Note Structural analysis of the lipoteichoic acids isolated from bovine mastitis Streptococcus uberis 233, Streptococcus dysgalactiae 2023 and Streptococcus agalactiae 0250 Anna Czaban ´ ska a,  , Olga Neiwert a,  , Buko Lindner b , James Leigh c , Otto Holst a , Katarzyna A. Duda a,⇑ a Division of Structural Biochemistry, Research Center Borstel, Leibniz-Center for Medicine and Biosciences, Airway Research Center North (ARCN), Member of the German Center for Lung Research (DZL), Parkallee 4a/c, D-23845 Borstel, Germany b Division of Immunochemistry, Research Center Borstel, Leibniz-Center for Medicine and Biosciences, Borstel, Germany c School of Veterinary Medicine & Science, University of Nottingham, Nottingham, UK article info Article history: Received 20 July 2012 Received in revised form 4 September 2012 Accepted 5 September 2012 Available online 15 September 2012 Keywords: Streptococcus uberis Streptococcus dysgalactiae Streptococcus agalactiae LTA structure NMR analysis abstract Lipoteichoic acid (LTA) is an amphiphilic polycondensate located in the cell envelope of Gram-positive bacteria. In this study, LTAs were isolated from the three bovine mastitis species Streptococcus uberis 233, Streptococcus dysgalactiae 2023, and Streptococcus agalactiae 0250. Structural investigations of these LTAs were performed applying 1D and 2D nuclear magnetic resonance experiments as well as chemical analyses and mass spectrometry. Compositional analysis revealed the presence of glycerol (Gro), Glc, ala- nine (Ala), and 16:0, 16:1, 18:0, 18:1. The LTAs of the three Streptococcus strains possessed the same structure, that is, a lipid anchor comprised of a-Glcp-(1?2)-a-Glcp-(1?3)-1,2-diacyl-sn-Gro and the hydrophilic backbone consisting of poly(sn-Gro-1-phosphate) randomly substituted at O-2 of Gro by D-Ala. Ó 2012 Elsevier Ltd. All rights reserved. Bovine mastitis, inflammation of the udder, usually arises as a result of intramammary infection by bacteria. 1 The signs of masti- tis vary according to factors in the host and the invading pathogen and intramammary infection may result in sub-clinical or clinical disease. 2,3 Sub-clinical mastitis, by definition, shows no obvious signs of disease. 4 Those associated with clinical disease can range from visible abnormalities in the milk (protein aggregates or clots) accompanied by pain and swelling in the affected gland to produc- tion of a secretion which is composed solely of aggregated protein in a serous fluid. In severe cases there may be systemic signs such as elevated temperature and loss of appetite which may develop to bacteremia, septicemia and death of the animal. 1 Milk from the uninfected gland contains leukocytes, including macrophages, neutrophils, and lymphocytes typically below 200,000 cells/ml. 5 Infection usually results in an inflammatory re- sponse which leads to an increase in cell count, primarily due to the influx of neutrophils. Milk from sub-clinically infected quarters usually has a cell count in excess of 250,000 cells/ml but this figure may vary widely. That from clinically infected quarters usually contains in excess of 2000,000 cells/ml. 5 The inflammatory reac- tion and the increase in the numbers of neutrophils in the gland re- sult in a lower rate of milk production and a gross deterioration of the quality of the secretion. Such milk is unmarketable. Over 135 infectious agents have been associated with clinically apparent episodes of mastitis 6 but the vast majority of cases are due to infection with one of five bacterial species. Those most com- monly implicated are Streptococcus uberis, 7 Escherichia coli, 8 Staph- ylococcus aureus, 8 Streptococcus dysgalactiae, 9 and Streptococcus agalactiae. 10 Virulence determinants associated with the bacterial cell sur- face play an important role in the persistence and pathogenesis of bacterial infections. This process includes colonization through adherence and invasion of host cells and the subversion of host response. The proteins involved in such processes are often candidates for inclusion in vaccines or targets for new therapeutic agents. However, the surface of Gram-positive bacteria is decorated with many different macromolecules, including carbohydrates, teichoic acids, lipoteichoic acids (LTAs), and polysaccharides. 11 LTA comprises in most cases glycerol phosphate (GroP) repeating units which are bound to a glycolipid anchor and can be further substituted by sugars and amino acids, in particular D-Ala. Ribitol was also found to be present in certain species. LTA seems to be indispensable for viability, 12 and is important in cell division and in binding cell-wall proteins. During infection it contributes to host cell binding. LTA of different bacteria 0008-6215/$ - see front matter Ó 2012 Elsevier Ltd. All rights reserved. http://dx.doi.org/10.1016/j.carres.2012.09.007 ⇑ Corresponding author. Tel.: +49 4537 188 2490; fax: +49 4537 188 7450. E-mail address: kduda@fz-borstel.de (K.A. Duda).   These authors contributed equally to this work. Carbohydrate Research 361 (2012) 200–205 Contents lists available at SciVerse ScienceDirect Carbohydrate Research journal homepage: www.elsevier.com/locate/carres