Levels of stx2a expression in STEC strains with different q genotypes

Abstract

Although STEC pathogenicity is a multifactorial phenomenon, the main virulence factors are Shiga toxins 1 and 2 (Stx1 and Stx2), which are directly related with the development of HUS. Stx2a is the subtype most frequently associated with HUS both in Argentina and worldwide. Additionally, it has been suggested that the pathogenic potential of STEC also correlates with the amount of Stx2 produced. In general, stx genes are located in the late region of prophages. The anti-terminator protein Q allows transcription of late genes and influences stx expression. The aims of this study were to evaluate the stx2a expression levels of STEC strains, both in basal and induced states, and to compare them taking into account the different q genes carried by each strain.METHODS: In total 35 STEC strains that carry stx2a (alone or in combination with another stx subtype) were chosen for this study. They have been isolated from different sources and belonged to serotypes O26:H11, O91:H21, O113:H21, O145:H- and O157:H7. The presence of q933, q21 and qO111 variants and their location close to stx2 were determined by PCR. The cultures were grown in LB and the lytic cycle of the phages was induced adding mitomycin C. The stx2a expression levels were measured by qPCR and the results were analyzed based on the ΔΔCT method. The differences in stx2a transcription were compared by analysis of variance (ANOVA).RESULTS: All strains excepting one showed higher levels of stx2a expression under mitomycin C induction. Additionally, a strain with no detectable stx2a expression was excluded from the analysis. Nineteen strains carried only q933 subtype, 11 carried both q933 and q21, and one strain was positive for qO111. Proximity to stx genes was confirmed in all cases. Three strains were negative for all of the q variants analyzed. These strains showed lower basal expression levels than those with q9333/q21 genotype and lower induced expression than those with q933 or q9333/q21 genotypes. In addition, a significant lower basal expression was detected in strains carrying only q933 in comparison with strains of genotype q933/q21. Nevertheless, in the induced state no significant differences were observed.CONCLUSIONS: Strains negative for the studied q genes showed low levels of stx2a expression and a low response to induction. However, a differential trend in stx2a expression was not clear between q933 and q933/q21 strains.