, D.J., J.R. Roberson, and F. Elvinger. 2001. Use of eosin methylene blue agar to differentiate Escherichia coli from other gram-negative mastitis pathogens. Jour. Vet. Diag. Invest. 13:273-275. Leininger, D.J., J.R. Roberson, F. Elvinger, D. Ward, and M. Akers. Efficacy of frequent milk-out for treatment of cows with experimentally induced Escherichia coli mastitis. Accepted for publication July 23, 2002. Roberson, J. R. Establishing treatment protocols for clinical mastitis. Accepted to Veterinary Clinics of North America: Food Animal Practice. Louisianna Boddie, R. L. and S. C. Nickerson. 2002. Reduction of mastitis caused by experimental challenge with Staphylococcus aureus and Streptococcus agalactiae by use of a quaternary ammonium and halogen-mixture teat dip. J. Dairy Sci. 85:258-262. Boddie, R. L., W. E. Owens, C. H. Ray, S. C. Nickerson, and N. T. Boddie. 2002. Germicidal activities of representatives of five different teat dip classes against three bovine Mycoplasma species using a modified excised teat model. J. Dairy Sci. 85:1909-1912. Nickerson, S. C. 2001. Mastitis therapy and control: Management control options. in Encyclopedia of Dairy Sciences. Roginski, H., J. W. Fuquay, and P. F. Fox (eds). Academic Press, London. (In press). Nickerson, S. C. 2001. Mammary resistance mechanisms: Anatomical. in Encyclopedia of Dairy Sciences. Roginski, H., J. W. Fuquay, and P. F. Fox (eds). Academic Press, London. (In press). Nickerson, S. C. 2001. Mammary gland anatomy: Anatomy. in Encyclopedia of Dairy Sciences. Roginski, H., J. W. Fuquay, and P. F. Fox (eds). Academic Press, London. (In press). Nickerson, S. C. 2001. Effective strategies for controlling mastitis in heifers. in Proceedings of the Manitoba Dairy/Forage/Beef Opportunities Conference. Winnipeg, Manitoba, Canada. (In press). Owens, W. E. 2001. Mastitsi therapy and control: Medical therapy options in Encyclopedia of Dairy Sciences. (In press). Owens, W. E. 2001. Microbiological procedures for the diagnosisi of bovine udder infections in Microbiological procedures for the diagnosis of bovine udder infectons. (In press). Owens, W. E. 2001. The Staphylococci in Microbiological procedures for the diagnosis of bovine udder infections. (In press). Owens, W. E., S. C. Nickerson, and C. H. Ray. 2002. Effect of pour-on and fly tag insecticide combination controlling horn flies and Staphylococcus aureus mastitis in dairy heifers. Louisiana Dairy Report. Michigan Publications Burton, J.L. A. Chaiyotwittayakun, K.L. Smith, R. Darch, and R.J. Erskine. 2002. Novel applications for coliform vaccine programs. Proc. 41st Annual Mtg. Natl. Mastitis Council. pp. 89-110. Burton, J.L., and R.J. Erskine. 2002. Mastitis and immunity: some new ideas for an old disease. Veterinary Clinics of North America Food Animal Practice (in press). Burton, J.L., S.A. Madsen, and P.S.D. Weber. 2002. Why are dairy cows susceptible to mastitis around calving? Michigan Dairy Review 7(2):11-13. Burton, J.L., S.A. Madsen, J. Yao, S.S. Sipkovsky, and P.M. Coussens. 2001. An immunogenomics approach to understanding periparturient immunosuppression and mastitis susceptibility in dairy cows. Acta Vet. Scand. 42:407-424. Chaiyotwittayakun, A.C., R.J. Erskine, P.C. Bartlett, T. Herdt, R.. Harmon. 2002. The effect of ascorbic acid and L-Histidine therapy on acute mammary inflammation in dairy cattle. J. Dairy Sci. 85:60-67. Darch, R.A., R.J. Erskine, A.P. Belschner, P.M. Saama, and J.L. Burton. 2002. Lactational Immunization of Dairy Cows With Escherichia coli J5 Bacterin Maintains High Serum Antibody Responses. J. Dairy Sci. (submitted Sept., 2002). Erskine R.J., P.C. Bartlett, J.L. VanLente, C.R. Phipps. 2002. Efficacy of systemic ceftiofur as a therapy for severe clinical mastitis in dairy cattle. J. Dairy Sci. 85:2571-2575. Erskine RJ, R.D.Walker, C.A. Bolin, P.C. Bartlett, and D.G. White. 2002. Trends in antibacterial susceptibility of mastitis pathogens during a seven-year period. J. Dairy Sci. 85:1111-1118. Madsen, S.A., G.J.M. Rosa, P.M. Coussens, and J.L. Burton. 2002. Microarray analysis of mRNA abundance profiles in neutrophils from periparturient cows reveals a novel survival phenotype. Vet. Immunol. Immunopathol. (submitted Nov., 2002). Madsen, S.A., P.S.D. Weber, and J.L. Burton. 2002. Altered blood neutrophil gene expression in periparturient dairy cows. Vet. Immunol. Immunopathol. 86:159-175. Perkins K.H., M.J. VandeHaar, J.L. Burton, J.S. Liesman, R.J. Erskine, and T.H. Elsasser. 2002 Clinical responses to intramammary endotoxin infusion in dairy cows subjected to feed restriction. J Dairy Sci 85:1724-1731. Perkins, K.H., M. J. VandeHaar, R. J. Tempelman, and J. L. Burton. 2001. Negative energy balance does not decrease expression of leukocyte adhesion or antigen presenting molecules in cattle. J. Dairy Sci. 84:421-428. Tempelman, R.J., P.M. Saama, A.E. Freeman, S.C. Kelm, A.L. Kuck, M.E. Kehrli, Jr., and J.L. Burton. 2002. Genetic variation in bovine neutrophil sensitivity to glucocorticoid challenge. Acta Agric. Scand. (in press). Weber, P.S.D., S.A. Madsen, G.W. Smith, J.J. Ireland, and J.L. Burton. 2001. Pre-translational regulation of neutrophil CD62L in glucocorticoid-challenged cattle. Vet. Immunol. Immunopathol. 83:213-240. Yao, J., J.L. Burton, P. Saama, S. Sipkovsky, and P.M. Coussens. 2001. Generation of EST and cDNA Microarray Resources for the Study of Bovine Immunobiology. Acta Vet. Scand. 42:391-405. Outreach Erskine R.J. J-5 vaccines and E. coli mastitis: Efficacy and economic realties. American Association of Bovine Practitioners, Madison, WI; Sept, 2002. Erskine R.J. Mastitis therapy update. Ontario Veterinary College Short Course for Dairy Herd Health Certificate Program, Guelph, Ont; May, 2002. Erskine R.J. and P.M. Sears. Update on mastitis therapy and applied pharmacology. Short course for Dairy Herd Management Certificate Program Alumni, East Lansing, MI; March, 2002. Erskine R.J., M. Masterson, D. Morin, A. Godkin. Seminar on mastitis therapy. Short course presented at the American Association of Bovine Practitioners, Rapid City, S.D. (2000), Vancouver, B.C. (2001), Madision, WI. (2002). New York Rivas AL, Quimby FW, Coksaygan O, Alba A, Arina A, Arrobas MJ, Gonzalez RN, Mohammed HO, Lein DH. Expression of CD3 and CD11b antigens on blood and mammary gland leukocytes and bacterial survival in milk of cows with experimentally induced Staphylococcus aureus mastitis. 2001. Am J Vet Res. 62:1840-1851 van Schaik G, Lotem M, Schukken YH. Trends in somatic cell counts, bacterial counts, and antibiotic residue violations in New York State during 1999-2000. 2002. J Dairy Sci. 85:782-789. van Schaik G, Schukken YH, Nielen M, Dijkhuizen AA, Barkema HW, Benedictus G. Probability of and risk factors for introduction of infectious diseases into Dutch SPF dairy farms: a cohort study. 2002. Prev Vet Med. 54:279-289. White LJ, Evans ND, Lam TJ, Schukken YH, Medley GF, Godfrey KR, Chappell MJ. The structural identifiability and parameter estimation of a multispecies model for the transmission of mastitis in dairy cows with postmilking teat disinfection. 2002 Math Biosci. 180:275-281. White LJ, Evans ND, Lam TJ, Schukken YH, Medley GF, Godfrey KR, Chappell MJ. The structural identifiability and parameter estimation of a multispecies model for the transmission of mastitis in dairy cows. 2001 Math Biosci. 174:77-90. White LJ, Schukken YH, Lam TJ, Medley GF, Chappell MJ. A multispecies model for the transmission and control of mastitis in dairy cows. 2001. Epidemiol Infect. 127:567-576. Zadoks RN, Allore HG, Hagenaars TJ, Barkema HW, Schukken YH. A mathematical model of Staphylococcus aureus control in dairy herds. 2002 Epidemiol Infect 129:397-416. Zadoks RN, Van Leeuwen WB, Kreft D, Fox LK, Barkema HW, Schukken YH, Van Belkum A. Comparison of Staphylococcus aureus Isolates from Bovine and Human Skin, Milking Equipment, and Bovine Milk by Phage Typing, Pulsed-Field Gel Electrophoresis, and Binary Typing. 2002. J Clin Microbiol. 40:3894-3902. Ohio Takemura, K., J.S. Hogan, J. Lin, and K.L. Smith. 2002. Efficacy of immunization with ferric citrate receptor FecA from Escherichia coli on induced coliform mastitis. J. Dairy Sci. 85:774-781. Wise, A. J., J. S. Hogan, V. B. Cannon, and K. L. Smith. 2002. Phagocytosis and serum susceptibility of Escherichia coli cultured in iron-deplete and iron-replete media. J. Dairy Sci. 85:1454-1459. Tennessee Journal Articles Almeida, R. A., and S. P. Oliver. 2001. Interaction of coagulase-negative Staphylococcus species with bovine mammary epithelial cells under in vitro conditions. Microbial Pathogenesis, 31:205-212. Almeida, R. A., and S. P. Oliver. 2001. Role of collagen in adherence of Streptococcus uberis to bovine mammary epithelial cells. J. Vet. Med. B 48:759-763. Brown-Crutchfield, M. E., H. G. Kattesh, S. P. Oliver, and J. F. Schneider. 2002. Mitogen-induced lymphocyte blastogenic response in piglets weaned at different ages. J. Animal and Veterinary Advances, 1:127-128. Gillespie, B. E., H. Moorehead, H. H. Dowlen, D. L. Johnson, K. C. Lamar, M. J. Lewis, S. J. Ivey, and S. P. Oliver. 2002. Efficacy of extended pirlimycin therapy for treatment of chronic environmental Streptococcus species intramammary infections in lactating dairy cows. Veterinary Therapeutics, Accepted. Murinda, S. E., L. T. Nguyen, S. J. Ivey, B. E. Gillespie, R. A. Almeida, and S. P. Oliver. 2002. Prevalence and molecular characterization of Escherichia coli O157:H7 in bulk tank milk and fecal samples from cull dairy cows: a 12-month survey of dairy farms in East Tennessee. J. Food Prot. 65:752-759. Murinda, S. E., L. T. Nguyen, S. J. Ivey, B. E. Gillespie, R. A. Almeida, F. A. Draughon, and S. P. Oliver. 2002. Molecular characterization of Salmonella spp. isolated from bulk tank milk and cull dairy cow fecal samples. J. Food Prot. 65:1100-1105. Murinda, S. E., L. T. Nguyen, S. J. Ivey, R. A. Almeida, and S. P. Oliver. 2002. Development of a novel single tube agar-based test system for motility enhancement and immunocapture of Escherichia coli O157:H7 by H7 flagellar-specific antibodies. J. Clin. Microbiol. Accepted. Oliver, S. P., M. J. Lewis, B. E. Gillespie, H. H. Dowlen E. C. Jaenicke, and R. K. Roberts. 2002. Milk production, milk quality and economic benefit associated with prepartum antibiotic treatment of heifers. J. Dairy Sci. Accepted. Park, Hee-Myung, R. A. Almeida, and S. P. Oliver. 2002. Identification of lactoferrin-binding proteins in Streptococcus dysgalactiae subsp. dysgalactiae and Streptococcus agalactiae isolated from cows with mastitis. FEMS Microbiol. Lett. 207:87-90. Park, Hee-Myung, R. A. Almeida, D. A. Luther, and S. P. Oliver. 2002. Binding of bovine lactoferrin to Streptococcus dysgalactiae subsp. dysgalactiae isolated from cows with mastitis. FEMS Microbiol. Lett. 208:35-39. Book Chapters Oliver, S. P. and G. M. Pighetti. 2002. Mastitis Pathogens (b) Environmental pathogens. In: Encyclopedia of Dairy Science, Hubert Roginski, Patrick F. Fox and John W. Fuquay, Eds., Academic Press, London. Proceedings Almeida, R. A. L. F. Calvinho, D. Dopfer, and S. P. Oliver. 2001. Internalization of mastitis pathogens: A bacterial strategy to survive and cause intramammary infection. In: Proc. 2nd Intl. Mastitis and Milk Quality Symposium, pp. 426-427. Almeida, R. A. L. F. Calvinho, D. Dopfer, and S. P. Oliver. 2001. Adherence and internalization of mastitis pathogens: bacterial strategies to survive and cause intramammary infection. In: Department of Animal Science Annual Report, The University of Tennessee, Knoxville. http://www.agriculture.utk.edu/ansci.annualreports. htm#AnimalHealth. Almeida, R. A. L. F. Calvinho, D. Dopfer, and S. P. Oliver. 2002. Internalization of mastitis pathogens: A bacterial strategy to survive and cause intramammary infection. In: Proc. Natl. Mastitis Counc. pp. 235-236. Almeida, R. A., and S. P. Oliver. 2001. Adherence strategies of Streptococcus uberis. In: Proc. 2nd Intl. Mastitis and Milk Quality Symposium, pp. 424-425. Almeida, R. A., and S. P. Oliver. 2002. Adherence strategies of Streptococcus uberis. In: Proc. Natl. Mastitis Counc. pp. 237-238. Almeida, R. A., and S. P. Oliver. 2002. Virulence factors of Streptococcus uberis and their relationship with the pathogenesis of bovine mastitis. In: Proc. Korean Association for Buiatrics, 7:105-142. Almeida, R. A., S. E. Murinda, and S. P. Oliver. 2002. Identification and validation of critical control points during the harvesting of milk. In: Proc. Intl. Workshop on the Safety of Foods of Animal Origin, Zhejiang University, Hanghou, China, pp. 9-10. Gillespie, B. E., and S. P. Oliver. 2001. DNA fingerprinting of mastitis pathogens: Useful tools for bacterial identification and epidemiological evaluation. In: Proc. 2nd Intl. Mastitis and Milk Gillespie, B. E., B. M. Jayarao, A. G. Mathew, F. A. Draughon, and S.P. Oliver. 2001. PCR-ELISA procedure for Salmonella serovars A, B, C1, C2 and D. In: Proc. 2nd Intl. Mastitis and Milk Quality Symposium, pp. 460-461. Gillespie, B. E., B. M. Jayarao, A. G. Mathew, F. A. Draughon, and S. P. Oliver. 2001. PCR-ELISA procedure for identification of Salmonella serovars A, B, C1, C2 and D. In: Department of Animal Science Annual Report, The University of Tennessee, Knoxville. http://www.agriculture.utk.edu/ansci.annual reports.htm#AnimalHealth. Gillespie, B. E., H. Moorehead, H. H. Dowlen, D. L. Johnson, K. C. Lamar, M. J. Lewis, S. J. Ivey, J. W. Hallberg, S. T. Chester, and S. P. Oliver. 2001. Efficacy of extended pirlimycin therapy for treatment of chronic intramammary infections in lactating dairy cows. In: Proc. 2nd Intl. Mastitis and Milk Quality Symposium, pp. 458-459. Gillespie, B. E., H. Moorehead, H. H. Dowlen, D. L. Johnson, K. C. Lamar, M. J. Lewis, S. J. Ivey, J. W. Hallberg, S. T. Chester, and S. P. Oliver. 2002. Efficacy of extended pirlimycin therapy for treatment of chronic intramammary infections in lactating dairy cows. In: Proc. Natl. Mastitis Counc. pp. 207-208. Hockett, M. E., F. N. Schrick, R. A. Almeida, and S. P. Oliver. 2001. Endocrine and ovarian alterations during experimentally-induced mastitis. In: Proc. 2nd Intl. Mastitis and Milk Quality Symposium, pp. 203-207. Hockett, M. E., F. N. Schrick, R. A. Almeida, and S. P. Oliver. 2001. Endocrine and ovarian alterations during experimentally-induced Streptococcus uberis mastitis. In: Department of Animal Science Annual Report, The University of Tennessee, Knoxville. http://www.agriculture.utk.edu/ansci.annualreports.htm#AnimalHealth. Murinda, S. E., L. T. Nguyen, S. J. Ivey, R. A. Almeida, F. A. Draughon, and S. P. Oliver. 2001. Prevalence of Salmonella species in bulk tank milk and fecal samples from cull dairy cows. In: Proc. 2nd Intl. Mastitis and Milk Quality Symposium, pp. 498-499. Murinda, S. E., L. T. Nguyen, S. J. Ivey, R. A. Almeida, F. A. Draughon, and S. P. Oliver. 2002. Prevalence of Salmonella species in bulk tank milk and fecal samples from cull dairy cows. In: Proc. Natl. Mastitis Counc. pp. 174-175. Murinda, S. E., L. T. Nguyen, S. J. Ivey, R. A. Almeida, F. A. Draughon, and S. P. Oliver. 2001. Evaluation of antimicrobial agents for inhibiting fungal growth on agar media for isolation of Campylobacter in fecal samples from cull dairy cows. In: Proc. 2nd Intl. Mastitis and Milk Quality Symposium, pp. 494-495. Murinda, S. E., L. T. Nguyen, S. J. Ivey, R. A. Almeida, M. J. Montgomery, F. A. Draughon, and S. P. Oliver. 2001. Isolation of mastitis and foodborne pathogens in bulk tank milk and fecal samples from cull dairy cows. In: Proc. 2nd Intl. Mastitis and Milk Quality Symposium, pp. 496-497. Murinda, S. E., L. T. Nguyen, S. J. Ivey, R. A. Almeida, M. J. Montgomery, F. A. Draughon, and S. P. Oliver. 2002. Isolation of mastitis and foodborne pathogens in bulk tank milk and fecal samples from cull dairy cows. In: Proc. Natl. Mastitis Counc. pp. 178-179. Murinda, S. E., L. T. Nguyen, S. J. Ivey, R. A. Almeida, B. E. Gillespie, M. J. Montgomery, F. A. Draughon, and S. P. Oliver. 2001. Isolation of mastitis and foodborne pathogens in bulk tank milk and fecal samples from cull dairy cows. In: Department of Animal Science Annual Report, The University of Tennessee, Knoxville. http://www.agriculture.utk.edu/ansci.annualreports. htm#AnimalHealth. Murinda, S. E., L. T. Nguyen, S. J. Ivey, R. A. Almeida, F. A. Draughon, and S. P. Oliver. 2001. Prevalence of Escherichia coli O157:H7 in bulk tank milk and fecal samples from cull dairy cows. In: Proc. 2nd Intl. Mastitis and Milk Quality Symposium, pp. 500-501. Murinda, S. E., L. T. Nguyen, S. J. Ivey, R. A. Almeida, F. A. Draughon, and S. P. Oliver. 2002. Prevalence of Escherichia coli O157:H7 in bulk tank milk and fecal samples from cull dairy cows. In: Proc. Natl. Mastitis Counc. pp. 176-177. Oliver, S. P. 2001. Mastitis control during the dry period: a brief review and future prospects. In: Proc. Southeast Dairy Herd Management Conference, pp. 81-91. Oliver, S. P., and F. A. Draughon. 2001. Establishment of The University of Tennessee Food Safety Center of Excellence. In: Department of Animal Science Annual Report, The University of Tennessee, Knoxville. http://www.agriculture.utk.edu/ansci.annualreports.htm#AnimalHealth. Oliver, S. P., and F. A. Draughon. 2002. Food safety research and educational programs at The University of Tennessee Food Safety Center of Excellence. In: Proc. Intl. Workshop on the Safety of Foods of Animal Origin, Zhejiang University, Hanghou, China, pp. 5-6. Oliver, S. P., and R. A. Almeida. 2002. Mastitis control, food safety and quality milk production. In: Proc. Korean Association for Buiatrics, 7:67-104. Oliver, S. P., B. E. Gillespie, M. J. Lewis, E. C. Jaenicke, R. K. Roberts, and H. H. Dowlen. 2001. An effective strategy for the control of mastitis in heifers. In: Proc. 2nd Intl. Mastitis and Milk Quality Symposium, pp. 506-507. Oliver, S. P., B. E. Gillespie, M. J. Lewis, E. C. Jaenicke, R. K. Roberts, and H. H. Dowlen. 2001. An effective strategy for the control of heifer mastitis. In: Department of Animal Science Annual Report, The University of Tennessee, Knoxville. http://www.agriculture.utk.edu/ansci.annualreports.htm#AnimalHealth. Oliver, S. P., F. N. Schrick, M. E. Hockett, and H. H. Dowlen. 2001. Mastitis during early lactation impairs reproductive performance of dairy cows. In: Proc. Southeast Dairy Herd Management Conference,. pp. 47-64. Oliver, S. P., M. J. Lewis, B. E. Gillespie, and H. H. Dowlen. 2002. Preventing and controlling heifer mastitis. In: Proc. Professional Dairy Heifer Grower‘s Association. pp. 100-112. Oliver, S. P., R. A. Almeida, and S. E. Murinda. 2002. Mastitis control, food safety and quality milk production. In: Proc. Intl. Workshop on the Safety of Foods of Animal Origin, Zhejiang University, Hanghou, China, pp. 18. Park, H. M., R. A. Almeida, and S. P. Oliver. 2001. Identification of lactoferrin-binding proteins in Streptococcus dysgalactiae isolated from mastitic milk. In: Proc. 2nd Intl. Mastitis and Milk Quality Symposium, pp. 508-509. Park, H. M., R. A. Almeida, D. A. Luther, and S. P. Oliver. 2001. Identification and characterization of lactoferrin-binding proteins in Streptococcus species isolated from cows with mastitis. In: Department of Animal Science Annual Report, The University of Tennessee, Knoxville. http://www.agriculture.utk.edu/ansci.annualreports.htm#AnimalHealth. Quality Symposium, pp. 456-457. Schrick, F. N., M. E. Hockett, A. M. Saxton, M. J. Lewis, H. H. Dowlen, and S. P. Oliver. 2001. Influence of subclinical mastitis during early lactation on reproductive parameters. In: Department of Animal Science Annual Report, The University of Tennessee, Knoxville. http://www.agriculture.utk.edu/ansci. annualreports.htm# AnimalHealth. Schrick, F. N., M. E. Hockett, H. H. Dowlen, M. J. Lewis, and S. P. Oliver. 2001. Association of mastitis with reproductive failure in dairy cows. In: Proc. 2nd Intl. Mastitis and Milk Quality Symposium, pp. 526-527. Schrick, F. N., M. E. Hockett, H. H. Dowlen, M. J. Lewis, and S. P. Oliver. 2002. Association of mastitis with reproductive failure in dairy cows. In: Proc. Natl. Mastitis Counc. pp. 170-171. Zadoks, R. N., B. E. Gillespie, H. W. Barkema, O. C. Sampimon, Y. H. Schukken, and S. P. Oliver. 2001. Clinical, epidemiological and molecular characteristics of Streptococcus uberis. In: Proc. 2nd Intl. Mastitis and Milk Quality Symposium, pp. 83-87. Abstracts Almeida, R. A., and S. P. Oliver. 2002. Trafficking of Streptococcus uberis in bovine mammary epithelial cells. Proceedings of the Conference of Research Workers in Animal Diseases (In press). Batson, S. D., S. E. Murinda, L.T. Nguyen, and S. P. Oliver. 2002. Phenotypic and genetic characterization of Escherichia coli O26 isolates from North America using carbohydrate fermentation and genes encoding virulence factors. Proceedings of the Conference of Research Workers in Animal Diseases (In press). Ebner, P. D., S. E. Murinda, B. E. Gillespie, S. P. Oliver, and A. G. Mathew. 2002. Virulence attributes of E. coli O157:H7 isolated from dairies in East Tennessee. Intl. Assoc. Food Prot., Abstract PO. Evans, M. R., F. A. Draughon, S. P. Oliver and V. Ling. 2002. Genotypic diversity of Listeria isolates from dairy cows, calves and the farm environment. 89th annual meeting of the Intl. Assoc. Food Prot., Abstract PO75. Hockett, M. E., N. R. Rohrbach, R. A. Almeida, H. Dowlen, K. Lamar, S. P. Oliver, and F. N. Schrick. 2002. Effects of experimentally-induced clinical mastitis on estrous behavior, luteal function, and establishment of pregnancy during early lactation of dairy cows. J. Anim. Sci. 80(1)/J. Dairy Sci. 85(1): 264. Hockett, M. E., N. R. Rohrbach, R. A. Almeida, H. E. Blackmon, F. Scenna, S. P. Oliver, and F.N. Schrick. 2002. Effects of experimentally-induced clinical mastitis during the preovulatory period on endocrine function, follicular growth and ovulation of lactating dairy cows. J. Anim. Sci. 80(1)/J. Dairy Sci. 85(1): 265. Lamar, L. D., M. Evans, V. Ling, S. P. Oliver, D. A. Golden and F. A. Draughon. 2002. Geographic Information System (GIS) and Listeria isolates recovered from dairy cows, calves and farm environments. Intl. Assoc. Food Prot., Abstract PO70 (Note: Ms. Lamar won 1st place in the Developing Scientist Competition). Murinda, S. E., L. T. Nguyen, T. L. Landers, F. A. Draughon, A. G. Mathew, J. S. Hogan, K. L. Smith, D. D. Hancock, and S. P. Oliver. 2002. Multiplex PCR for detection and confirmation of Shiga-toxin producing Escherichia coli. Proceedings of the Conference of Research Workers in Animal Diseases (In press). Pighetti, G.M., S.M. Youngerman, and S.P. Oliver. 2002. A novel and promising candidate gene for mastitis resistance. Proceedings of the Conference of Research Workers in Animal Diseases. In press. Rambeaud, M., G. M. Pighetti, and S. P. Oliver. 2002. Dynamics of leukocytes and cytokines during experimentally-induced Streptococcus uberis mastitis. Proceedings of the Conference of Research Workers in Animal Diseases (In press). Youngerman, S. M., J. L. Edwards, F. N. Schrick, M. van Amstel, H. H. Dowlen, S. P. Oliver, and G. M. Pighetti. 2002. Leukocyte phenotypes expressed by a calf cloned from a mastitis susceptible dairy cow. Proceedings of the Conference of Research Workers in Animal Diseases (In press). Popular Press Articles Oliver, S. P. 2001. Heifer mastitis: strategies for control and economic implications. Hoardms Dairymen West. Oliver, S. P. 2002. Dairy food safety starts on the farm. Dairy Herd Management. August Issue 2002, p. 18. Washington Ebling, T.L., L.K. Fox, K.W. Bayles, G. A. Bohach, K.M. Byrne, W.C. Davis, and W.A. Ferens. 2001. Bovine mammary immune response to an experimental intramammry infection with a Staphylococcus aureus strain containing a gene for staphylococcal enterotoxin C1. J. Dairy Sci. 84:2044-2050. Joo, Y. S., L. K. Fox, W. C. Davis, G. A. Bohach, and Y. H. Park. 2001. Staphylococcus aureus associated with mammary glands of cows: Genotyping to distinguish different strains among herds. Vet Micro 80:131-138. Middleton J.R., and L.K. Fox. 2002. Influence of Staphylococcus aureus strain on May; mammary quarter milk production. Vet Rec. 150(13):411-412. Middleton J.R., L.K. Fox, and T.H. Smith. 2001. Management strategies to decrease the prevalence of mastitis caused by one strain of Staphylococcus aureus in a dairy herd. JAVMA. 218:1615-1618. Middleton J.R., L.K. Fox, J.M. Gay, J.W. Tyler, and T.E. Besser. 2002. Use of pulsed-field gel electrophoresis for detecting differences in Staphylococcus aureus strain populations between dairy herds with different cattle importation practices. Epidemiol. Infect. 129:387-395. Middleton J.R., L.K.Fox, J.M. Gay, J.W. Tyler, and T.E. Besser. 2002. Influence of Staphylococcus aureus strain-type on mammary quarter milk somatic cell count and N-acetyl-beta-D-glucosaminidase activity in cattle from eight dairies. J. Dairy Sci. 85:1133-40. Middleton JR, L.K. Fox, and T.H. Smith. 2001. Management strategies to decrease the prevalence of mastitis caused by one strain of Staphylococcus aureus in a dairy herd. JAVMA. 218:1615-1618. Middleton, J. R., and L. K. Fox. 2001. Therapeutic cessation of lactation of Staphylococcus aureus-infected mammary quarters. J. Dairy Sci., 84:1976-1978. Oura, L.Y., L.K. Fox, C.C. Warf, and G. K. Kemp. 2002. Efficacy of two acidified chlorite postmilking teat disinfectants with sodium dodecylbenzene sulfonic acid on prevention of contagious mastitis using an experimental challenge protocol. J. Dairy Sci. 85:252-257. Zadoks, R. N., W. B. van Leeuwen, D. Kreft, L. K. Fox, H. W. Barkema, Y. H. Schukken, and A. van Belkum. 2002. Comparison of Staphylococcus aureus isolates from bovine and human skin, milking equipment, and bovine milk by phage typing, pulsed-field gel electrophoresis, and binary typing. J. Clin. Microbiol. 40:3894-3902.

Impact Statements

Substantial savings to the producer resulting from an improved understanding how cows resist and bacteria cause disease.
Improved milk quality and safety as a result of enhanced understanding of the impacts of current therapies and prevention strategies.
Potential identification of new strategies or therapies against mastitis by identifying specific genes/proteins of cows and pathogenic organisms that protect against or cause mastitis.

Date of Annual Report: 11/24/2003

Report Information

Annual Meeting Dates: 11/13/2002 - 11/15/2002
Period the Report Covers: 10/01/1997 - 10/01/2002

Participants

Andrew, Sheila (University of Connecticut);Barkema, Herman (University Prince Edward Island);Barlow, John (University of Vermont);Burton, Jeanne (Michigan State University);Cullor, Jim (UC Davis);Dingwell, Randy (University of Prince Edward Island);Erskine, Ron (Michigan State University);Fox, Larry (Washington State University);Godden, Sandra (University of Minnesota);Guidry, Al (USDA-Beltsville);Harmon, Bob (University of Kentucky);Hogan, Joe (Ohio State University);Hurley, Walt (University of Illinois);Kelton, David (University of Guelph);Kerr, David (University of Vermont);Kerr, Kirklyn (University of Connecticut);Leslie, Ken (University of Guelph);Madsen, Sally (Michigan State Uiversity);Middleton, John (Washington State University);Morin, Dawn (University of Illinois);Oliver, Steve (University of Tennessee);Owens, Bill (Louisiana State University);Pighetti, Gina (University of Tennessee);Roberson, Jerry (Kansas State University);Ruegg, Pam (University of Wisconsin);Sargeant, Jan (Kansas State University);Schukken, Ynte (Cornell);
Schultz, Bruce (Kansas State University);Sears, Phil (Michigan State University);Sordillo, Lorraine (PSU);Timms, Leo (Iowa State University);van Dorp, Renate (Michigan State University);Wenz, John (Colorado State University)

Brief Summary of Minutes

Accomplishments

Objective 1. Characterize host responses or factors that affect resistance of the mammary gland to mastitis. IA, KS, Cornell, Guelph. Overall, 11% of quarters developed new IMI in the dry period. The majority of new IMI were caused by environmental streptococci and coliform organisms (34% and 30%, respectively). The proportions of new IMI were different among herds, parity levels, and season of drying-off. Quarters with cracked teat-ends were more likely to develop new infections than those without cracks (15% and 10%, respectively). Quarters that closed, through formation of a keratin plug early in the dry period, were significantly less likely to develop a new infection, compared to quarters that did not close (10% and 14%, respectively). At the end of the six-week dry period, 23% of quarters were still classified as open. The risk of quarters closing was significantly associated with the level of milk production on the day before drying-off. Management strategies aimed at reducing milk production, improving teat integrity, and enhancing closure of the teat streak canal should be evaluated and implemented. IL. Exposing cows to short days (8 hr light, 16 hr darkness) during the dry period enhanced neutrophil chemotaxis and lymphocyte blastogenesis at calving and was associated with a 50% decrease in new intramammary infections during the dry period and a 50% decrease in clinical mastitis incidence the first 10 days of lactation, compared with long days (16 hr light, 8 hr darkness). However, cows exposed to short days appeared to have a slightly increased local inflammatory response to experimental Streptococcus uberis mastitis. IA. Three trials evaluating a salve and 2 high emollient (>70%) winter teat dips showed no statistical difference in teat end health compared to a commercial non-barrier dip and BMP. LA. Efficacy of 2 iodine teat dips and 2 sodium chloride teat dips was evaluated using NMC protocol B guidelines. A barrier teat sealant was evaluated for use as a dry cow product. MI. Over 310 genes in bovine neutrophils respond to parturition with changed expression. For ~ 60% of these genes, expression levels were dramatically increased, especially the first 12 hr following parturition. Remaining gene expression changes included pronounced down-regulation at parturition or for at least one day postpartum. The majority of these changes suggested parturition induced survival of blood neutrophils. This hypothesis is supported by pronounced neutrophilia observed for ~48 hr around parturition and survival of neutrophils from parturient cows ~24 hr longer in culture than those from mid-lactation cows. Preliminary experiments suggest that steroids of bovine parturition induce this survival with concurrent down-regulation of cellular metabolic processes that could otherwise lead to severe systemic inflammation. Given these findings, we preliminarily conclude that the parturient blood environment induces an adaptive response in bovine neutrophils that may be critically important to overall animal health. NY. The immune response to E.coli mastitis was evaluated in both a field and challenge study. The results indicate a significant increase in IgG2 antibody titer after challenge both in the natural challenge and the experimental challenge. OH. Immunization site reactions were negatively correlated with clinical signs in cows immunized with E. coli J5 and challenged with a heterologous strain of E. coli. TN. A Interleukin-8 receptor single nucleotide polymorphisms are present at defined frequencies and may be linked to mastitis development. B Experimental Strep. uberis IMI induced local production of TNF-alpha, IL-1, and IL-8, which may play a role in the pathogenesis of Strep. uberis mastitis. Other mediators may be involved in initial leukocyte recruitment to the mammary gland since increases in milk somatic cells occurred earlier than adhesion molecule expression or cytokine production. C A cloned calf from a cow susceptible to mastitis exhibited a significantly greater percentage of gd-T cells overall when compared to age-matched controls. These values also were within the upper range expressed by all calves when evaluated on a single day. Further investigation using cloned animals could result in identification of potential genes, immune components, and/or other factors associated with mastitis resistance. Objective 2. Characterization of virulence factors of mammary gland pathogens. NY. S. uberis infections in dairy cows are increasing. In this project we aim to better understand the differences in pathogenesis of S. uberis strains isolated from cows and the environment. First a genetic classification system will be developed, and second further studies into the pathogenesis of S. uberis infections will be undertaken. OH. Culture of E. coli in medium to induce FecA expression resulted in decreased susceptibility of bacteria to phagocytosis. TN. Strep. uberis avoids intracellular bactericidal mechanisms by exploiting preferential intracellular trafficking pathways in bovine mammary epithelial cells. This strategy would allow Strep. uberis to persist intracellularly and may help explain establishment and persistence of chronic intramammary infections observed in lactating and nonlactating dairy cows. WA. In a study of the shedding pattern of Mycoplasma sp. in cows with mastitis, mycoplasma was not recovered in 29% of composite samples, while 54% of the samples had more than 1 million colonies per ml in milk. In 43% of the quarter milk samples mycoplasma was not recovered while in 39% of the samples there were greater than 1 million colonies per ml. On effects of storage and sample handling, freezing and prolonged storage had a negative impact on organism recovery. Objective 3. Development and evaluation of techniques for modulation of host responses to mastitis pathogens. WA, LA, TN, OH, Cornell, Guelph, CT, KY. A greater number of intramammary infections in heifers were cured by treatment during the peripartum period. As a result, treated cows had less IMI at calving, and also less new IMI at calving. IMI were reduced in all herds by treatment, and CNS were the pathogens accounting for the vast majority of treatable IMI. Postpartum milk production was equivalent in treated and control heifers, and milk somatic cell count slightly lower in treated heifers. IL. Antimicrobial susceptibility test results, using the disk diffusion method, were not correlated with clinical or bacteriological outcome of clinical mastitis episodes treated with intramammary cephapirin and/or intravenous oxytetracycline. A DNA amplification method was developed for detection of Mycoplasma species in bulk milk and can detect 1,000 cfu/ml at a similar cost to traditional culture. Intramuscular administration of ceftiofur HCl (2.2 mg/kg q 24 hr for 3 treatments) to cows with experimentally induced coliform mastitis was associated with a more rapid return to normal of milk albumin, lactose, sodium, and chloride concentrations, compared with no treatment. IA. A 27 new teat dip sealant prototypes for dry cows were evaluated for persistency, with 3 prototypes showing 40% increased persistency compared to commercially available products. B Mean % and mean total number of cytokeratin positive cells/ml milk before and 12 hr after endotoxin challenge were 4.18% and 7711, and 40.63% and 2,954,554, respectively. Treatment with steroidal or NSAID did not mitigate these changes. C Treatment with isoflupredone acetate (20 mg IV 1X) following an endotoxin challenge model and clinical signs resulted in statistically similar milk production as healthy untreated controls. KS. A study was conducted that determined the reliability of a documented test procedure to assess teat canal closure during the dry period. Although fairly reliable when a single technician used the procedure, reliability decreased when multiple technicians were employed. These data should be considered for studies employing teat end closure techniques. LA. Various methods of treating teat injuries in lactating cows were described. MI. A The current 3-dose J-5 vaccination regime increases serum and milk anti-coliform IgG1, but is relatively short-lived and does not promote opsonic phagocytosis of E. coli by bovine neutrophils. In contrast, at least 6 vaccinations given monthly throughout periparturition and lactation elicited significant anti-coliform IgG2 responses in serum and milk. IgG2 antibodies also interacted with multiple coliform mastitis pathogens in ELISA assays and recognized a predominant ~ 38 kDa protein in cell membrane proteins. Thus, we may have identified a coliform outer membrane protein that could be used to improve coliform mastitis vaccines, because opsonizing IgG2 antibodies that recognize this protein also promote effective phagocytosis of E. coli by neutrophils. B We continue to evaluate specific antigens of S. aureus cell wall and capsules for specific activity towards staphylococcal organisms. Several subsets of antigens have been evaluated as a prelude to antibiotic treatment. C Experimental infection models of streptococcal mastitis have demonstrated efficacy of intramammary infusions of a nisin preparation. Additionally, safety studies were completed that suggested this product will not affect milk quality or cow health. NY. Chronic coliform mastitis was studied in three large dairy herds. Incidence of cases was evaluated and isolates were obtained. These isolates will be studied in more detailed in the years to come. OH. Efficacy of an oil and water adjuvant was greater than water soluble adjuvant in a polyvalent E. coli bacterin. WA. A study was done to determine the effectiveness of removal of udder and teat hair on new IMI and bacterial counts in milk. Hair removal did not have a significant affect on milk bacterial counts. However, hair removal might be effective in improving milk quality when less stringent pre- and/or post-milking preparation procedures are used. Objective 4. Characterize dairy food safety issues relevant to bovine mastitis. CT. A The objective of this study was to evaluate the effect of antibodies, milk fat, milk protein, mastitis pathogens and SCC in colostrum and transition milk from Holstein heifers on the specificity rates of several commercially available B-Lactam antibiotic residue screening tests. Specificity rates ranged from 0.16 to 0.88 for colostrum and 0.60 to 1.0 for transition milk. Increased milk protein and IgG1 concentrations in milk were associated with increased probability of false positive outcomes for the Charm Cowside, CITE Snap and Penzyme tests. Milk should not be tested for antibiotic residues prior to the sixth milking after parturition to avoid high rates of false positive outcomes. B Fifty multiparous Holstein and Jersey dairy cows at the University of Connecticut dairy herd are currently being enrolled in a study to determine if the combined effects of NEB and ketosis increase the risk of cows developing mastitis during early lactation. C A new formulation of a chlorohexidine teat disinfectant is being evaluated. NY. The genome of S. agalactia isolated from cows and from humans was compared using a number of classification methods including antimicrobial resistance genes, pcr based methods and sequencing. An important overlap was present between human and bovine isolates. Further studies are needed to look at the potential importance of this finding. TN. Rhamnose may be useful in differential isolation media for Shiga toxin-producing E. coli (STEC)O26. Presence of shiga toxin 1 gene sequences may aid in rapid identification of E. coli O26. About 98% (148 of 151) of E. coli O157 strains (largely serotype H7) were stx+ and almost all elaborated Shiga toxins. Acid-resistance (~96% of STEC) and tellurite-resistance (~86% of STEC) were common characteristics.

Publications

Publications Attachment:

Attachment

Impact Statements

<li>A combination of basic and applied science aimed at reducing the incidence and severity of mastitis in dairy cows in order to improve animal health, milk quality, and productivity.<li>Improved mammary health and milk quality of first lactation cows through the use of antibiotic therapy in heifers to reduce current and prevent new intramammary infections.<li>A greater understanding of cow factors that contribute to mastitis development has led to improved management strategies for controlling
mastitis.<li>Continued evaluation of new teat dip formulations to provide consumers with reliable information and allow them to choose the best product for reducing mastitis.<li>Development of new vaccination protocols, including adjuvants and delivery strategies, to improve the effectiveness of coliform vaccines.<li>Greater understanding of genes modified during the periparturient period when cows are more susceptible to mastitis. A better understanding of these genes will eventually allow de
velopment of novel preventive and therapeutic strategies against mastitis. Impact 7. Enhanced understanding of the genetic make-up of E. coli, Strep. uberis, other coliforms and streptococci. This information then can be used to identify specific components of the bacteria that help it infect an animal. This basic research can lead to new methods of preventing and treating mastitis.; Impact 8. Providing a milk supply safe for human consumption is critical. As such, research has been
conducted to determine organisms present in milk that are human health pathogens, to compare human and bovine pathogens, to determine the most sensitive tests and times for evaluating the presence of antibiotic residues in milk, and to reduce antimicrobial use through the use of antimicrobial susceptibility tests.