PCR Detection of Microbial Pathogens PCR Detection of Microbial ...

164 Gallien
The first description of STEC as a source of HC was published in 1983 (11).
Meanwhile, STEC have been isolated from food, feces, stool, sewage, drinking
water, and various other habitats connected with human infectious diseases.
The toxins produced by STEC were shown to be cytotoxic for a number of
tissue culture cell lines, e.g. kidney cells from the green African monkey known
as Vero cells (12). This led to the term verotoxin for shiga toxin and, consequently,
verocytotoxin-producing E. coli (VTEC) as a synonym for STEC, with
both terms being equally used at present.
Enterohemorrhagic E. coli (EHEC) form a subgroup of STEC (VTEC). However,
it is still unclear which factors and/or mechanisms actually make an EHEC
out of an STEC. Obviously, all EHEC strains possess shiga toxin genes (stx).
The shiga toxins themselves exhibit RNA-glycosidase activity and inhibit the
synthesis of proteins in eukaryotic cells (13). The activity of the toxins is associated
with subunit A, which consists of two parts (structural data of stx genes are
given below). The A1 fragment harbors the specific N-glycosidase activity,
whereas fragment A2 links A1 to five B subunits. In a eukaryotic host cell, the
enzyme cleaves at the adenine at position 4324 in a loop structure of the 28S
rRNA of the 60S ribosomal subunit (13).
A schematic presentation of transmission routes to man is given in Fig. 1.
Direct transmission of EHEC from ruminants as the main reservoir for STEC
or from raw foods, such as milk, raw or undercooked meat, sausage containing
beef, cheese produced from raw milk, to humans, as well as transmission from
man to man are observed frequently (14).
The classification of E. coli into serogroups O, K, and H is well established.
Serogroup O157 occurs very often in human EHEC infections, but other
serogroups such as O22, O26, O103, O111, and O145 also figure prominently
(15–18). As stx genes are located on temperent phages, many E. coli serotypes
are subject to infection by these phages (19). A World Health Organization
(WHO) list contains more than 50 E. coli serotypes connected with EHEC
infections in man (20,21). Moreover, other bacteria, e.g., Citrobacter freundii
are also known to harbor stx genes (22).
Virulence genes of STEC/EHEC can be located on the DNA of temparent
phages, on plasmids, or on the chromosome. The Shiga-toxin genes are located
on the DNA of phages. At present, the following subtypes are known: stx 1; stx 2;
stx 2c; stx 2d; stx 2e; and stx 2f (23–30).
Pathogenicity islands like the locus of enterocyte effacement (LEE) or a
high- pathogenicity-island (HPI) (31) and the ast A gene (32) are located on the
chromosomal DNA of the cells. LEE contains a gene cluster coding for a type
III secretion system. Some proteins expressed by genes of LEE form a channel
to connect a bacterial cell and a host cell. Subsequently, other proteins encoded
by genes of LEE can pass this channel.