Veterinary Immunology and Immunopathology 143 (2011) 125–130 Contents lists available at ScienceDirect Veterinary Immunology and Immunopathology j ourna l ho me pag e: www.elsevier.com/locate/vetimm Research paper The abundance of milk cathelicidin proteins during bovine mastitis G.A. Smolenski, R.J. Wieliczko, S.M. Pryor, M.K. Broadhurst, T.T. Wheeler, B.J. Haigh ∗ AgResearch Ltd, Ruakura Research Center, East St, Hamilton, New Zealand a r t i c l e i n f o Article history: Received 5 November 2010 Received in revised form 23 June 2011 Accepted 27 June 2011 Keywords: Mastitis Cathelicidin Neutrophil Diagnostic a b s t r a c t Current on-farm methods for detecting mastitis in dairy cows have limitations with their specificity and sensitivity, particularly at an early stage of infection. There is therefore a need to explore new approaches for detecting early and subclinical mastitis. This study examined the expression of a group of neutrophil-specific proteins, the cathelicidins, in milk samples from naturally occurring as well as experimentally induced mastitis infections. Immunoblot analysis indicated that cathelicidin proteins are only observed in infected quarters and demonstrate a high correlation with somatic cell count (SCC) during the onset of infection. In most of the infections examined, cathelicidin was detected prior to the observation of clinical symptoms and at SCC counts as low as 6.2 × 10 3 cells/mL. In naturally occurring mastitis the correlation between cathelicidin and infection status is not as strong, with 25% of pathogen-positive milk samples containing no detectable cathelicidin. This may reflect the varying levels of neutrophil concentration and activity at different stages or severities of infection. Our results indicate that milk cathelicidin levels increase following intramammary infection and cathelicidin-based biomarkers may assist in the detection of preclinical mastitis or determining the stage of infection. © 2011 Elsevier B.V. All rights reserved. 1. Introduction Mastitis can be difficult to detect prior to the onset of clinical symptoms. This is currently achieved through the measurement of milk somatic cell counts (SCC) and cul- ture based tests for microorganisms. Both tests require laboratory analysis at off-farm facilities and are not rou- tinely performed on individual cows throughout lactation. As a consequence, cows with subclinical infections may remain undetected in the milking herd, contributing to a high SCC in the bulk milk and to lowered milk production. As such, there is considerable interest in developing on- farm tests which are simple, fast and potentially amenable to in-line monitoring as part of milking systems. Current on-farm tests include the California mastitis test (CMT) (Schalm and Noorlander, 1957) or measuring the electri- cal conductivity of the milk (Fernando et al., 1982). Both ∗ Corresponding author. Tel.: +64 7 838 5099; fax: +64 7 838 5628. E-mail address: brendan.haigh@agresearch.co.nz (B.J. Haigh). of these tests are qualitative in their nature with reports showing moderate sensitivity and specificity when dealing with subclinical samples (Middleton et al., 2004; Norberg et al., 2004). There is therefore a need to investigate and develop new approaches for detecting early and subclinical mastitis on the farm. One approach for detecting infection in the mammary gland is to measure the proteins in milk that are part of the inflammatory response to infection. Proteins such as haptoglobin (Akerstedt et al., 2009), serum amyloid A (Eckersall et al., 2006), N-acetyl--d-glucosaminidase (Pyorala, 2003), and lipopolysaccharide-binding protein (Schroedl et al., 2001) have been identified as potential markers for mastitis. In a previous study we identi- fied cathelicidin-1 as one of a group of proteins which are increased in abundance in milk from mastitic cows (Smolenski et al., 2007). Cathelicidin proteins are a major component of the neutrophil secondary granule and increased levels of these proteins are observed in a range of inflammatory conditions (Zanetti, 2005). Fol- lowing degranulation or secretion, cathelicidin proteins 0165-2427/$ – see front matter © 2011 Elsevier B.V. All rights reserved. doi:10.1016/j.vetimm.2011.06.034