Streptococcus uberis ST439 and ST475 induce differential inflammatory responses in a mouse intramammary infection model Susweta Das Mitra a,c , Bibek Ranjan Shome a, ⁎, Bhuvana Mani a , D. Velu a , Apala Banerjee a , Kiran Bankar b , Sankar Kumar Ghosh c , Sandip Santra a , K.P. Suresh a , Habibur Rahman a a ICAR-National Institute of Veterinary Epidemiology and Disease Informatics (NIVEDI), Ramagondanahalli, Yelahanka, Bengaluru, Karnataka, India b BionividPvt Ltd, Kasturinagar, Bengaluru, Karnataka, India c Department of Biotechnology, Assam University, Silchar, Assam, India abstract article info Article history: Received 11 January 2016 Received in revised form 4 March 2016 Accepted 28 March 2016 Available online xxxx Streptococcus uberis causing mastitis is a growing challenge to the dairy industry. Molecular, epidemiological and population structure studies have revealed clonal diversity among the infecting strains. In this study, mouse intramammary infection model was used to uncover the host immune response to two epidemiologically impor- tant live strains of S. uberis (SU1and SU2) obtained from subclinical case of mastitis possessing specific and unique multi locus sequence types (ST), pulsed field gel electrophoresis (PFGE) pulsotypes and virulence pro- files. Temporal (2 h, 4 h, 8 h, 12 h, 24 h and 48 h) expression of key inflammatory mediators (IL2, IL4, IL6, IL12, TNFα, IFNγ, GMCSF, TLR2, TLR4, TLR9, TLR11, TLR12, CD14, IL1β, RANTES, Lactoferrin, and CXCl1) by reverse transcription and probe-based quantitative real-time PCR showed relative mRNA levels higher (p b 0.05) in re- sponse to SU2 compared with SU1 with 24 h PI serving as a critical point for the deviating behavior (SU1 versus SU2). Further employing the predicted biological processes under the influence of this pool of tested genes, the delineation of gene regulatory networks suggested SU1 - favoring its persistence in the host environment; in con- trast, SU2 - which elevated gene expression indicating towards pathogen clearance or immune surveillance. This study suggested how these unique strains could manipulate the host immune response to influence the severity of mastitis; our results expand the available information on host pathogen interaction and provide a firm foun- dation needing further investigations to gain control over this pathogen. © 2016 Published by Elsevier B.V. Keywords: Mastitis Mice model S. uberis Immune response 1. Introduction Bovine mastitis is the most serious economically significant disease affecting the dairy industry. Several factors complicate the disease con- dition, including the multiple causative agents, poor understanding of the early immune response and the complexities associated with mam- mary epithelial cell damage by both the agent and host factors (Awale et al., 2012). Bacteria are the main etiological agents, with Staphylococcus aureus, Coagulase Negative Staphylococci (CoNS), Escherichia coli and Streptococ- cus uberis being the most common pathogens (Schukken et al., 2011; Kromker et al., 2014). Recently, S. uberis has been detected more fre- quently in a growing number of dairy herds and has caused both sub- clinical and clinical infections of the udder (Kromker et al., 2014; Swanson et al., 2009; Smolenski et al., 2014). Due to the importance of Streptococcal mastitis, several epidemiological studies have been per- formed across the globe to investigate its virulence potential and clonal diversity and to elucidate the pathogen biology (Wang et al., 2013; Shome et al., 2012; Rato et al., 2008; Zadoks, 2007). S. uberis possesses different virulence factors (i.e., CAMP factor (cfu), R-plasminogen acti- vator (pauA/skc), and the adhesion molecule gene (sua)) that empower its adherence, internalization and persistence in the host environment and enable it to establish infection (Shome et al., 2012; Patel et al., 2009; Chen et al., 2010). The protein encoded by sua gene is immuno- genic and plays a key role in adherence, thereby aiding persistent infec- tion (Chen et al., 2010). Over time, pathogens have evolved to be more ingenious than ever anticipated. Due to the existing difficulty in eradication and the increas- ing concern over antibiotic usage, alternative therapeutics are needed for mastitis control programs. In our previous study, we encountered novel clones of S. uberis that were prevalent in the southern province and caused subclinical mastitis (Shome et al., 2012) that required atten- tion from the mastitis control program. These clones were unique and were reported for the first time in mastitis surveillance. This finding augmented the need to unravel the bacteria's strategic interactions Gene xxx (2016) xxx–xxx Abbreviations: IMI, Intramammary Infection; PI, Post inoculation; ST, Sequence Type; TLR, Toll Like Receptor; IF, Interferon; TNF, Tumor Necrosis Factor; IL, Interleukin; CXCl, Chemokine. ⁎ Corresponding author at: ICAR-NIVEDI (formerly PD_ADMAS), Ramagondanahalli, Yelahanka, Bengaluru 560064, Karnataka, India. E-mail address: brshome@gmail.com (B.R. Shome). GENE-41260; No. of pages: 9; 4C: http://dx.doi.org/10.1016/j.gene.2016.03.054 0378-1119/© 2016 Published by Elsevier B.V. Contents lists available at ScienceDirect Gene journal homepage: www.elsevier.com/locate/gene Please cite this article as: Mitra, S.D., et al., Streptococcus uberis ST439 and ST475 induce differential inflammatory responses in a mouse intramammary infection model, Gene (2016), http://dx.doi.org/10.1016/j.gene.2016.03.054