[ frequency of toxigenic strains of Clostridium difficile in hospitalized patients with diarrhea in Tehran/Iran by PCR method, 2010]

Clostridium difficile has been identified as a pathogen in antibiotic associated diarrhea [AAD], pseudomembranous colitis and also nosocomial diarrhea. The present study was performed to find the prevalence of toxigenic strains of C .difficile isolated from diarrhea patients hospitalized in Tehran hospitals. A total of 98 fecal samples obtained during July to December 2010 were studied. Samples were rapidly cultured on the CCFA medium and incubated at the anaerobic conditions. Then ELISA was done to detect toxin A and B in the stool. Molecular identification of C.difficile was done by cdd3 universal primer and toxin A gene [tcdA], toxin B gene [tcdB] and binary toxin profiles were determined by PCR method. From a total of 98 fecal samples, 15 samples [15.3%] were positive of which, 12 strains [21.2%] were A+B+, 2 strains [2%] were A+B-, and 1 strain [1%] was A-B+. This study showed that Clostridium difficile is an important pathogen in the development of nosocomial diarrhea. Therefore, routine detection of C.difficile in suspected cases is recommended

Antimicrobial resistance profile and presence of class I integrongs among Salmonella enterica serovars isolated from human clinical specimens in Tehran, Iran

Salmonella is one of the leading causes of food-borne diseases. Increasing occurrence of antimicrobial resistance, especially multidrug-resistance, in Salmonella serovars is a major public health problem worldwide. This study was carried out to detect class I integrons and antibiotic resistance profiles in clinical isolates of Salmonella serovars collected from seven hospitals in Tehran during November 2009 to June 2010. Antibiotic susceptibility profile of 19 antibiotics against 58 Salmonella isolates commonly used in humans was determined using disk diffusion assay. Minimum inhibitory concentration against ceftriaxone and ciprofloxacin was studied. PCR assays were used to detect class I integrons. Among 58 Salmonella isolates, 72.4% were Salmonella enterica serovar Enteritidis, 8.7% were Salmonella enteric serovar Typhimurium and 18.9% were other serovars. Of the total 58 Salmonella serovars, 43 [74.1%] were multidrug resistant and showed resistance to three or more antibiotic families. Class I integrons were identified in 38 [88.3%] MDR Salmonella isolates. Ciprofloxacin minimum inhibitory concentration ranged between 0.125-2 microg/ml for four isolates and other four isolates exhibited resistance to ceftriaxone [MIC 64-256 microg/ml]. The high prevalence of class I integrons was seen in our MDR Salmonella isolates and class I integrons might play an important role in the dissemination of antimicrobial resistance determinants

[ role of typical enteroaggregative escherichia coli [tEAEC] strains in diarrhea among children]

Enteroaggregative Escherichia coli [EAEC] strains have been implicated as causative agents of acute, chronic and persistent diarrhea in children. EAEC have special aggregative adherence pattern for HEp-2 or HeLa cells. PCR methods based on the presence of essential virulence factors would facilitate the diagnosis of EAEC strains. The aim of this study was to assess the usefulness of the PCR assay targeting the pCVD432 gene for the detection of EAEC strains and the presence of aggR, agg, aafA, aap and astA genes which can be used for the differentiation of typical and atypical EAEC strains. One hundred and forty faecal samples were collected from children of less than 12 years of age with diarrhea at the Besat Hospital, in Hamadan, between July 2007 and May 2008. The faecal samples were plated onto MacConkey agar and incubated at 37 oC for 18 hours. E.coli colonies were identified by biochemical tests. EAEC strains were identified by ues of pCVD432 primer which is specific for EAEC strains. Then their virulence factors were evaluated by use of specific primers of aggR, aggA, aafA, aap and astA genes. Strains of E. coli harboring the pCVD432 gene were found in the faecal samples from 15 [10.8%] patients. The aggR gene was present in 11[73.3] strains. The aggA gene was found in 3 isolates and 4 strains had the aafA gene. The aap and astA genes were found in 9 and 7 strains, respectively. Several different combinations of the genetic markers were found among the EAEC strains. The most prevalent combinations were aggR/aap, found in 8 [53.3%] and aggR/aafA detected in 4[26.7%] strains. The results of this study showed that EAEC strains were one of the most prevalent enteric pathogens in children in this region. The pCVD432 gene PCR assay is the most common method for detection of EAEC strains. Typical EAEC strains with aggR gene had a high prevalence in this region and considering the pattern of virulence factors, they were heterogenous

[HeLa-cell adherence patterns of shiga toxin- producing Escherichia coli [STEC] serotypes isolated from faecal samples]

Shiga toxin-producing Escherichia coli [STEC] has been recognized as an important cause of diarrheal diseases. The ability to colonize the human intestine is an essential part of the infection. To determine the importance of the adherence among non-O157 STEC serotypes isolated from diarrheal and asymptomatic healthy persons, HeLa cell adherence pattern was studied. Polymerase chain reaction-restriction fragment length polymorphism [PCR-RFLP] analysis of the flagellin gene [fliC] was performed in 35 strains of STECO serogroups for determining their flagellar antigen [H] status. The entire coding sequence of fliC was amplified by PCR, the amplicon was restricted with HhaI, and the restriction fragment pattern was examined after gel electrophoresis. The adherence property of 35 Shiga toxin-producing Escherichia coli [STEC] not belonged to O157:H7 serotype was checked on HeLa cell adherence pottern. Three to five individual colonies from each case were assessed. These strains were isolated from patients with bloody, non-bloody diarrhea and healthy cases. Of the 35 STEC colonies, 2 strains [5.7%] were isolated from bloody diarrheal cases, 25 strains [71.5%] from non-bloody diarrheal and 8 strains [22.8%] from healthy persons. None of the strains belonged to O157: H7 serotype but 22 strains belonged to O126 and O128, O26 and O111 serogroups. Out of 27 strains isolated from bloody and non-bloody diarrheal cases 3 strains [11.1%] were non-adherent [NA] and 24 strains [88.9%] were adherent. However, aggregative adherence [AA] was observed in 10 serotypes [O128: H2, O128: H9, O125: H12 and O125: H15] and diffuse adherence [DA] and localized adherence [LA] were exhibited by 1 serotype, respectively. The remaining 12 strains showed non-specific adherence [NSA]. Among adherence patterns, LA and AA were found to be significantly associated with diarrhea [p < 0.001]. Human pathogenicity of E. coli strains belonging to STEC group varied according to serotypes, virulence attribute and adherence patterns. Based on the observation obtained in this study, it can be concluded that STEC strains with adherence property are more virulent than the non-adherence ones and further detail studies on adherence factors and their mechanisms of pathogenesis are needed

Serotypes of enteropathogenic Escherichia coli isolated from children under 5 years of age

The purpose of this study was to find out the frequency of different serotypes of enteropathogenic Escherichia coli [EPEC] among healthy/ diarrheal cases. A total of 191 strains, 111 from diarrheal and 80 from asymptomatic persons were examined. Determination of the EPEC serogroups was performed by agglutination tests using polyvalent and monovalent O antiserum. PCR-RFLP analysis of the flagellin-encoding [fliC] gene and agglutination tests using polyvalent and monovalent sera against H antigens [H1 to H56] according to the instructions of the manufacturer was performed. Seventeen [8.9%] strains were non-motile and untypable with conventional serotyping method that showed as H[-]. Forty-three fliC restriction patterns were found for motile and non-motile serotypes. Each motile serotype was characterized by one or two fliC specific restriction patterns. O142:H48 [6.8%], O86:H48 [6.35], O111:H21[4.7%] and O127:H21 [4.2%] were the most prevalent serotypes, and O55:H12/45, O86:H48, O127:H21, O142:H48, O126:H19 serotypes were the most frequently agents in diarrheal cases, compared to asymptomatic children [P<0.05]. There were common EPEC serotypes in diarrheal and asymptomatic children, however some serotypes either found only in diarrheal cases or isolated from asymptomatic persons. Some serotypes were isolated more frequently from diarrheal cases than asymptomatic persons. The conventional serological method using antisera is the basis for the H typing system in E-coli, but it is impossible to serotype non-motile bacteria. PCR-RFLP analysis of fliC gene is a practical method in identifying the H variant in motile and non-motile EPEC serotypes and is useful for epidemiological studies

Serogroup Distribution of Shigella in Tehran

In recent years, the importance of Shigella as an enteric pathogen with global impact has been increasingly recognized. In this study, serogroup distribution of Shigella isolated from clinically diagnosed cases of gastroenteritis and acute diarrhea in Tehran, capital of Iran was investigated between December 2002 and November 2003. Fecal specimens and rectal swabs were cultured for Shigella spp. using st and ard microbiological techniques. The isolates of Shigella were identified by biochemical assay and serological testing. From a total of 302 Shigella isolates, 178, 110, 10 and 4 strains were identified as S.sonnei [58.9%; 95% CI: 53.2-64.5], S. flexneri [36.4%; 95% CI: 31.0-42.2], S.boydii [3.3%], and S. dysenteriae [1.3%], respectively. The peak of infection occurred during summer. Overall, 167 patients [55.3%] were males and 135 [44.7%] were females

Role of Aeromonas hydrophila in Diarrhea

Aeromonas hydrophila is a causative agent of a number of human infections. Aeromonads have been isolated from patients with diarrhea. In spite of a number of virulence factors produced by Aeromonas species, their association with diarrheal diseases has not been clearly linked. In current study, 1546 fecal samples of a randomly selected population were screened for presence of A. hydrophila. Out of the total number of cases, 20 were suffering from diarrhoea and the rest were asymptomatic healthy individuals. The result showed that 3.4% of the samples were positive for Aeromonas spp. A. hydrophila was isolated as the sole enteropathogen from 80% diarrheal and 20% asymptomatic cases. A significant association was found between the A. hydrophila and diarrhoeal diseases in rural and urban areas [P<0.05], while no difference was revealed between genders. The highest rates of A.hydrophila [3.9%] was detected in children <6 years of age. A significant association was also found between A. hydrophila and different age groups [P< 0.05]. In conclusion, our epidemiological study showed that Aeromonas spp. as a sole enteropathogen could be responsible for diarrhea

Rapid identification of group a streptococci by pyrrolidonyl beta-naphthylamide hydrolysis

In a clinical trial a new pyrrolidony Beta -naphthylamide [PYR] hydrolysis test was compared with the bacitracin disk susceptibility test for accuracy in the presumptive identification or group A streptococci [GAS]. Among 128 isolates of beta-hemolytic streptococci 93 group A isolates were found. The sensitivity of the PYR and bacitracin tests were similar [98.9%], but the bacitracin test had a lower specificity [80%] than the PYR test [100%]. The efficiency of the PYR and bacitracin test were 99.2% and 93.7%, respectively- All bacitracin tests were performed on subcultures of the isolates from the primary plate, whereas PYR testing was performed on colonies from the primary plate. This shortened the turnaround time for the PYR test compared ro the bacitracin test by at least 24 hours