Short report: high prevalence of serine protease autotransporter cytotoxins among strains of enteroaggregative Escherichia coli

Abstract

Enteroaggregative Escherichia coli (EAEC) pathogenesis is thought to comprise intestinal colonization followed by the release of enterotoxins and cytotoxins. Here, we use a polymerase chain reaction (PCR) to determine the prevalence of 10 genes encoding serine protease autotransporter toxins (SPATEs) in a collection of clinical EAEC isolates. Eighty-six percent of EAEC strains harbored genes encoding one or more class I cytotoxic SPATE proteins (Pet, Sat, EspP, or SigA). Two Class II, non-cytotoxic, SPATE genes were found among EAEC strains: pic and sepA, each originally described in Shigella flexneri 2a. Using a multiplex PCR for five SPATE genes (pet, sat, sigA, pic, and sepA), we found that most of the Shigella isolates also harbored more than one SPATE, whereas members of most other E. coli pathotypes rarely harbored a cytotoxic SPATE gene. SPATEs may be relevant to the pathogenesis of both EAEC and Shigella spp.

Figure 1. Secretion of SPATE proteins from EAEC strains

Supernatants from 5 representative EAEC strains are shown. Supernatant proteins were precipitated with 10% TCA and separated by SDS-PAGE followed by Coomassie Staining. SPATE proteins represented by bands directly identified by mass spectrometry were SepA from strains H194-2 and 239-1; Pic from strains H77-1 and 239-1; Sat from strain 232-1-1 (indicated by arrows). MW, Molecular weight; HB101, negative control strain.

Figure 2. Distribution of Class I and Class II SPATEs among EAEC strains

The distribution of the SPATE genes among 55 EAEC strains was determined by PCR. (A) Distribution of Class-I and Class-II SPATEs among the EAEC collection. (B) Frequency of Pet, Sat, SigA, and EspP.

Figure 3. Identification of SPATE sequences from EAEC and Shigella isolates by multiplex PCR

PCR products representing genes for Sat, SepA, Pic, SigA and Pet were separated in 1.2% agarose gels. (A) PCR products from 11 EAEC strains (B) PCR products from 12 Shigella strains. EAEC strain 042 is the positive control for pet and pic; S. flexneri 2a strain 2457T is the positive control for sat, sepA, pic and sigA.