Rapid detection and simultaneous identification of the Mycoplasma and Ureaplasma species by real-time PCR and melt curve analysis among fertile and infertile females

Objective(s): Mycoplasma and Ureaplasma species threaten reproductive health and fertility worldwide. Due to the lack of sensitive, accurate, and affordable diagnostic tools, the simultaneous contributions of these agents in infertility have been overlooked. This study aims to detect and identify Mycoplasma and Ureaplasma species in the genital tracts of fertile and infertile females simultaneously.Materials and Methods: In a case-control study, cervicovaginal clinical samples were collected from patients referred to two teaching hospitals in Isfahan from July 2019 to February 2019. The initial screening was by using Real-time PCR and designed primer to evaluate the presence of Mycoplasma and Ureaplasma species including fertile and infertile women. The bacteria species were then detected and differentiated by using the melt curve and sequenced to confirm and identify. Finally, the standard curve was used to measure and compare the copy number of each species in each group. The isolates also were detected in clinical samples using the commercial PCR method.Results: The frequencies of Mycoplasma genitalium and Mycoplasma hominis were (0.0, 10.0%) in the fertile group and (4.3%, 34.3%) in the infertile group, respectively. Ureaplasma parvum and Ureaplasma urealyticum species in the fertile group (7.1%, 5.7%) and in the infertile group (32.9%, 24.3) were determined, respectively. The comparison of the results obtained from PCR and Real-time PCR showed that the recent technique has the ability to track 101–103 copy numbers.Conclusion: The present method allows differential diagnosis and quantification of Mycoplasma and Ureaplasma species in a short time and simultaneously

Frequency of mecA Gene in the Clinical Isolates of Staphylococcus epidermidis in Isfahan, Iran

and people with permanent prostheses. Its increased resistance to antibiotics has created a serious challenge for healthcare system. This study was conducted to determine the pattern of antibiotic-resistance of S. epidermidis isolates from clinical samples. Methods:   During nine months, 251 clinical samples isolated from strains of S. epidermidis were examined. Following identification of isolates, their antibiotic sensitivity was determined using disc diffusion method. Resistance to vancomycin was assessed using agar screening method, and its MIC values were measured using episilometry (E-test). Methicillin-resistant gene (mecA) was traced using PCR.        Results:    A total of 120 S. epidermidis strains were isolated from the 251 clinical samples, mostly associated with urine samples. In this study, 95 isolates (79%) were found resistant to cefoxitin, 66 (55%) to vancomycin, and 94 (78.33%) to multiple drugs. In molecular assessment, 37 isolates (54.41%) contained mecA gene, of which, 32 isolates showed resistance to vancomycin. Conclusion:   Increased resistance to methicillin and vancomycin in S. epidermidis isolates represents a serious warning to the healthcare system. Thus, careful and appropriate choice of treatment is imperative for reducing medication resistance

Comparative Analysis of Three Methods for Determination of Imipenem Resistance in Pseudomonas aeruginosa

Background: These days, the antibiotic resistance of Pseudomonas aeruginosa isolates toimipenem has significantly increased. Therefore the study of resistance to imipenem in thisorganism to imipenem in determining the appropriate treatment is crucial and necessary. The goalof this study is to compare three phenotypic methods of E-test, disk diffusion and micro brothdilution in the study of resistance to imipenem in clinical isolates of P. aeruginosa.Methods: Within a 6-month interval, 120 clinical specimens were collected and evaluated. Allisolates were identified as P. aeruginosa by standard biochemical tests and amplification of 16SrRNA gene. Three phenotypic methods of E-test, disk diffusion, and micro broth dilution wereused to determine imipenem resistance in P. aeruginosa isolates.Results: Of the 96 P. aeruginosa isolates studied for their resistance to imipenem by the use ofE-test, disk diffusion and micro broth dilution methods, 38.5% of the strains in micro broth dilutionmethod and 33.3% in the two methods of E-test and disk diffusion were resistant to imipenem. Therate of sensitivity and specificity of disk diffusion and E-test methods were 100%, 90.1%, and theywere 100% and 83.1% for micro broth dilution, respectively.Conclusions With regard to the results obtained from the comparison of the three methods100% agreement were observed among the antimicrobial susceptibility results obtained by the Etest and disk diffusion methods (P ≥ 0.05). Therefore, the use of disk diffusion method can bean appropriate replacement for E-test method with regard to its being easy and cost-effective.</span

Evaluation of the Simultaneous Effects of Lactobacillus delbrueckii and L. lactis on Biofilms of Isolates from Chronic Ulcer Infections with Multiple-drug Resistance

Background: Bacterial biofilm is a major barrier to chronic wound healing. Therefore, the prevention of biofilm formation has an effective role in accelerating the healing of these wounds. Today, probiotics’ anti-biofilm and antibacterial activity have been proven, and bacteriotherapy by probiotics is a new strategy for treating chronic ulcer infections. Objectives: The present study aimed to investigate the synergistic effects of Lactobacillus delbrueckii and L. lactis on biofilms of bacterial agents isolated from these ulcers in the human plasma biofilm model (hpBIOM). Methods: This study examined 82 specimens of chronic ulcer biofilms and identified bacterial isolates using phenotypic and molecular methods. After preparing the hpBIOM, 50 µL of each probiotic (109 CFU/mL) was added in two doses separately and simultane-ously. After 24 hours, 1 mL of bromelain (0.1 g/mL) was added to the complex and incubated at 37°C for two hours. Then, the surviving bacterial cells were counted by serial dilutions. Results: Among 119 bacterial isolates, Staphylococcus aureus (19%), Escherichia coli (17.0%), and Pseudomonas aeruginosa (14%) were the most common bacterial isolates. Lactobacillus delbrueckii showed anti-biofilm activity against multiple-drug resistance pathogens, Staphylococcus, P. aeruginosa, and K. pneumoniae. Although L. lactis had anti-biofilm activity against these three pathogens, its effect was less than that of L. delbrueckii. The two probiotics did not have any synergistic effect on the biofilms of the isolates. Conclusions: The results of the present study emphasized the potential of probiotics in destroying biofilms of isolates with multiple-drug resistance; however, their simultaneous use for this purpose requires further investigation

Phylogenetic analysis of human bocavirus in children with acute respiratory infections in Iran

Human bocavirus (HBoV) was first characterized in nasopharyngeal aspirates from young children with acute respiratory infections. It is prevalent among children with acute wheezing. This study was carried out in order to analyze the infection frequency and coinfection rates of HBoV with respiratory syncytial virus (RSV) and to perform phylogenetic analysis of HBoV in samples of children with acute respiratory infection in Isfahan, Iran. During the time period 2016–2017, altogether 75 respiratory samples from children hospitalized with acute respiratory infection were collected. The samples were first screened for RSV by direct immunofluorescence method and then subjected to detect HBoV DNA by PCR. Genotyping of HBoV-positive samples was conducted by direct sequencing of PCR products using NP and VP1/VP2 genes. Out of 75 respiratory samples, 20 (26.7%) and 10 (13.3%) were positive for RSV and HBoV, respectively. The coinfection rate was 40% (p = 0.048). Considering the seasonal distribution, winter has the highest extent outbreak (p = 0.036). Sequence analysis of positive samples exhibits that all of the isolated HBoV were related to genotype 1 (HBoV-1) with minimal sequence variations. Increasing frequency of HBoV suggests that the virus is related to acute respiratory infection in children. A single genetic lineage of HBoV1 seems to be the major genotype in Iran