Iranian Journal of Microbiology
Volume:2 Issue: 1, Mar 2010

Enteropathogenic Escherichia coli (EPEC) strains can be detected by serogrouping and the presence of enterocyte attaching- effacing (eae) gene. Most EPEC strains belong to a certain O antigenic group. Locus of enterocyte effacement (LEE) Pathogenicity Island contains the eae gene and secretory proteins (ESPs) that introduce the attaching-effacing lesion. LEE inserted in tRNA genes include the SelC, PheU and PheV sites. The aim of the present study was to genetically characterize EPEC strains isolated from children with diarrhea. Serogrouping was performed by EPEC antisera in 321 E. coli isolates. The presence of eae, stx, espB, and eaf genes and detection of insertion sites of LEE was done by PCR using specific primers. Seventeen (5.3%) isolates belonging to 7 EPEC serogroups were identified among the whole material and all carried the eae gene. None of the 321 isolates showed presence of stx gene indicating that all 17 isolates classified as EPEC by O serogrouping did not belong to the enterohaemorrhagic E. coli (EHEC) group. Of these, 8 (53%) isolates carried the eaf and 16 (94.1%) carried the espB gene. The insertion sites of LEE in serogrouped isolates were selC (in 6 isolates), pheU (in 7 isolates) and pheV (in 2 isolates). The insertion site in 2 isolates was not determined by PCR. Serogrouping and detection of the eae gene showed to be reliable for detection of EPEC strains. No Shigatoxin- producing E. coli (STEC) strain was found among the isolates. Detection of the insertion site of LEE showed that selC, pheU or PheV are insertion sites of LEE in the EPEC strains.

Shiga toxin-producing Escherichia coli (STEC) strains are human pathogens linked to hemorrhagic colitis and hemolytic uremic syndrome. Shiga toxins (Stx1 and Stx2) are the major virulence factors of these strains. The aim of this work was to study the prevalence and distribution of stx1 and stx2 gene in E. coli O157:H7 and non-O157:H7 strains isolated from cattle in Shiraz, Iran. Four hundred and twenty samples consisted of recto-anal mucosal swabs were collected from cattle. They were checked for the presence of the stx1 and stx2 gene using multiplex-PCR every 1 week over a 1-year period (2007-2008). A total of 146 strains carrying the stx1 and stx2 gene were isolated from 51 (12.14%) cattle. Overall, 15 (3.57%) were identified as O157:H7 and 131 (31.19%) revealed to be non-O157:H7. Both stx2 and stx1 genes were detected in 51 (34.93%) STEC isolates. Genotypes stx1 and stx2 were detected in 15 (10.27%) and 78 (53.42%) respectively. Seasonal distribution of stx genes revealed high percentage of positive animals in warm seasons. The gene sequence similarity ranged from 94 to 100%. Frequency of stx1 and stx2 in animals and its relation to human disease is not well understood in Iran. The high prevalence of STEC in cattle seems to parallel that which is usually observed in warm seasons and it also parallels occurrence of human STEC. The higher prevalence of the stx2 gene than stx1 in strain populations isolated from cattle indicates a risk alert of E. coli O157:H7 being shed by cattle in these populations. Appropriate measures are now needed to prevent the spread of this life-threatening foodborne disease in our country.

Infection with vancomycin-resistant enterococci (VRE) has caused a therapeutic problem. VanA and VanB resistant types are the predominant phenotypes among vancomycin resistant enetrococci. Transposon 1546 (Tn1546) harboring the vanA gene cluster, plays an important role in the horizontal transfer of vanA gene. In this study, we examined the phenotypic and genotypic diversity of a number of clinical VRE. Twenty-four clinical VRE isolated from two university hospitals in Tehran were examined based on their antimicrobial susceptibility, Tn1546 related element organization and pulsed-field gel electrophoresis (PFGE) patterns. Integration of well-studied insertion sequence elements IS1216V, IS1542 and IS1251 was examined by PCR mapping and sequencing. From 24 isolates, 15 isolates with VanA phenotype and 9 isolates with VanB phenotype were identified which both groups interestingly possessed the vanA gene. According to PCR mapping, our isolates were assigned to 6 main groups. In 14 (58.3%) isolates, IS1216V was inserted in vanX-vanY region and/or in truncated left-hand of Tn1546-like elements. In 11 (45.8%) isolates, both IS1216V and IS1542 were inserted in vanX-vanY and orf2-vanR regions, respectively and none of them harbored IS1251. Interestingly, PFGE of the isolates showed a high degree of diversity. PCR mapping revealed that VanA elements in our isolates were highly heterogeneous. Overall, we found no correlation between transposon type and PFGE pattern. Genetic diversity of VRE provides practical information for epidemiological studies and our data showed horizontal transfer of VRE in this region.

To investigate microorganisms causing bacteremia in Jordanian children and to assess their sensitivity to various groups of antimicrobials. A retrospective study was conducted on positive blood cultures taken from 378 children aged below 15 year, who sought medical attention at Princess Rahmah Hospital between January and December/2008. Out of 4475 tested blood samples, 378 isolates were recovered from blood cultures. The male to female isolate ratio was (1.26:1.0). The most frequent pathogen found was Staphylococcus aureus (86.2%), followed by Klebsiella spp. (9%), Escherichia coli (1.9%), Streptococcus spp. (1.9%), Pseudomonas spp. (0.8%), and Acinetobacter sp. was found in only one culture (0.3%). The susceptibility rate of S. aureus was recorded the highest (99.6%) for vancomycin, and the lowest susceptibility rate (3.2%) was recorded for aztreonam. Staphylococcus aureus was the main isolate in bacteremic children, with all isolates demonstrating susceptibility to vancomycin. Overall, aztreonam resistance was near 97%, and this rate was not affected by sex and blood isolate type. This information should be considered when empirical therapy is recommended or prescribed for children with bacteremia.

Non-tuberculosis mycobacteria (NTM), as certain species of mycobacteria, can affect human in several ways. In the preceding years, the rate of NTM detection has risen in both immunocompromised and immunocompetent patients. On the other hand, several reports have debated the possibility of co-infection of both Mycobcateriu tuberculosis (MTB) and NTM in individuals that puts the role of NTM in disease manifestations under question. Moreover, it is now proven that some of the cases that are identified as anti-TB treatment failure or suspected for drug resistance are actually NTM.

The properties of neuraminidase produced by P. aeruginosa strain PAO1 during growth in a defined medium (BHI) was examined and compared with some neuraminidase features of K. pneumoniae in this investigation. The enzyme was isolated from concentrated culture supernatants of P. aeruginosa which was used in a sensitive fluorometric assay by using 2''-(4-methylumbelliferyl) α-D-N acetylneuraminic acid as substrate. Neuraminidase production in P. aeruginosa PAO1 paralleled bacterial growth in defined medium (BHI) and was maximal in the late logarithmic phase of growth but decreased during the stationary phase, probably owing to protease production or thermal instability. Highest production of P. aeruginosa PAO1 neuraminidase was in BHI culture media. The neuraminidase of P. aeruginosa PAO1 possessed an optimum temperature of activity at 56 °C and the activity was maximal at pH 5. Heating the enzyme to 56 °C for 45 min., in the presence of bovine serum albumin destroyed 33.1% of it''s activity and addition of Ca+2, EDTA and NANA also decreased activity markedly. The results revealed that the highest specific activity is for p. aeruginosa PAO1

Acetic acid bacteria (AAB) are useful in industrial production of vinegar. The present study aims at isolation and identification of acetic acid bacteria with characterization, optimization, and evaluation of their acetic acid productivity. Samples from various fruits were screened for presence of acetic acid bacteria on glucose, yeast extract, calcium carbonate (GYC) medium. Carr medium supplemented with bromocresol green was used for distinguishing Acetobacter from Gluconobacter. The isolates were cultured in basal medium to find the highest acetic acid producer. Biochemical tests followed by 16S rRNA and restriction analyses were employed for identification of the isolate and phylogenic tree was constructed. Bacterial growth and acid production conditions were optimized based on optimal inoculum size, pH, temperature, agitation, aeration and medium composition. Thirty-seven acetic acid bacteria from acetobacter and gluconobacter members were isolated. Acetic acid productivity yielded 4 isolates that produced higher amounts of acid. The highest producer of acid (10.03%) was selected for identification. The sequencing and restriction analyses of 16S rRNA revealed a divergent strain of Acetobacter pasteurianus (Gene bank accession number # GU059865). The optimum condition for acid production was a medium composed of 2% glucose, 2% yeast extract, 3% ethanol and 3% acid acetic at inoculum size of 4% at 3L/Min aeration level in the production medium. The isolate was best preserved in GYC medium at 12oC for more than a month. Longer preservation was possible at -70oC. The results are suggestive of isolation of an indigenous acetic acid bacteria. Pilot plan is suggested to study applicability of the isolated strain in acetic acid production.

Screening is a routine procedure for isolation of microorganisms which are able to produce special metabolites. Purified thermostable α-amylase from bacterial sources is widely used in different industries. In this study we analyzed samples collected from three different hot springs in Iran to detect any strains capable of producing thermostable α-amylase. Hot water samples from Larijan (67°C, pH 6.5), Mahallat (46°C, pH 7), and Meshkinshahr (82°C, pH 6), were cultivated in screening starch agar plates and incubated at 65°C for 24 hours. Thereafter, the plates were stained with Gram''s iodine solution. The bacterial colonies from the Meshkinshahr hot-spring produced the largest haloforming zone. Based on the phenotypic tests, the strain was identified as Bacillus sp. The culture condition was optimized for biosynthesis of α-amylase. The enzyme was produced at maximum level when it was incubated at 70 °C in the presence of soluble starch (1%) at pH 6. The addition of calcium (10 mM) and peptone (1%) to the mineral medium, shortened the lag period and improved the growth and α-amylase synthesis. The addition of glucose (1%) to the culture greatly diminished the syntheses of α -amylase. Importantly, the enzyme extract retained 100% activity when incubated for 45 minutes at 100°C. The Meshkinshahr hot-spring is rich in the Bacillus spp thermostable α-amylase producing strain of the thermophilic bacterial population. Iranian hot-springs like Meshkinshahr, have large microbial storages and can be used as sources of different biological products like enzymes. The enzyme which was produced with Bacillus sp. could hydrolyse polymers like starch and was used at laboratory scale successfully.