Iranian Journal of Microbiology
Volume:7 Issue: 3, Jun 2015

Antibiotic resistance is increasing, especially in healthcare-associated infections causing significant public health concerns worldwide. National information is required to make appropriate policies, update list of essential drugs for treatment, and evaluate the effects of intervention strategies. A nationwide surveillance of antimicrobial resistant bacteria in nosocomial infections was established in Iran in 2008, so that the data obtained through the surveillance would enable us to construct a database. Seven major teaching hospitals in Shiraz, Tabriz, Sari, Mashhad, Sanandaj, Ahwaz and Isfahan participated in this study. A total of 858 strains isolated from blood and other sterile body fluids were tested. Identification at the species level was performed with conventional biochemical methods and the API system. Susceptibility tests were done using disk diffusion method. The methicillin-resistance in S. aureus (MRSA) was determined by the oxacillin agar screen plate and respective MIC values were assessed using the E-test strips. The confirmatory disk diffusion methods were applied for phenotypic identification of extended-spectrum β- lactamase (ESBL) production for E. coli and K. pneumoniae, according to CLSI guidelines. Cultivation and re-identification of the strains yielded 858 isolates, consisting of 224 S. aureus, 148 Klebsiellaspp., 105 Serratia spp., 146 E. coli, 67 Acinetobacter spp., 38 Enterobacter spp., 95 Pseudomonas spp., 71 P.aeruginosa.35 Stenotrophomonas sp., and 8 other organisms. MRSA was detected in 37.5% of the isolates. No vancomycin-resistant or vancomycin-intermediate resistant S. aureus was detected. With the exception of Acinetobacter and Stenotrophomonas, 85% of the Gram-negative isolates were found to be susceptible in vitro to imipenem. Overall, about 61% of K. pneumoniae and 35% of E. coli isolates were ESBL producing. Multidrug resistant isolates of Gram-negative organisms and methicillin-resistant strains of S. aureus have been detected in many hospitals in this study.

Haemophilus influenzae type b (Hib) is divided into two distinct genotypes, type I and type II, based on the structure of capsular polysaccharides. The capsulation locus of Haemophilus influenzae type b consists of three functionally distinct regions, designated regions 1 to 3. Region III contains hcsA and hcsB genes; however, notable sequence variation in this region can be used to recognize different Hib genotypes. The purpose of this study was to investigate the prevalence and genotype of the Hib strains isolated from patients with invasive disease in Iran. In the present study, 8 pairs of primers were used for identification and serotyping of encapsulated Haemophilus influenzae strains, as well as confirmation of species identification. Additionally, in order to identify the cap- sular genotypes of Haemophilus influenzae type b (type I and II), two additional primer pairs were used to amplify the hcsA gene. Out of 50 isolates of H. influenzae, four were found to be type b. Interestingly, among these 4 Hib isolates, 2 strains belonged to the type-II category. Our study shows that the prevalence of both Hib types I and II seems to be high in Iran.

Klebsiella pneumoniae is an opportunistic pathogen responsible for up to 10% of nosocomial infections. The emergence and spread of multidrug resistant K. pneumoniae, mostly due to the production of extended-spec- trum β-lactamases (ESBL) and carbapenemases, is often responsible for antibiotic treatment failure of these infections. We compared the antibiotic resistance profiles, ESBL and carbapenemase production as well as presence of KPC-type genes in burn and non-burn clinical isolates of K. pneumoniae. Fifty five clinical isolates were collected from Shahid Motahari (25 burn isolates) and Shariati (30 non-burn isolates) hospitals between August 2011 to January 2012. Antibiotic susceptibility was determined to 12 antibiot- ics using disc diffusion. The phenotypic confirmatory test (PCT) was used to screen for ESBL production. Carbapenemase activity was measured by the modified Hodge test (MHT) and KPC-type carbapenemases were further sought by PCR using specific primers. Both groups were highly resistant to cefotaxime and ceftazidime (>92%). Burn isolates were significantly more resistant to cefepime, amoxiclav, imipenem, meropenem, gentamicin and ciprofloxacin compared to the non-burn strains (p<0.05). No significant differences were observed in ESBL production between the two groups. Carbapenem resistance was only observed among the burn isolates (n=5, 9.1%). Five carbapenem-resistant isolates produced carbapenemases. However, none of the isolates harbored the KPC-type genes. Higher rates of drug resistance were observed in burn isolates of K. pneumoniae compared to the non-burnstrains. Carbapenemase phenotype was only observed among the burn isolates but KPC-type gene was not detected.

Escherichia coli O157:H7, an emerging pathogen, causes severe enteritis and the extraintestinal complication of hemolytic-uremic syndrome. The goal of this study was to evaluate the conjugate of E. coli O157: H7 lipopolysaccharide (LPS) with diphtheria toxoid (DT) as a candidate vaccine in mice model. LPS from E. coli O157:H7 was extracted by hot phenol method and then detoxified. Purified LPS was coupled to DT with adipic acid dihydrazide (ADH) as a spacer and 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDAC) as a linker. The coupling molar ratio of LPS to DT was 3:1. Clinical evaluation of E. coli O157:H7 LPS-DT conjugate was also performed. The conjugate was devoid of endotoxin activity and indicated 0.125 U/ml of D-LPS. Mice immunization with D-LPS DT conjugate elicited fourfold higher IgG antibody in comparison to D-LPS. Also, in vivo protection of mice with conjugate provided high protection against the LD50 of E. coli O157:H7, which indicated a good correlation with the IgG titer. Our results showed that the suggested vaccine composed of E. coli O157:H7 LPS and DT had a significant potential to protect against E. coli infections.

Many surface proteins are implicated in nasopharyngeal colonization and pathogenesis of Streptococcus pneumoniae. Some of these factors are candidate antigens for protein based vaccines. New vaccine designs focus on the surface proteins (e. g., pspA and pspC) and also cytolysin, and pneumolysin. In this study, 3 key virulence genes, lytA, pspC, and rrgA, which encoded surface proteins, were detected among S. pneumoniae isolates. A total of 260 nasopharyngeal swabs were collected from healthy children under 6 years old attending day care centers in Mashhad, Iran. Isolates of S. pneumoniae were confirmed by optochin susceptibility and colony appearance and also by PCR for cpsA gene. The presence of lytA, pspC, and rrgA genes were also detected by PCR. A total of 59 isolates were confirmed as S. pneumoniae. Among these isolates, 50 (84.74%), 19 (32.20%), and 2 (3.38%) were positive for lytA, rrgA, and pspC genes respectively. The presence of these genes among S.pneumoniae isolates were as follows: 1) rrgA, lytA, pspC (1 isolate), 2) rrgA, lytA(17isolates), 3) pspC (2 isolate), 4) lytA (50 isolates). cpsA gene was specific for detection of S. pneumoniae isolates which were colonized in nasopharynx. The lytA gene was the most frequent gene among the S. pneumoniae isolates, and combination of rrgA, lytA was the most observed pattern. Thus, it is important for future monitoring of vaccine formulation in our country.

Staphylococcus aureus is one of the main causatives of bovine mastitis. Resistance of some strains to methicillin, can complicate the treatment of its infections. On the other hand, enterotoxin production is also im- portant. Therefore, the aim of our study was to investigate the methicillin resistance and enterotoxin production in S. aureus isolates caused bovine mastitis. Four hundred and fifty milk samples were collected. After isolation of Staphylococcus aureus, MRSA strains were detected by cefoxitin disc diffusion and oxacillin agar screening methods. DNA was extracted by phenol– chloroform method and PCR was applied for mecA, sea and seb genes. SCCmec types of mecA gene were identified using multiplex-PCR. Fifty-four (12%) S. aureus were isolated. Out of these, 10 and 9 MRSA strains identified by cefoxitin disc diffusion and oxacillin agar screening methods, respectively. All 10 MRSA isolates identified by cefoxitin disc diffusion, were positive for mecA gene and all of them belonged to SCCmec type IV. The sea genes were detected in 19 isolates and only two isolates were positive for seb genes. One isolate possessed both sea and seb genes. Findings of this study indicated that results of cefoxitin disc diffusion test is in concordance with the PCR for mecA gene and has a higher sensitivity compared to oxacillin agar screening method. Finally, Our findings suggest that enterotoxin A is the dominant type.

Bacillus anthracis is one of the most homogenous bacteria ever described. Bacillus anthracis 17JB is a laboratory strain. It is broadly used as a challenge strain in guinea pigs for potency test of anthrax vaccine. This work describes genetic characterization of B. anthracis 17 JB strain using the SNPs and MLVA genotyping.Results and In SNPs typing, the originally French 17JB strain represented the A. Br. 008/009 subgroup. In Levy's genotyping method, 843, 451 and 864 bp long fragments were identified at AA03, AJ03 and AA07 loci, respectively. In the vaccine manufacturer perspective these findings are much valuable on their own account, but similar research is required to extend molecular knowledge of B. anthracis epidemiology in Persia.

The aim of the study was to evaluate the clinical effects of a mouthwash containing Sage (Salvia officinalis) extracts on Streptococcus mutans (SM) causing dental plaque in school-aged children. A double blind clinical trial study was conducted in a dormitory on 70 girls aged 11-14 years having the same socioeconomic and oral hygiene conditions. These students were randomly divided into 2 groups; the first group (N=35) using Sage mouthwash, and the second group (N=35) using placebo mouthwash without active any ingredients. At the baseline, plaque samples obtained from the buccal surfaces of teeth were sent to laboratory to achieve SM colony count. These tests were reevaluated after 21 days of using the mouthwashes. Statistical data analysis was performed using t-student tests with p<0.05 as the level of significance. Sage mouthwash significantly reduced the colony count (P=0.001). Average number of colonies in test group was 3900 per plaque sample at the baseline, and 300 after mouthwash application. In the control group, pre-test colony count was 4400 that was reduced to 4000; although this reduction wasn’t significant. The Sage mouthwash effectively reduced the number of Streptococcus mutans in dental plaque.

Photobacterium damselae subsp. damselae is a marine pathogenic bacterium which causes disease in marine animals and human. This bacterium mostly found in coastal shallow seawater. So, the aim of this study was isolation and characterization of Photobacterium damselae subsp. damselae from edible fish of Persian Gulf, Bandar Abbas. Totally 100 fish from different species were evaluated and out of that 5 different types of fish with external symptoms including: Caranx sexfasciatus, Lethrinus olivaceus, Scomberoid tol, Auxis thazard and Liza macrolepis, were collected from Bandar Abbas local fish market in September 2013. The samples were cultured on Marin Agar 2216 and Thiosulfate Citrate Bile salts Sucrose Agar media and incubated at 25˚C for 48 hrs. Then the isolates were characterized using biochemical (API 20 NE system) and molecular techniques. In addition, antibiotic susceptibility, presence of poly β hydroxy butyrate and hemolysis activity of isolates were evaluated. Entirely, 30 Gram negative bacterial colonies were isolated from the selected fish. Among the isolates, two suspected colonies were identified as Photobacterium damselae from Caranx sexfasciatus with API 20NE bio- chemical test. This results confirmed by 16s rRNA sequencing method. Both isolates showed α hemolytic with existence of β hydroxyl butyrate. Furthermore, the isolates were susceptible to ciprofloxacin, chloramphenicol and nalidixic acid. Overall, the study indicated first time isolation of this bacterium from one type of fish caught from Persian Gulf, which warns us to pay more attention to fishery in this geographical area.