Ovine subclinical mastitis: Proteomic analysis of whey and milk fat globules unveils putative diagnostic biomarkers in milk Elisabetta Chiaradia a, ⁎ , Andrea Valiani b , Micaela Tartaglia a, 1 , Fausto Scoppetta a, 1 , Giovanni Renzone c , Simona Arena c , Luca Avellini a , Simona Benda b , Alberto Gaiti a , Andrea Scaloni c , ⁎⁎ a Laboratory of Biochemistry, Department of Pathologic, Diagnostic and Clinical Veterinary Medicine, University of Perugia, 06126 Perugia, Italy b Laboratory of Dairy Production and Control, Istituto Zooprofilattico Sperimentale dell'Umbria e delle Marche, 06126 Perugia, Italy c Proteomics and Mass Spectrometry Laboratory, ISPAAM, National Research Council, 80147 Naples, Italy ARTICLE INFO ABSTRACT Article history: Received 18 January 2013 Accepted 19 March 2013 Available online 2 April 2013 Subclinical mastitis is one of the main causes of alteration in milk content and has a major impact on both animal welfare and economy in the dairy industry. A better knowledge is needed to understand the ovine mammary gland metabolism and its response to bacterial infection. In this study, the proteomic changes in ovine milk as a result of subclinical mastitis were investigated by comparing both whey and fat globule membrane profiles of samples from Staphylococcus chromogenes-positive individuals, with those from non-infected counter- parts having high or low somatic cell count; the latter were used as control. 2-DE and combined MS procedures were utilized for this purpose. Although sample bromatological parameters were very similar, proteomic analysis highlighted significant differences between the three experimental groups. Most relevant changes were observed between samples of infected milk and control. Modifications related to the defense response of the mammary gland to the pathogen were evident, with important consequences on nutritional and technological properties of milk. On the other hand, quantitative protein changes between non-infected samples with low and high levels of somatic cells indicated that the latter may result as a consequence of a probable unpaired cellular metabolism due to cellular stress, hormonal variations or previous infections. Putative biomarkers useful for the monitoring of sheep mammary metabolism and for the careful management of ovine subclinical mastitis to avoid its clinical degeneration are proposed and discussed. Biological significance Proteomics has been here applied to the differentiation of healthy and subclinical mastitic sheep milk samples, evidencing the response of the mammary gland to S. chromogenes infection. Presented results propose useful protein biomarkers for the detection of ewe mammary infection at its subclinical stages and, subsequently, mastitis recognition and treatment. Differently from bovine, these data confirm that the increase in somatic cell Keywords: Subclinical mastitis Ovine Proteomics Somatic cell count Whey Milk fat globule JOURNAL OF PROTEOMICS 83 (2013) 144 – 159 ⁎ Corresponding author. Tel.: + 39 075 5857751; fax: + 39 075 5857747. ⁎⁎ Corresponding author. Tel.: + 39 081 5966006; fax: + 39 081 5965291. E-mail addresses: elisabetta.chiaradia@unipg.it (E. Chiaradia), andrea.scaloni@ispaam.cnr.it (A. Scaloni). 1 Both authors contributed equally to this work. 1874-3919/$ – see front matter © 2013 Elsevier B.V. All rights reserved. http://dx.doi.org/10.1016/j.jprot.2013.03.017 Available online at www.sciencedirect.com www.elsevier.com/locate/jprot