“Characterization of the PhoP/PhoQ system in S. marcescens and its role in pathogenesis”

BARCHIESI, JULIETA; CASTELLI, MARÍA EUGENIA; GARCÍA VÉSCOVI, ELEONORA

Villa Carlos Paz, Córdoba

Congreso; VI Congreso de la Sociedad Argentina de Microbiología General (SAMIGE); 2009

SAMIGE

The opportunistic human pathogen <i>Serratia marcescens</i> is a Gram-negative bacterium that causes disease in a wide range of both invertebrate and vertebrate hosts and in plants. <i>S. marcescens</i> is a significant cause of hospital-acquired infection, especially in patients with impaired immunity. As many <i>S. marcescens</i> strains are also resistant to multiple antibiotics, it represents a growing problem for public health. However, relatively little is known about the factors that contribute to <i>S. marcescens</i> pathogenesis within its host. In our laboratory, with the aim of finding regulatory mechanisms that would participate in <i>S. marcescens</i> pathogenesis, we undertook the characterization of PhoP/PhoQ two component system. This system controls transcription of key virulence genes in diverse bacterial pathogens. In order to study the physiological role of <i>phoP</i> in <i>S. marcescens</i>, we have constructed a <i>phoP</i> mutant strain. This mutant exhibited impaired growth in minimal broth limited in Mg²⁺, in acid pH, and showed increased sensitivity to antimicrobial peptides than the wild type strain. Furthermore, the <i>phoP</i> strain was attenuated in the survival inside epithelial cells. These results suggest that the <i>phoP</i> gene is required for <i>S. marcescens</i> invasion of epithelial cells. In addition, &beta-galactosidase assays revealed that <i>phoP</i> transcription was modulated by the Mg²⁺ and Polimixin B levels, predicted to be environmental signals detected by the system. To further explore the PhoP regulon, we perform a random mutagenesis strategy selecting for Mg²⁺ transcriptional regulated clones. One of the novel PhoP-activated genes identified was <i>mgtE</i>. MgtE is a Mg²⁺ transport protein and it has never been reported before as a PhoP regulated gene. Conversely, MgtA and MgtB, members of another Mg²⁺ transport family, have been extensively characterized as PhoP regulon member in <i>Salmonella</i>. This result adds a new Mg²⁺ transporter as a PhoP target, reinforcing the importance of PhoP/PhoQ system in Mg²⁺ homeostasis control. On the other hand, we carried out an <i>in silico</i> search of PhoP binding sites in the <i>S. marcescens</i> genome by MEME/MAST programs. We identified several predicted gene members of the PhoP regulon, such as <i>phoP</i>, <i>mgtA</i>, <i>mgtCB</i> and <i>pmrG</i>. The other PhoP binding sites identified corresponded to genes previously uncharacterized as PhoP regulon members in others bacteria. Among them, genes implied in copper resistance (<i>cueR</i>), citrate metabolism (<i>citB</i> and <i>citC</i>), oxidative stress resistance and antibiotics resistance (<i>ramA</i>). To our knowledge, this represents the first study of the virulence PhoP/PhoQ system in <i>S. marcescens</i>.