Genome and transcriptome scale portrait of sigma factors in Mycobacterium avium subsp. paratuberculosis Leonardo A. Sechi a,b, * , Giovanna E. Felis a , Niyaz Ahmed b,c , Daniela Paccagnini a , Donatella Usai a , Silvia Ortu a , Paola Molicotti a , Stefania Zanetti a,b a Dipartimento di Scienze Biomediche, Sezione di Microbiologia Sperimentale e Clinica, Universita ` degli studi di Sassari, Viale S. Pietro 43/B, 07100 Sassari, Italy b ISOGEM, International Society of Genetic and Evolutionary Microbiology, Working Group on Mycobacterial Disease, Sassari, Italy c Pathogen Evolution Group, Centre for DNA Fingerprinting and Diagnostics, Hyderabad, India Received 9 October 2006; accepted 8 January 2007 Available online 12 January 2007 Abstract Mycobacterium avium subsp. paratuberculosis (MAP) is the etiological agent of Johne’s disease (JD), a chronic gastroenteritis of ruminants and other animals, including primates. Many evidences suggested association of MAP to Crohn’s disease, a chronic granulomatous gastrointestinal disease of humans with strong similarities with JD. The present study attempts to evaluate global gene regulation in MAP, which has not been addressed previously, despite the availability of MAP genome sequence. For this purpose, we investigated: (i) the presence of sigma factors and their relationship to sigma factors of other mycobacteria (M. avium subsp. avium, M. tuberculosis, M. bovis, M. leprae and M. smegmatis), and (ii) their expression during different growth conditions and in vitro infection of intestinal epithelial Caco2 cells. MAP genome contains 19 putative sigma factor, but only 12 belong to gene families common to other mycobacteria. Gene expression was evaluated with Real-Time PCR during growth in 7H9 medium and mycobactin J, in 7H9 medium plus mycobactin J and lisozyme, and during infection of Caco2 cells: very different expression patterns were observed and, on the whole, only 7 sigma factors were found to be expressed. sigJ was upregulated during the infection of Caco2 cells. Even if only few sigma factors were expressed in the three conditions tested, the overall high numbers of MAP sigma factors suggests a noteworthy flexibility of this pathogen. Thus, this first report on expression of MAP sigma factors opens the way to an extensive characterization of global gene regulation, as a key to understand strategies of survival and mechanisms of infections used by this organism. # 2007 Elsevier B.V. All rights reserved. Keywords: In silico analysis; Phylogenetic tree and expression of Sigma factor; Mycobacterium avium subspecies paratuberculosis; Caco2 cell infection 1. Introduction MAP bacilli are the etiological agents of Johne’s disease, a world-wide chronic gastroenteritis of ruminants (Chacon et al., 2004). It has been reported in non ruminant animals such as rabbits (Judge et al., 2005) foxes and stoats (Rowe and Grant, 2006) as well as primates (Zwick et al., 2002). Recent reports describe a strong association of MAP with Crohn’s disease, a chronic granulomatous gastrointestinal disease of humans which resembles Johne’s disease in several aspects (Bull et al., 2003; Naser et al., 2004; Sechi et al., 2005, 2001). The complete genome sequence of Mycobacterium avium subsp. paratuber- culosis (MAP) has been recently published (Li et al., 2005), as those of other mycobacterial species such as M. tuberculosis (Cole et al., 1998), M. bovis (Garnier et al., 2003), M. leprae (Cole et al., 2001), M. smegmatis [available to the public since October 2004 (http://www.tigr.org)]. Despite the availability of the complete genome sequence and several studies have been carried out on pathogen virulence and host responses, very little is known about the global gene regulation of MAP www.elsevier.com/locate/meegid Infection, Genetics and Evolution 7 (2007) 424–432 * Corresponding author at: Dipartimento di Scienze Biomediche, Sezione di Microbiologia Sperimentale e Clinica, Universita ` degli studi di Sassari, Viale S. Pietro 43/B, 07100 Sassari, Italy. Tel.: +39 079 228303; fax: +39 079 212345. E-mail address: sechila@uniss.it (L.A. Sechi). 1567-1348/$ – see front matter # 2007 Elsevier B.V. All rights reserved. doi:10.1016/j.meegid.2007.01.001