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Abstract The main purpose was to determine the abundance of dominant phyla, Bifidobacterium spp., and Lactobacillus in breast milk of obese mothers versus normal-weights in fourth month of lactation in Iranian population. Sixty health women at the fourth month of breastfeeding, aged 18–40 years, were included and categorized based on body mass index (BMI) to the obese (BMI ≥ 30 kg/m2) and normal-weights (18.5 ≤ BMI ≤ 24.9). Bacterial DNA was extracted and qPCR of the 16S region was performed after human milk donation in a sterile condition. A multiple linear mixed model was used to determine the effective factors on the phyla population. Bifidobacterium spp. was significantly higher in milk of normal-weight group than the obese. The current weight showed a significant effect on the Actinobacteria abundance in milk. The Bacteroidetes and Firmicutes were significantly lower in mother’s milk with cesarean section (p = 0.04). Pre-pregnancy obesity decreased the Firmicutes and Lactobacillus abundance in maternal milk (p = 0.04 and p = 0.01). The Actinobacteria and Bifidobacterium spp. showed a significant effect on infant’s height (p = 0.008 and p = 0.04). The maternal current and pre-pregnancy weight showed an important effect on abundance of Actinobacteria and Bifidobacterium spp., as the good phyla and genus in milk which are associated with the infant’s height.

Background: Enterococci are main normal microbial flora of both humans and animals and can survive in a diverse range of environments. These bacteria carry out aminoglycoside and vancomycin resistance genes and spread them in environment by many routs such as chicken meat products. The present study was aimed to determine the frequency of aminoglycoside and vancomycin resistant genes in Enterococcus species isolated from chicken meat specimens. Methods: A total of 250 chicken raw meat specimens was prepared from slaughterhouses at Zanjan province, cultured at BHI broth and incubated at 37°C for 24h. The positive cultures were sub-cultured in blood agar plates and grown colonies identified using phenotypical and biochemical tests. Antibiotic susceptibility was determined according to the Clinical and Laboratory Standards Institute (CLSI) standards and PCR assays were performed to detect vanA, vanB, aph (2")1c, aph (2")1b, aph (2")1d, ant(3'), aph(3')IIIa, ant(4')1a, ant(6') and aac(6') genes. Results: In total, 100 Enterococcus species isolated from 250 specimens and 35% of them belonged to E. faecalis and the others were E. faecium (65%). The prevalence of the vanA, vanB, aph (2")1c, aph (2")1b, aph (2")1d, ant(3'), aph(3')IIIa, ant(4')1a, ant(6') and aac(6') genes among the 100 Enterococcus species was 14%, 12%, 10%, 1%, 2%, 50%, 26%, 9%, 18% and 22%, respectively. Conclusion: The current study revealed that the rate of antimicrobial resistance genes to aminoglycosides and vancomycin was worrying and health measurements in meat products industries must be performed to prevent spread of antimicrobial resistance elements among bacteria.

Background: Efflux pumps such as MexEF-OprN and mexXY-OprM play an important role in the resistance of Pseudomonas aeruginosa (P. aeruginosa) to antibiotics. The present study aimed to assess the reduced expression of efflux pump genes of P. aeruginosa with Satureja khuzistanica essential oil (SKEO).Methods: The present cross-sectional study was conducted in 2016 at the Microbiology Laboratory of Baqiyatallah University of Medical Sciences, Tehran, Iran. The disk diffusion method was used for susceptibility testing of gentamicin and norfloxacin. Minimum inhibitory concentration (MIC) was determined for gentamicin and norfloxacin. The antibacterial efficacy of SKEO was defined by determining the MIC values using the microdilution method. In vitro, the synergistic interaction of SKEO combined with gentamicin or norfloxacin was examined via checkerboard assay and defined as a fractional inhibitory concentration index. The reverse transcription-polymerase chain reaction technique was used to measure changes in the expression of the efflux pump genes. The data were analyzed using SPSS software version 16.0, and p Results: The MIC values of SKEO were in the range of 6 to 12 µg/mL. In the presence of sub-inhibitory concentrations (1.16 to 2 MIC) of SKEO, synergistic effects were revealed using the checkerboard method. The effect of norfloxacin and gentamicin increased up to 8-fold. The expression of mexY and mexE was reduced after treatment with SKEO.Conclusion: SKEO reduced the expression of efflux pumps and the MIC values of norfloxacin and gentamicin in vitro.

Abstract Background Bladder cancer is one of the leading causes of cancer death in adults worldwide. There are various risk factors described for the bladder cancer development including genetic background as well as environmental exposure. Currently, infectious agents such as human papilloma virus (HPV) has also been linked to bladder cancer risk. The current study aimed to evaluate the potential correlation between HPV infection and the oncological outcome in urothelial bladder cancer. Methods Totally 106 tissue samples of histopathologically confirmed transitional cell carcinoma (TCC) of the urinary bladder were included in this study. The presence of high risk (types 16 and 18) and low risk (types 11 and 6) types of HPV was evaluated using polymerase chain reaction (PCR) followed by in situ hybridization. Results Out of 106 bladder cancer patients, a total of 24 cases (22.6%) were positive HPV infection. The most common type of HPV detected was type 16 followed by types 11 and 18, and 6. According to independent T-test results, there was a significant association between mean age and HPV infection (P = 0.015). Moreover, our findings showed a significant relation between infection with HPV and tumor stage, tumor grade, muscle invasion of the tumor, as well as tumor recurrence. The results of Chi-square Test indicated that there is significant statistical association between types of HPV and tumor grade (P-Value = 0.044). Conclusion Our findings indicated that a family history of cancer and HPV infection can be potential independent predictive factors for tumor recurrence in bladder cancer. Overall, the results of this study strongly indicate a significant relationship between HPV infection and an aggravated outcome of the disease and a higher risk of recurrence in patients with bladder cancer.

Background and Objectives: The ever-increasing of antibiotic resistance in methicillin-resistant Staphylococcus aureus (MRSA) has become a major threat to public health worldwide. Molecular typing is used to determine the source of MRSA infections as well as to control and prevent the spread of these pathogens. The present study aimed to investigate the characteristics of staphylococcal cassette chromosome mec (SCCmec) types and antibiotic resistance of community- acquired (CA-) and hospital acquired (HA-) MRSA isolates. Materials and Methods: In this cross-sectional study, the antibiotic susceptibility patterns of 109 clinical S. aureus isolates were determined by the Kirby-Bauer disk-diffusion and microdilution broth methods. MRSA isolates were confirmed using the polymerase chain reaction (PCR) method for the detection of the mecA gene. SCCmec typing was performed by a multiplex PCR assay among MRSA isolates. Results: The prevalence of MRSA isolates was 39.4%. Linezolid, vancomycin, and ceftaroline were the most effective agents against MRSA isolates. The incidence of multidrug-resistant (MDR) and resistance to most antibiotics were significantly higher in MRSA than methicillin-susceptible S. aureus (MSSA) isolates (P

Background: The loop-mediated isothermal amplification (LAMP) method is frequently used for identifying many microorganisms. The present review aimed to evaluate the sensitivity and specificity of LAMP method for detection of food-borne bacteria and to compare these features with those of polymerase chain reaction (PCR), as an alternative molecular diagnostic procedure, and with cultivation method, as the gold standard method. Methods: The literature was searched in electronic databases (PubMed, Scopus, Web of Science, and EMBASE) for recruiting publications within Jan 2000 to Jul 2021. We used the combinations of keywords including foodborne disease, LAMP, PCR, Loop-mediated isothermal amplification, and polymerase chain reaction. Meta-analysis was used to adjust the correlation and heterogeneity between the studies. The efficiency of the methods was presented by negative likelihood ratio, positive likelihood ratio, sensitivity, specificity, and odds ratio using forest plots. A P-value less than 0.05 was considered as statistical significance cut off. The confidence intervals were presented at the 95% interval. Results: Overall, 23 relevant studies were analyzed. The sensitivities of LAMP and PCR methods were estimated to be 96.6% (95% CI: 95.0-97.7) and 95.6% (95%CI: 91.5-97.8), respectively. The specificities of LAMP and PCR were also estimated to be 97.6% (95%CI: 92.6-99.3) and 98.7% (95%CI: 96.5-99.5), respectively. Conclusion: The specificities of LAMP and PCR assays were determined by comparing their results with cultivation method as the gold standard. Overall, the specificity of both PCR and LAMP methods was low for detection of fastidious bacteria. Nevertheless, LAMP and PCR methods have acceptable specificities and sensitivities, and their application in clinical practice necessitates more studies.

This study aimed to explore the prevalence, antimicrobial resistance levels, and serotype distribution of S. pneumoniae in the Middle East region. We conducted a systematic literature review by searching several databases including PubMed, ISI Web of Science, Scopus, Google scholar through 2000 to 2017 by using the following keywords: “Streptococcus pneumoniae”, “pneumococcus”, “serotype”, “Antibiotic resistance,” and “Middle East “in combination with “OR” and “AND” Boolean Operators within Title/Abstract/Keywords fields. We used a random-effects model to calculate the pooled prevalence and 95% confidence intervals (CIs) for binomial variables. All statistical analyses were done using STATA 12.0 (STATA Corp, College Station, TX). We found 73 articles appropriate, on the word of inclusion and exclusion criteria, for inclusion in this systematic review and meta-analysis. The result revealed that the pooled prevalence of S. pneumoniae carriage was 35% (95% CI: 26-44%). The most frequent pneumococcal serotypes were19, 19F, 6, 23 and 6A/B which were found in 19%,12%, 11%, 10% and 10% of isolates respectively. Pneumococcal resistance reported for azithromycin, cefaclor, clarithromycin, chloramphenicol, erythromycin, and tetracycline were 24%, 37%, 23%, 11%, 26%, and 29% respectively, while vancomycin resistance was not reported. The highest resistant prevalence was reported against co-trimoxazole (Trimethoprim/sulfamethoxazole). For this antibiotic, a pooled resistance prevalence of 43% was identified. The present review demonstrates that the prevalence of S. pneumoniae carriage was high in the Middle East region. Surveillance must be continued in this region to evaluate. The resistance pattern and serotype distribution.

Background: Listeria monocytogenes show high mortality among pregnant women and newborns. This study aimed to detect L. monocytogenes in pregnant women with a history of abortion and assess the serotypes, antibiotic susceptibility patterns, and its resistance genes. Methods: Overall, 400 vaginal swabs were taken from pregnant women with a history of abortion in the past few years in a tertiary care hospital in Tehran, Iran, during 2015-2018. Antibiotics susceptibility to a panel of 10 antibiotics was determined using the standard disk diffusion method and the isolates serotyped by the agglutination method. The antimicrobial-resistant isolates were also screened for the presence of tetM, ermB and dfrD genes by PCR. Results: Overall, 22 L. monocytogenes isolates were identified. High rates of resistance were observed for trimethoprim (50%; n=11), sulphamethoxazole (50%; n=11), tetracycline (45.45%; n=10) and gentamicin (36.36%; n=8). From 22 L. monocytogenes isolates, 13 (59.10 %), 5 (22.73%), 3 (13.63%) and 1 (4.54%) belonged to serotypes 4b, 1/2a, 1/2b, and 3c, respectively. The genetic determinant tetM was detected in 70% of the tetracycline-resistant isolates. Out of 11 trimethoprim-resistant isolates, 27.27% isolates contained dfrD. Moreover, the ermB gene was found in 83.33% of the erythromycin-resistant isolates. Conclusion: Ampicillin and partly penicillin consider to be suitable antimicrobial agents to treat human listeriosis. Moreover, due to resistance against many antibiotics, it is necessary to continue monitoring and managing antimicrobial resistance.

Background: Efflux pumps are involved in resistance of Acinetobacter baumannii isolates to antimicrobial agents. AdeABC efflux pump is one of the RND superfamily efflux pump and consists of adeA (membrane fusion), adeB (multidrug transporter) and adeC (outer membrane) genes. In this study, the frequency of adeA, adeB and adeC genes among A. baumannii isolates with resistance to erythromycin, trimethoprim, meropenem and imipenem was investigated. Methods: Overall, 79 strains of A. baumannii were isolated from patients admitted to two major hospitals in Tehran during 2016. Antibiotic susceptibility testing was determined by disc diffusion and microdilution methods according to Clinical and Laboratory Standards Institute (CLSI) guideline. The presence of adeA, adeB and adeC genes was also determined using Multiplex PCR assay. Results: The highest and the lowest resistance among A. baumannii isolates were to trimethoprim (93%) and erythromycin (53%), respectively. The frequency of adeA, adeB and adeC genes was 96.2%, 96.2% and 91.1 % respectively. There was a significant relationship between imipenem resistance and presence of efflux pump genes (P

Background and Objectives: Streptococcus pneumoniae causes many lethal infections. Due to its reduced sensitivity to commonly used antibiotics, development of new strategies against pneumococcal infections seems to be necessary. We aimed to investigate immunodominant antigens in S. pneumoniae culture supernatant in order to develop novel targets for pneumococcal vaccines. Materials and Methods: In this study S. pneumoniae ATCC49619 was sub-cultured into BHI broth from overnight culture at 37°C for 4 h. The supernatant proteins were precipitated using acetone precipitation method. A rabbit was intramuscularly immunized with alum adjuvant and 100 μg pneumococcal supernatant proteins, 6 times at 14 days' intervals to produce hyperimmune serum. ELISA assay was performed to determine the antibody level response to pneumococcal secretory proteins. Then dot blot applied for rapid evaluation of hyperimmune serum reactivity to pneumococcus supernatant proteins. The western blot was also used to determine the interaction of supernatant proteins with immunogenic rabbit's hyperimmune-serum. Results: According to the western blot analysis, the immunodominant protein had 140KDa molecular weight and designated as pneumococcal secretory protein140 (Psp140). Conclusion: The Psp140 protein in the supernatant of S. pneumoniae culture is an immunodominant protein and it is likely related to pneumococcal secretory protein or surface exposed protein which released into culture supernatant during bacterial growth.

Background: Traditional dairy products are the main sources for probiotic bacteria. This study aimed to isolate and characterize the potentially probiotic Lactobacillus strains isolated from traditional dairy products in Iran. Methods: Microbial population of each dairy product was enriched and screened for acid- and bile- resistant strains. The isolates were primarily characterized by conventional phenotypic and biochemical tests. The species were finally identified using restriction analysis and 16S rDNA sequencing. In addition, the antibacterial activity of the four most common species was evaluated against seven gastrointestinal pathogens. Results: From a total of nine yogurt and cheese samples, 17 acid-resistant Lactobacillus strains were isolated (survival>50% in PBS buffer, pH 2.5 for 3 h). Eight of them also showed high tolerance to bile. Also, the superiority of molecular techniques over phenotypic tests for effective identification of species was observed. Biochemical tests were useful in combination with 16S-amplified ribosomal DNA restriction analysis (ARDRA) technique to discriminate the isolates at species level. Conclusion: 16S–ARDRA in combination with biochemical tests is suggested as a potentially useful method for accurate identification of Lactobacillus species where the 16S-rDNA sequencing could not discriminate between two closely related species. Considering the acid and bile resistance capacity of isolated strains, traditional dairy products may be used as important sources of probiotic strains.

Background and Objectives: Brucellosis is a widespread zoonotic disease with a high prevalence in both animals and humans. The present study was aimed to evaluate the susceptibility of Brucella strains isolated from human clinical specimens against commonly used antimicrobial agents. Materials and Methods: A total of 360 blood specimens were collected during 2016-2018 and subjected to culture and Brucella spp. identification. The classical biotyping for Brucella isolates was performed according to Alton and coworker's guidelines. Antimicrobials susceptibility test carried out using disk diffusion and minimal inhibitory concentration (MIC) methods. Results: In this study, sixty B. melitensis strains were isolated from blood samples (16%) and all them belonged to biovar 1. Majority of the tested antibacterial agents, excepting ampicillin-sulbactam had an effective activity against B. melitensis isolates in E-test (MIC) and disk diffusion method. Moreover, probable resistance to rifampin and ampicillin-sulbactam were observed in 60 (100%), 1 (1.7%), 11 (18.4%) and 2 (3.4%) isolates, respectively. Conclusion: Our data suggest that the efficacy of commonly used antibiotics for brucellosis treatment should be regularly monitored. In conclusion, appropriate precaution should be exercised in the context of antibiotic administration to prevent future antibiotic resistance.

Background: Klebsiella pneumonia is one of the common causes of pneumonia and bacteremia in intensive care patients. The present study was aimed to determine the capability of (GTG) 5-PCR assay for molecular typing of K. pneumonia strains isolated from patients with urinary tract infections. Methods: In this descriptive-sectional study, K. pneumoniae strains were collected from hospitalized patients with urinary tract infection in Baqiyatallah Hospital, Tehran, Iran during 2017-2018. Isolates were identified by conventional microbiological tests. Bacterial DNA was extracted using boiling method and (GTG) 5-PCR assay was used for subtyping of the isolates. For clustering of isolates, dendrogram was generated according to the unweighted pair group method with arithmetic (UPGMA). Results: Overall, 88 K. pneumoniae isolates were isolated and subjected to the molecular typing study. The (GTG) 5–PCR assay was able to differentiate the K. pneumoniae strains into 9 clusters including G1-G9. Genotype clusters G4 and G9 consist of highest (26) and lowest (1) number isolate, respectively. Conclusion: The K. pneumonia strains isolated under the study belonged to various clones and the (GTG) 5-PCR assay as simple and rapid method can be a powerful tool for molecular typing of K. pneumoniae strains.

Background: Mycoplasma pneumoniae is a common cause of community-acquired pneumonia (CAP) worldwide, especially among children and debilitated populations. The present study aimed to investigate a loop-mediated isothermal amplification (LAMP) technique for rapid detection of M. pneumoniae in clini-cal specimens collected from patients with pneumonia. Methods: Throat swabs were collected from 110 outpatients who suffered from pneumonia. Throat swab samples were obtained from patients referred to the hospital outpatient clinics of Tehran University hospitals, Iran in 2017. The presence of M. pneumoniae in the clinical specimens was evaluated by LAMP, PCR and culture methods. Sensitivity and specificity of the LAMP and PCR assays were also determined. Results: Out of 110 specimens, LAMP assay detected M. pneumoniae in 35 specimens. Detection limit of the LAMP assay was determined to be 33fg /μL or ~ 40 genome copies/reaction. Moreover, no cross-reaction with genomic DNA from other bacteria was observed. Only 25 specimens were positive by the culture method. The congruence between LAMP assay and culture method was ‘substantial’ (κ=0.77). Specificity and sensitivity of LAMP assay were 88.2%, 100% in compare with culture. However, the con-gruence between LAMP assay and PCR assay was ‘almost perfect’ (κ=0.86). Specificity and sensitivity of LAMP assay were 92.5%, 100% in compare with PCR. Conclusion: Overall, the LAMP assay is a rapid and cost-efficient laboratory test in comparison to other methods including PCR and culture. Therefore, the LAMP method can be applied in identification of M. pneumoniae isolates in respiratory specimens.

Background and Objectives: Expressions of lasA and lasB genes of Pseudomonas aeruginosa are associated with bacterium pathogenicity. The present study was aimed to assess the effect of Satureja khuzistanica essential oil (SKEO) extract on expression of lasA and lasB genes in P. aeruginosa. Materials and Methods: Pseudomonas aeruginosa isolates were cultured in Mueller Hinton broth containing sub-inhibitory concentrations of SKEO and total RNA extracted using Trizol method. cDNA was synthesized using random Hexamer primer and finally the expression of lasA and lasB genes carried out by real-time PCR. Results: The MICs of SKEO extract for PA9, PA10, PA11, PA13, PA41 and PA42 isolates were 8, 8, 8, 9, 7 and 12 µg/ml, respectively. Statistical analysis for 6 isolates revealed that the reduction in expression of lasA and lasB genes under SKEO treatment was significant (P

Background and Objectives: Leptospirosis is a zooanthroponosis caused by the genus of Leptospira. It is an emerging public health problem due to its increasing incidence. The achievement to a vaccine that prevent from entrance of Leptospira interrogans to the deeper tissues of the host is needed. This study aimed to investigate the immunogenicity of LcpA (rLcpA) and LenA (rLenA) recombinant proteins in combination with LTB (rLTB) recombinant protein as an adjuvant against leptospiral infection in hamsters. Materials and Methods: The genes encoding these proteins were cloned into pGH cloning vector and then lenA, lcpA and ltb genes subcloned into pET-15b and pET-28a expression vectors, respectively. The hamsters were immunized with the purified recombinant proteins and challenged with Leptospira interrogans for evaluation of their survival. The antibody responses to the recombinant proteins were determined by ELISA. Then, data entered into SPSS software. Statistical Kruskal–Wallis test was used to compare the significant differences among different groups. The groups with significant differences were further analyzed by post hoc tests. The p value < 0.05 statistically was considered significant. Results: Immunized hamsters with rLenA-plus-rLTB, rLcpA-plus-rLTB and rLenA-plus-rLcpA-plus-rLTB proteins showed 60%, 74%, and 80% survival rates, respectively. A significant amount of interleukin-17 (IL-17), interleukin-4 (IL-4) and gamma interferon (IFNγ) cytokines were produced in immunized hamsters. Conclusion: Based on our findings, rLcpA and rLenA proteins in combination with rLTB can protect the hamsters against L. interrogans and effectively induce a protective antibody response. Thus, these proteins can be used as an additional prophylactic tool against leptospira.

Background: Choline-binding proteins (CBPs) are a group of surface-exposed proteins, which play crucial and physiological roles in Streptococcus pneumoniae. The novel member of CBPs, choline-binding protein M (CbpM) may have binding activity to plasma proteins. This study aimed to clone and express CbpM and demonstrate its interaction with plasma proteins and patients’ sera.Methods: The total length of cbpM gene was cloned in pET21a vector and expressed in BL21 expression host. Verification of recombinant protein was evaluated by Western blot using anti-His tag monoclonal antibody. Binding ability of the recombinant protein to plasma proteins and the interaction with patients’ sera were assessed by Western blot and ELISA methods.Results: The cbpM gene was successfully cloned into pET21a and expressed in BL21 host. Binding activity to fibronectin and fibrinogen and antibody reaction of CbpM to patients’ sera was demonstrated by Western blot and ELISA methods, respectively.Conclusion: CbpM is one of the pneumococcal surface-exposed proteins, which mediates pneumococcal binding to fibronectin and fibrinogen proteins.

Introduction: Escherichia coli O157:H7, an important foodborne pathogen, can cause serious renal damage, which can also lead to mortality. Since a rapid and sensitive method is needed to identify this pathogenic agent, we evaluated Loop-Mediated Isothermal Amplification Assay (LAMP) to detect Escherichia coli O157:H7. Methods: We used six primers that specifically identified the rfbE gene. To examine the sensitivity of the method, different dilutions were subjected to the LAMP reaction. Other bacterial strains also were investigated to determine the specificity of the test. The turbidity of the amplified products was assayed by visual detection. The amplified products were detected by addition of SYBR Green II to the reaction tubes. Results: Amplification products were observed as a ladder-like pattern on the agarose gel. A white turbidity emerged in the positive tubes. Under UV light, the positive samples were green, whereas the negative samples were orange. The detection limit of the LAMP was 78 pg/tube, and this indicated that it was 100 times more sensitive than PCR for the detection of EHEC. No LAMP products were detected when template DNA of non- EHEC strains were used, suggesting high specificity of the LAMP assay. Conclusion: The results indicated that the LAMP assay is a valuable diagnostic assay to identify EHEC O157:H7. In addition, the simplicity, sensitivity, specificity, and rapidity of this assay make it a useful method to diagnose pathogens in primary labs without any need for expensive equipment or specialized techniques.

Among many pneumococcal antigens, choline-binding proteins (CPBs) display a high immunogenicity in animal models. This study aims to determine the immunogenicity of CbpM, CbpG and CbpL proteins of Streptococcus pneumoniae in a mice model. The genes were cloned into pET21a expression vector and the recombinant proteins were produced. Mice were immunized with the purified recombinant proteins. Subsequently, the mice were challenged with S. pneumoniae ATCC 49619 (2×106 CFU) and their survival and bacterial clearances were followed 24 hours after infection. The antibody responses of the mice were determined by ELISA assay. The opsonophagocytosis assay was performed using rabbit’s sera. Passive immunization was carried out using two doses of anti-CbPs antibodies. Finally, these mice were experimentally infected with virulent bacteria and the protective effects of two doses of 10 and 100 µg/mL by monitoring the survival rate and bacterial clearance were determined at 2, 3 and 7 days after bacterial challenge. The mice actively immunized with CbpM, CbpG and CbpL recombinant proteins showed survival rate of 100%, 85% and 75%, respectively. The survival rates among passively immunized mice groups which received 100 µg/mL dose of anti-CbpM, anti-CbpG and anti- CbpL were 50%, 50% and 25%, respectively. The rates of opsonization with rabbit’s antibodies against CbpM, CbpG, and CbpL at 100 µg/mL doses was 45.6%, 14.7% and 82.3%, and at 10 µg/mL was 12.9%, 12.2% and 9.35%, respectively. Our findings suggest that the recombinant proteins particularly CbpM and CbpG can protect the mice against pneumococcus19F serotype and effectively induce a protective antibody response. Thus, CbpG and CbpM proteins might be used as suitable vaccine candidate in pneumococcal vaccine formulations.

Background and Objectives: Extended-spectrum β-lactamases (ESBLs) and fluoroquinolones are generally used to treat invasive Salmonella infections, but emergence of antibiotic-resistant strains are increasing worldwide. This study was aimed to investigate the distribution of ESBLs among Salmonella serogroups isolated from pediatric patients in Tehran, Iran. Materials and Methods: The study included all Salmonella isolates recovered from pediatric patients admitted to Children’s Medical Center, Tehran, Iran during 2015-2016. Bacterial isolation and identification were performed by standard biochemical and agglutination tests. Antimicrobial susceptibility testing was done according to the Clinical and Laboratory Standards Institute (CLSI). Polymerase chain reaction was used to identify the genetic determinants responsible for ESBL phenotypes. Results: A total of 138 S. enterica serovars were isolated from stool specimens, including serogroup A (1), serogroup B (18), serogroup C (41) and serogroup D (78). Forty isolates out of 138 Salmonella strains had shown ESBL-positive phenotype. All ESBL-positive isolates showed multiple resistant phenotype. Resistance to more than 3 antimicrobial agents was observed among ESBL-positive strains. The frequency of Salmonella strains carrying the blaCTX, blaTEM and blaSHV genes was 17 (12.3%), 40 (29.9%) and 4 (2.89%) respectively. Conclusion: The high rates of ESBLs positive-Salmonella strains recovered from pediatric patients is alarming and indicates a necessity to substitute the cephalosporins with a proper alternative.

Background: Probiotics mainly belong to Lactobacillus spp. and they are useful for humans in appropriate amounts. The present study aimed to determine antibiotic resistance patterns of such bacteria isolated from probiotic yogurts of Iran. Methods: Probiotic bacteria were isolated from 7 yogurt samples and the isolates were identified by conventional methods and then confirmed using polymerase chain reaction (PCR) technique. Antibiotic susceptibility test was performed by disc diffusion test in order to determine isolates antibiotic resistance patterns. Results: From 7 yogurt samples, 8 isolates were recovered and PCR assay also showed that the isolates belong to Lactobacillus delbrueckii subsp. bulgaricus. Antibiotic susceptibility testing showed that three isolates from three brands were resistant to vancomycin and gentamicin. Other strains were susceptible to all tested antibiotics. Conclusion: Low resistance rate to antibiotics among Iranian probiotic bacteria indicate that the consumption of their products is safe but their clearance following antibiotic therapy can be unfavorable. Hence, applying probiotic strains with non-transferrable resistance elements in probiotic yogurts can be a useful strategy to make stable probiotic products.

Introduction: Brucella spp. are gram-negative, facultative intracellular bacteria pathogens responsible for brucellosis, a zoonotic disease that can cause abortion, fetal death, and genital infections in animals and undulant fever in humans. Lipopolysaccharide (LPS) is known as a major virulence factor of Brucella spp. The wboA gene is capable of encoding a glycosyltransferase that appears to play a major role in LPS biosynthesis. Hence, the characterization of this gene can help in the clarification of the pathogenicity of Brucella spp. Methods: This study was carried out at Razi Vaccine and Serum Research Institute in 2011. Briefly, the wboA gene in B. abortus biovar 3 and B. melitensis biovar 1, the predominant biovars in Iran, were amplified by using two pairs of specific primers. Polymerase chain reaction (PCR) products were cloned into a thymine–adenine (TA) cloning vector and transformed into an E. coli DH5α before being sequenced. Multiple alignments of identified sequences were performed, with all wboA sequences deposited in the GenBank sequence database. Results: This study showed that a mismatch has occurred in B. melitensis biovar 1; this biovar is predominant in Iran. In contrast, the wboA gene from B. abortus biovar 3 was similar to that of other B. abortus variations. Conclusion: The comparison and alignment of the wboA gene of native Brucella strains in Iran to all wboA sequences deposited in GenBank revealed that the wboA gene has changed in the long term; hence, because of its unique nucleotide pattern, the gene can be used for specific diagnosis of B. abortus and B. canis.

No Abstract.

Abstract Background Identification of the microflora of the sand fly gut and the environmental distribution of these bacteria are important components for paratransgenic control of Leishmania transmission by sand flies. Methods Biotic and abiotic bacterial communities of four compartments of a hyper-endemic focus of Zoonotic Cutaneous Leishmaniasis (ZCL) were investigated using 16S ribosomal DNA sequencing and phylogenetic tree construction. These compartments include Phlebotomus papatasi’s gut, skin and intestinal tract of great gerbil Rhombomys opimus, the gerbil nest supplies, and plant food sources of the vectors and reservoirs. Results Sequence homology analysis using nine available 16S rDNA data bases revealed 40, 24, 15 and 14 aerobic bacterial species from the vector guts, the gerbil bodies, the gerbil nests, and the plants, respectively. The isolated bacteria belong to wide ranges including aerobic to facultative anaerobic, pathogen to commensals, sand fly oviposition inducers, land to air and ocean habitats, animal and human probiotics, and plant growth-promoting rhizobacteria. Matching data analysis suggested that the adult P. papatasi gut bacteria could be acquired from three routes, adult sugar feeding on the plant saps, adult blood feeding on the animal host, and larval feeding from nest supplies. However, our laboratory experiment showed that none of the bacteria of the reservoir skin was transmitted to female sand fly guts via blood feeding. The microflora of sand fly guts were associated with the sand fly environment in which the predominant bacteria were Microbacterium, Pseudomonas, and Staphylococcus in human dwellings, cattle farms, and rodent colonies, respectively. Staphylococcus aureus was the most common bacterium in sand fly guts. Presence of some sand fly ovipoisition inducers such Bacillus spp. and Staphylococcus saprophyticus support association between gut flora and oviposition induction. Conclusions Results of this study showed that Bacillus subtilis and Enterobacter cloacae particularly subsp. dissolvens are circulated among the sand fly guts, the plants, and the sand fly larval breeding places and hence are possible candidates for a paratransgenic approach to reduce Leishmania transmission.

Pneumococcal autolysins are enzymes involved in cell wall turnover and cellular division physiologically. They have been found to be involved in the pneumococcus pathogenesis. The aim of this study was to identify the autolytic activity of Spr1754 as a novel protein of Streptococcus pneumoniae. Moreover, the binding of the recombinant protein to plasma proteins was also determined. The spr1754 gene was amplified by PCR and cloned into the pET21a(+) prokaryotic expression vector. The constructed pET21a(+)/spr1754 recombinant plasmid was transformed into E. coli Origami (DE3) and induced using IPTG. The recombinant protein of Spr1754 was purified by Ni-NTA affinity chromatography and confirmed by SDS-PAGE and Western blot analysis using anti-His tag monoclonal antibody. Autolytic activity and the ability of the recombinant protein in binding to plasma proteins were performed using zymogram analysis and western blot, respectively. The spr1754 with expected size was cloned and overexpressed in Escherichia coli Origami (DE3), successfully. After purification of the Spr1754 recombinant protein, the autolytic activity was observed by zymography. Of the four plasma proteins used in this study, binding of lactoferrin to Spr1754 recombinant protein was shown. The Spr1754 recombinant protein has a bifunctional activity, i.e., as being autolysin and lactoferrin binding and designated as Aap (autolytic/ adhesion/ pneumococcus). Nevertheless, characterization of the Aap needs to be followed using gene inactivation and cell wall localization.

Background and Objectives: Loop-mediated isothermal amplification is a novel nucleic acid amplification assay providing as a simple diagnostic tool for rapid identification of microbial diseases in developing countries. In this study, a LAMP assay was established for Yersinia enterocolitica, a leading cause of acute enterocolitis in young children. Materials and Methods: LAMP assay was established with four primers targeting a specific locus for the detection of Y. enterocolitica. The assay was conducted at 65°C in thermo block for 90min. The sensitivity of LAMP was evaluated in com- parison to conventional PCR using pTZ57R containing the target locus. Finally, specificity was assessed using DNA from common enteropathogenic bacteria. Results: Results showed that the sensitivity of LAMP assay was 44-copy number, which was 10-fold higher than that of PCR. No cross-reactivity was observed when testing against other enteropathogenic pathogens. Conclusion: This study showed that LAMP assay is an alternative molecular diagnostic tool for infections with Y. enteroco- litica. In addition, this method may be useful in diagnosis at field or in laboratories without PCR machine.

Shigella species are among the common causes of bacterial diarrhoeal diseases. Traditional detection methods are time-consuming resulting in delay in treatment and control of Shigella infections thus there is a need to develop molecular methods for rapid and simultaneous detection of Shigella spp. In this study a rapid multiplex PCR were developed for simultaneous detection of three pathogenic Shigella species.For detection of Shigella spp., a pair of primers was used to replicate a chromosomal sequence. Three other sets of primers were also designed to amplify the target genes of three most common species of Shigella in Iran including S. sonnei, S. flexneri and S. boydii. The multiplex PCR assay was optimized for simultaneous detection and differentiation of three pathogenic Shigella species. The assay specificity was investigated by testing different strains of Shigella and other additional strains belonging to non Shigella species, but responsible for foodborne diseases.The Shigella genus specific PCR yielded the expected DNA band of 159 bp in all tested strains belonging to four Shigella species. The standard and multiplex PCR assays also produced the expected fragments of 248 bp, 503 bp, and 314 bp, for S. boydii, S. sonnei and S. flexneri, respectively. Each species-specific primer pair did not show any cross-reactivity.Both standard and multiplex PCR protocols had a good specificity. They can provide a valuable tool for the rapid and simultaneous detection and differentiation of three most prevalent Shigella species in Iran.

Background: Microbes particularly bacteria presenting in the gut of haematophagous insects may have an important role in the epidemiology of human infectious disease. Methods: The microbial flora of gut and surrounding environmental of a laboratory strain of Phlebotomus papatasi, the main vector of Zoonotic Cutaneous Leishmaniasis (ZCL) in the old world, was investigated. Biochemical reac- tions and 16s rDNA sequencing of the isolated bacteria against 24 sugars and amino acids were used for bacteria species identification. Common mycological media used for fungi identification as well. Results: Most isolates belonged to the Enterobacteriaceae, a large, heterogeneous group of gram-negative rods whose natural habitat is the intestinal tract of humans and animals. Enterobacteriaceae groups included Edwardsiella, Enterobacter, Escherichia, Klebsiella, Kluyvera, Leminorella, Pantoea, Proteus, Providencia, Rahnella, Serratia, Shigella, Tatumella, and Yersinia and non Enterobacteriaceae groups included Bacillus, Staphylococcus and Pseu- domonas. The most prevalent isolates were Proteus mirabilis and P. vulgaris. These saprophytic and swarming mo- tile bacteria were isolated from all immature, pupae, and mature fed or unfed male or female sand flies as well as from larval and adult food sources. Five fungi species were also isolated from sand flies, their food sources and colo- nization materials where Candida sp. was common in all mentioned sources. Conclusion: Midgut microbiota are increasingly seen as an important factor for modulating vector competence in insect vectors so their possible effects of the mirobiota on the biology of P. papatasi and their roles in the sandfly- Leishmania interaction are discussed.

Introduction: Mycobacterium avium subsp. paratuberculosis causes paratuberculosis (Johne's disease), a systemic infection and chronic inflammation of the intestine that affects many species, including bovine. Infection is widespread in livestock, and human populations are exposed. A possible association between MAP infection and Crohn's disease in humans has been also described. Effective control of paratuberculosis has hampered due to lake of rapid and accurate diagnostic test. Range of diagnostic tests is available, but all have inborn limitations. The present study was designed to develop a loop-mediated isothermal amplification (LAMP) assay for the rapid and simple detection of Mycobacterium avium subsp. paratuberculosis (MAP). Materials and Methods: Six primers were specially designed for recognizing eight distinct sequence of insertion sequence 900 (IS900). To determine the sensitivity of the LAMP assay, 10-fold serial dilutions were made from 431ng/ μl MAP stock solution and compared with Nested-PCR results obtained using similar templates at identical concentrations. Detection limit of the LAMP was defined as the last positive dilution and the reactions were performed four times to examine the reproducibility of the test. The specificity of the assays were evaluated by testing three Gram-positive bacteria including Mycobacterium bovis AN5, Mycobacterium tuberculosis DT and Mycobacterium avium avium. Results: Sensitivity of this assay for detection of DNA of MAP was 4fg/ μl and the specificity was 100%. This assay successfully detected MAP not only in the bacterial cultures but also in clinical fecal samples and the specificity of both PCR was 100%. This LAMP method is performed under isothermal conditions and no special apparatus is needed. In addition, its reactivity is directly observed with the naked eye without electrophoresis either as turbidity or in the form of a color change when SYBR Green 1, a fluorescent dsDNA intercalating dye, is employed. Conclusions: This assay is rapid which requires nearly 1 h for detection of MAP, low in cost and simple to perform, sensitive and practical tool for the detection of MAP and will be useful in facilitating the early diagnosis of paratuberculosis (Johne's disease) caused by the organism.

Background and Objective: The present study aimed to identify and prioritize the factors affecting the desire to leave the job of employees of a health center in Ahvaz City, Iran. Materials and Methods: A descriptive-analytical study was conducted in health centers in Ahvaz in 2023. The factors influencing the desire to leave a job were identified through a literature review and the fuzzy Delphi method. Additionally, 100 employees completed Kim's Turnover Tendency Scale. In the final step, the identified factors were weighted and prioritized using the Analytical Hierarchy Process (AHP). Results: A total of 32 effective factors influencing the desire to leave work were identified and categorized into four groups: individual factors, organizational factors, job factors, and job-related factors. The findings from the AHP method indicated that age (0.213), education level (0.176), and work experience (0.143) from the category of individual factors, as well as salary, type of employment, and lack of promotion opportunities from the category of organizational factors, leadership and management style (0.243), changes in job descriptions (0.243), and organizational position (0.198), along with the employment of unqualified individuals (0.147) from the category of managerial factors, were identified as the most significant risk factors predicting the intention to leave a job among employees in health centers. Conclusion: By identifying and prioritizing the factors affecting the desire to leave the job among the employees of health centers, preventive measures can be defined and implemented to maintain the human resources providing health services.