Cloning and Sequence Analysis of Genes Encoding Staphylococcus hyicu s Exfoliative Toxin Types A, B, C, and D

Abstract

Exfoliative toxins produced by certain strains of Staphylococcus hyicus mediate exudative epidermitis in pigs. In this study the genes coding for four different exfoliative toxin from S. hyicus (ExhA, ExhB, ExhC, and ExhD) were cloned and sequenced. The coding sequence of the four toxin genes ranged from 816 to 834 bp. The amino acid sequences of these four toxins were homologous to the earlier described exfoliative toxins SHETB from S. hyicus and ETA, ETB, and ETD from Staphylococcus aureus. The homology between the S. hyicus toxins was at the same level as the homology to the exfoliative toxins from S. aureus. The toxins showed similarity to serine proteases, including preservation of the catalytic tract in ExhA, ExhB, and ExhC. However, in ExhD, Asp in the putative catalytic tract was replaced with Glu. The recombinant toxins could be expressed in Escherichia coli, and three of the four toxins were recognized by monoclonal antibodies raised against native exfoliative toxins.Certain strains of Staphylococcus hyicus can cause the skin disease exudative epidermitis in pigs. Exudative epidermitis is characterized by separation of the cells in the epidermis in the upper stratum spinosum, exfoliation of the skin, erythema, and serous exudation (4, 32). Spontaneous exudative epidermitis in pigs is a generalized infection of the skin caused by strains of S. hyicus, which are able to produce a disease-causing factor designated exfoliative toxin (28). The exfoliative toxins responsible for the characteristic lesion of exudative epidermitis have been identified and purified from strains in Japan and Denmark. These toxins have been characterized as exoproteins of approximately 27 kDa or 30 kDa (6, 24). The toxins isolated in Japan were designated SHETA and SHETB (25), and the toxins isolated in Denmark were designated ExhA, ExhB, and ExhC. One additional toxin antigenically distinct from these was provisionally designated ExhD (5). The toxins differ in their antigenic properties (5, 26); however, no comparison of the toxins isolated in Japan and in Denmark has been published.The complex of skin lesions called the staphylococcal scalded skin syndrome in humans (7, 10, 16) has several aspects in common with exudative epidermitis in pigs. Staphylococcal scalded skin syndrome is caused by infection with Staphylococcus aureus strains that produce exfoliative toxin ETA, ETB, or both (14). The exfoliative toxins from S. aureus, ETA and ETB, have been cloned and their nucleotide sequences have been determined (12,17,20,22). Investigations have shown that the gene encoding ETA is located on the chromosome of S. aureus, while the gene encoding ETB is located on a 42-kb plasmid (reviwed by Arbuthnott [7]). The molecular masses of mature ETA and ETB as deduced from the amino acid sequences were 26,951 Da and 27,318 Da, respectively (17). The differences in the composition of the amino acid sequences reflected the previously observed antigenic differences of the two toxins (8,13,34). A third exfoliative toxin from S. aureus, designat...

Critical roles of amino acids Ser231, His107 and Asp156 of Staphylococcus sciuri exfoliative toxin C (ExhC) in the induction of skin exfoliations in neonate mice

“…Evidence for a homologous relationship is that the catalytic triad His72, Asp120 and Ser195 of ETA align respectively with His65, Asp114 and Ser186 of ETB (Bailey & Redpath 1992). In ExhA-ExhC, His115, Asp164 and Ser239 are considered to be the catalytic triad, although the Asp164 in ExhD is replaced with a glutamate (Ahrens & Andresen 2004). In this study, we found that Ser231, His107 and Asp156 formed the active site of ExhC, which supports the previous report (Ahrens & Andresen 2004).…”

“…From S. hyicus, a commonly seen causative agent for EE, four exfoliative toxins (ExhA-D) have been characterized (Ahrens & Andresen 2004). Although certain strain of S. hyicus code for more than one exfoliative toxins (Andresen 2005), ExhC is the only exfoliative toxin encoded by S. sciuri HBXX06 strain (Chen et al 2007).…”

“…It was reported that ExhA, ExhB, ExhC, ExhD, ETA and ETB show a certain similarity, exhibiting preservation of the catalytic tract of the active sites of the toxins (Ahrens & Andresen 2004). Evidence for a homologous relationship is that the catalytic triad His72, Asp120 and Ser195 of ETA align respectively with His65, Asp114 and Ser186 of ETB (Bailey & Redpath 1992).…”

“…In ExhA-ExhC, His115, Asp164 and Ser239 are considered to be the catalytic triad, although the Asp164 in ExhD is replaced with a glutamate (Ahrens & Andresen 2004). In this study, we found that Ser231, His107 and Asp156 formed the active site of ExhC, which supports the previous report (Ahrens & Andresen 2004). It was reported that mutant ETA, containing amino acid substitutions at any one of amino acid residues in the catalytic triad, are not active as exfoliations in the animal model of staphylococcal scalded skin syndrome (Papageorgiou et al 2000).…”

Serum cytokine modulation after Staphylococcus hyicus infection in BALB/c mice

“…S. hyicus, producing exfoliative toxins (ETs), is primarily known as the main causative agent of exudative epidermitis (EE) in piglets (Tanabe et al, 1996;Andresen, 2005). The disease is characterized by the separation of the cells in the upper stratum of the epidermis, exfoliation of the skin, erythema, and the formation of a thick, greasy, brown exudation (L'Ecuyer and Jericho, 1996;Ahrens and Andresen, 2004). S. hyicus has been isolated from pigs with septic polyarthritis and bovine mastitis and from White Leghorn laying chickens experiencing skin and feather problems (Phillips et al, 1980;Roberson et al, 1996;Chenier and Lallier, 2012).…”