Protective capacities of cell surface-associated proteins of Streptococcus suis mutants deficient in divalent cation-uptake regulators

Abstract

Many cell surface-associated, divalent cation-regulated proteins are immunogenic, and some of them confer protection against the bacterial species from which they are derived. In this work, two Streptococcus suis divalent cation uptake regulator genes controlling zinc/manganese and iron uptake (adcR and fur, respectively) were inactivated in order to study the protective capacities of their cell surface-associated proteins. The results obtained showed overexpression of a set of immunogenic proteins (including members of the pneumococcal histidine triad family previously reported to confer protection against streptococcal pathogens) in S. suis adcR mutant cell surface extracts. Likewise, genes encoding zinc transporters, putative virulence factors and a ribosomal protein paralogue related to zinc starvation appeared to be derepressed in this mutant strain. Moreover, protection assays in mice showed that although neither adcR-nor fur-regulated cell surface-associated proteins were sufficient to confer protection in mice, the combination of both adcR-and fur-regulated cell surface-associated proteins is able to confer significant protection (50 %, P50.038) against a challenge to mice vaccinated with them. INTRODUCTIONStreptococcus suis is an important pathogen that causes significant economic losses in the swine industry worldwide (Higgins et al., 1995). Infection caused by S. suis may vary from asymptomatic bacteraemia to fulminant systemic disease, with meningitis as the most outstanding feature (Higgins et al., 1995). It is also an important zoonotic agent for humans, who may acquire the infection through skin wounds upon contact with pigs and/or their by-products. Recently, during an outbreak in China, 215 human cases of S. suis infection were documented, and 39 of those patients died (Yu et al., 2006). More than 30 serotypes of S. suis, based on its capsular antigens, are currently known (Higgins et al., 1995). Serotype 2 is the most frequently isolated from diseased animals (Staats et al., 1997). Efforts to control infection are hampered by the lack of effective vaccines against S. suis. Furthermore, mechanisms involved in pathogenesis of this micro-organism are not yet completely understood (Gottschalk & Segura, 2000).Iron and zinc are essential components of many bacterial proteins either as structural or catalytic cofactors (Andrews et al., 2003;Babich & Stotzky, 1978). The concentration of both elements in the free state in the body fluids of mammals is extremely low in order to prevent bacterial proliferation. On the other hand, metals at high concentrations are toxic to micro-organisms. Toxicity occurs through the displacement of essential metals from their native binding sites or through ligand interactions (Babich & Stotzky, 1978; Chamnongpol et al., 2002). In addition, iron can be particularly toxic under aerobic conditions, interacting with either oxygen or oxygen-reduced species (Touati, 2000). Consequently, micro-organisms require homeostatic mechanisms to control intracellular metal levels.Fur (...

Lactococcus lactis ZitR Is a Zinc-Responsive Repressor Active in the Presence of Low, Nontoxic Zinc Concentrations In Vivo

“…Here, ZitR protein was one of the first members of the AdcR metallorepressor group to be characterized in vitro: purified rZitR protein was found to be a dimer able to bind up to four Zn(II) cations. Only two AdcR proteins were previously purified (1,3,44), and the S. pneumoniae AdcR protein has only recently been characterized as a dimer bound to three to five Zn(II) cations (44). An even number of bound cations is generally observed for dimeric forms of metalloregulators, in agreement with symmetric binding of both monomers (30,44).…”

“…adc-zit promoter regions display a conserved TTAACYRGTTAA palindrome (in one or two copies), proposed to be an AdcR/ZitR binding site (38). In pathogenic species, adcR is involved in biofilm formation (29), in the control of immunogenic surface proteins (1,3,34) and of zinc transport proteins (3), and in virulence (2). At the beginning of this work, little was known about AdcR/ZitR proteins (10,30,38).…”

“…At the beginning of this work, little was known about AdcR/ZitR proteins (10,30,38). S. pneumoniae and Streptococcus suis AdcR proteins were then found to be DNA-binding proteins (1,3), and S. suis AdcR was shown to be a metalloregulator specifically binding the TTAACYRGTTAA palindrome (1). While this work was submitted for publication, the S. pneumoniae AdcR protein was characterized in both apo-and Zn(II)-bound forms, and three of its Zn(II)-binding residues were identified (44).…”

Stress Physiology of Lactic Acid Bacteria

“…AdcR also regulates the expression of the four pneumococcal histidine triad (Pht) proteins, PhtA, PhtB, PhtD, and PhtE (77), involved in surface adhesion, as well as the Zn 2ϩ -specific AdcA homolog AdcAII and a Zn 2ϩ -dependent alcohol dehydrogenase (78,79). The adc operon is present in most streptococci and has also been studied in Streptococcus gordonii, where it is involved in Mn 2ϩ homeostasis (80), and in Streptococcus suis, where it regulates the expression of a Zn 2ϩ /Mn 2ϩ uptake system, ribosomal genes, and pht genes (81).…”

The Streptococcus suis transcriptional landscape reveals adaptation mechanisms in pig blood and cerebrospinal fluid

“…Research on gene regulation has focused mainly on protein regulators such as two-component or stand-alone regulators, including LuxS , CcpA , CovR ), CiaR/H , Ihk/Irr (Han et al 2012), AdcR (Aranda et al 2009b), ArgR , SalK/R (Li et al 2008), RevSC21 ), and Rgg . Recent studies, however, have revealed that regulatory RNAs (small RNAs from intergenic regions [sRNAs] and antisense RNA [asRNA]) have emerged as key regulators of virulence in several bacteria (Romby et al 2006;ToledoArana et al 2009;Waters and Storz 2009;Sharma et al 2010;Wurtzel et al 2012).…”