A statistical correlation between virulence genes and clinical features among patients with shigellosis in Mirzapur, Bangladesh

Abstract

Even though, the annual number of Shigella episodes or fatalities are not as terrifying as was reported in 1999, these numbers are still unacceptable. Shigella spp. is still contributing to about 800,000 deaths globally each year in concert with other diarrheagenic pathogens, affecting predominantly young children during the first 5 years of their lives, mostly in sub-Saharan and south Asian region. Therefore, it is still safe to consider Shigella as one of the major public health threats, specially in densely populated developing countries like, Bangladesh. This study was aimed at correlating the status of virulence genes among the Shigella strains and the corresponding clinical features observed in patients suffering from shigellosis. To achieve this goal, 61 different serotypes of S. flexneri strain, isolated from patients enrolled in Mirzapur study, during a period over 2009 to 2013, were randomly selected, and analyzed to detect the presence of 140 MD large virulence plasmid, and virulence (ipaH, ial, ipaH7.8), toxin (set, sen), and type 3 secretion system related genes (virB, ipaBCD, ipaB, ipaC, ipaD, ipgC, ipgB1, ipgA, icsB, ipgD, ipgE, ipgF, mxiH, mxiI, mxiK, mxiE, mxiC, spa15, spa47, spa32, spa24, spa). Simultaneously, two primers for the mxiE and spa15 genes, were designed and annealing temperatures were optimized. And the predictions were confirmed by PCR gel electrophoresis. Of these 61 strains, 140MD plasmid was detected in 79% (n=48) cases. The most prevalent gene was found to be ipaBCD (90%), followed by ial (89%), ipgC, ipgE (85%); virB, ipgA (82%); sen, mxiH, mxiI, spa15, spa47 (80%); ipgD (79%), ipgF (77%), spa32 (75%), ipgB1 (72%), mxiK, spa24 (70%); mxiE (66%), set (58%), icsB (44%) and mxiC (36%). This data was then compared against the clinical features for each corresponding strain, obtained from icddr,b Mirzapur study database, to detect any statistically significant association in between these two sets of data. Interestingly, 20 statistically significant correlation had been observed (p <0.05) while 5 of them were of high statistical significance (p <0.01). Briefly, the two Shigella enterotoxin genes (set, sen) were found to be significantly associated with multiple important clinical features frequently observed in shigellosis, such as, presence of blood in stool with a bloody mucoid consistency, rectal strain, dehydration, cough, fever and an overall increased state of disease severity, along with four other virulence genes (ial, ipaBCD, ipgD and spa24). The finding of this study elucidates the increased level of severity of moderate to severe diarrhea in patients infected with Shigella strains containing the enterotoxin genes, and also does it's part in building a gene library of this species.