Rapid Molecular Approach for Simultaneous Detection of Salmonella spp., Shigella spp., and Vibrio cholera

AbstractObjectivesGastrointestinal tract infection is still one of the serious public health problems in many geographic areas and is endemic in most countries including Iran. Early detection of the gastrointestinal tract pathogens can be extremely important. The aim of the current study was to apply a shortened time-multiplex polymerase chain reaction (PCR) for rapid and simultaneous detection of Salmonella spp., Shigella spp., and Vibrio cholera.MethodsThe standard and clinical strains of Salmonella spp., Shigella spp., and V. cholerae were used in the assay. Multiplex PCR was performed and optimized based on amplification of invA, putative integrase, and ompW genes for detecting Salmonella spp., Shigella spp., and V. cholerae, respectively. The specificity of the assay was evaluated by testing 12 different bacterial species.ResultsOnly Salmonella spp., Shigella spp., and V. cholerae strains had positive results when subjected to the assay using multiplex PCR. The assay showed a high sensitivity, and no amplification products were observed in multiplex PCR with any of the other microorganisms.ConclusionOur study indicated that the invA, putative integrase, and ompW-based multiplex PCR assay appears to be an efficient method for rapid and simultaneous detection of Salmonella spp., Shigella spp., and V. cholerae

Epidemiology of integrons among multidrug-resistant pathogens; an Asian update

Integrons are mobile genetic elements commonly found in the genome of bacteria, within plasmids, chromosomes, and transposons. They are recognized as important factors involved in genetic complexities, the formation of various phenotypes, and the emergence of bacterial adaptation. Integrons play a major role in capturing, expression, and transmission of antibiotic resistance genes and the resistance genes located on integrons can transmit from a strain to another one. Horizontal transmission of integrons is considered as the most important path of resistance genes transmission that leads to the emergence of multidrug resistance species. The genes carrying integrons are found in most Gram-negative pathogens. This review focuses on the epidemiology of classes 1, 2, and 3 integrons among clinically important bacteria in Asian countries

Isolation, Characterization and in vitro Evaluation of Specific Bacteriophages Targeting Extensive Drug Resistance Strains of Pseudomonas aeruginosa Isolated from Septic Burn Wounds

Background: Antibiotic resistant bacteria and various infections caused by them especially extensive drug resistance (XDR) strains and worrying statistics of mortality due to these strains and also the lack of a clear vision for development and production of new effective antibiotics have made the necessity of using alternative therapies more apparent. Materials and Methods: In this study, specific phages affecting the Pseudomonas aeruginosa XDR strain were extracted from hospital wastewater and their laboratory characteristics along with lysis effect on 40 XDR strains of P. aeruginosa were investigated. Results: The results indicated that three isolated phages (PaB1, PaBa2 and PaBa3) belonged to the Myoviridae and Pododoviridae families and were specific to Pseudomonas aeruginosa strains. More than 98% of phages absorbed their host in less than 10 minutes (Adsorption time <10 min) and completed their lytic cycle after 40 minutes (latent time = 40 min). Burst size of PaBa1, PaBa2 and PaBa3 was 240, 250 and 220 pfu/cell, respectively. PaBa1 lysed 62.5% of the XDR strains with the highest efficiency. The three Phage cocktail was effective against 67.5% of the studied strains. Conclusion: The results of this study indicate the significant potential of these phages for therapeutic use and prophylaxis of infections caused by this bacterium. Keywords: Bacteriophage; Pseudomonas aeruginosa; Drug resistance; Antibiotic resistanc

Isolation of new Klebsiella pneumoniae phage PSKP16

Background and Objectives: Klebsiella pneumoniae is a clinically relevant opportunistic pathogen belonging to the Enterobacteriaceae family. It is in the top three bacteria associated with antimicrobial resistance deaths globally, and one of the most dangerous bacteria causing nosocomial infections. Phage therapy offers a potential option for the treatment of drug-resistant bacterial infections. Materials and Methods: Phage PSKP16 was isolated against K. pneumoniae, capsular type K2 (isolated from a wound infection). PSKP16 is a new lytic phage with a Siphovirus-like morphology. Results: PSKP16 is a linear double stranded DNA phage with a GC content of 50% and genome size of 46,712 bp, for which we predicted 67 ORFs. PSKP16 belongs to the genus Webervirus and shows high evolutionary proximity to Klebsiella phages JY917, Sushi, and B1. Conclusion: Phage isolation is fast, cheap and efficient, but it requires time and characterization (which adds expense) to ensure that the isolated phages do not pose a health risk, which is essential to safely use phage therapy to treat life-threatening bacterial infections

Application of bacteriophage cocktails for reducing the bacterial load of nosocomial pathogens in hospital wastewater

Background and Objectives: In the third world and developing countries, hospital sewage is mixed with municipal wastewater. The treated effluent contains dangerous bacteria released into the environment and used in the irrigation of agricultural products, and eventually these bacteria may endanger the human health through foods. Antibiotic-resistant bacteria are mostly found in hospital wastewater. In water and wastewater treatment plants, large amounts of toxic and polluting substances are removed and destroyed, but this process does not eliminate bacteria. Materials and Methods: Wastewater samples from 22 hospitals in Iran were collected and in the meantime specific phages (against drug-resistant pathogenic bacteria) extracted using the bilayer agar technique. Phage amplification was performed by employing a fermenter after phage identification. Amplified phages were added to the primary sedimentation pond using New-Brunwick biofermenter BioFlo/Celligen®115 and the bacterial count was evaluated for the desired bacteria. Results: Our phage cocktail was able to reduce 99.8%, 99.4%, 99.5%, 99.8%, 99.7%, 99.8%, 99.6% and 99.9% of E. coli, E. faecium, E. faecalis, K. pneumoniae, A. baumannii, P. aeruginosa, S. maltophilia and S. aureus counts respectively. Conclusion: The application of phage cocktails can remarkably help improve personal hygiene, the environment, and the optimization of surface wate

Development of multiplex real-time quantitative PCR for simultaneous detection of Chlamydia trachomatis, Mycoplasma hominis, Ureaplasma urealyticum, and Mycoplasma genitalium in infertile women

Purpose: Sexually Transmitted Diseases (STDs) can cause sterility and many other problems for women planning pregnancy. Currently, almost 340 million people worldwide suffer from Sexually Transmitted Infections (STIs). This study made attempts to quickly identify STDs' most critical infectious agents using dedicated primers and probes. Methods: The present study was done on the cervical samples of 200 infertile women. After extracting the total DNA of Chlamydia trachomatis, Mycoplasma hominis, Ureaplasma urealyticum, and Mycoplasma genitalium, quantitative methods were employed to determine the rate of target bacteria using multiplex real-time PCR. Results: The multiplex qPCR showed the rates of 47%, 16%, 46%, and 16.5% for Chlamydia trachomatis, Mycoplasma hominis, Ureaplasma urealyticum, and Mycoplasma genitalium in infertile women, respectively. In some patients, there were co-infections with two or three bacteria. The diagnostic approach used in our research could be employed as an alternative detection tool to identify the four most common STD-associated bacterial agents while detecting mixed infections. Conclusions: Infertile women with no biological problems could have their genital tract checked using this newly designed identification technique and get proper treatment for their infections as quickly as possible

Evaluation of antibacterial activity of fve biocides and the synergistic efect of biocide/ EDTA combinations on bioflm-producing and non-producing Stenotrophomonas maltophilia strains isolated from clinical specimens in Iran

Background: The overuse of biocides in healthcare-facilities poses risk for emergence and spread of antibiotic resistance among nosocomial pathogens. Hospital-acquired infections due to S. maltophilia have been increased in the recent years and with its various resistance mechanisms contribute to patient morbidity and mortality in hospitals. The current study aimed to evaluate the susceptibility of biofilm-producing and non-producing S. maltophilia clinical isolates to five commonly used hospital biocides, alone and in combination with EDTA to examine the synergistic effect of combining EDTA on the bactericidal activity of them by microbroth dilution method. As well as the frequency of efflux genes encoding resistance to biocides among isolates. This study also intended to assess the effect of exposure of S. maltophilia isolates to sub-inhibitory concentrations of sodium hypochlorite upon the antimicrobial susceptibility patterns. Results: Based on minimum inhibitory and bactericidal concentrations of biocides sodium hypochlorite 5% (w/v) and ethyl alcohol 70% (v/v) were the strongest and weakest biocides against S. maltophilia isolates, respectively. The combination of EDTA with biocides significantly increased the effectiveness of the studied biocides. Exposure to subinhibitory concentration of sodium hypochlorite showed a significant change in the susceptibility of isolates towards ceftazidime (p = 0.019), ticarcillin/clavulanate (p = 0.009), and chloramphenicol (p = 0.028). As well as among the isolates examined, 94 (95%) were able to produce biofilm. The frequency of sugE1 resistance genes was found in 90.7% of our clinical S. maltophilia isolates. None of the isolates carried qacE and qacEΔ1 gene. Conclusions: The current study recommended that using the mixture of biocides with EDTA can be effective in reducing nosocomial infections. Also, this study demonstrated that exposure to sub-inhibitory concentrations of sodium hypochlorite leads to reduced antibiotic susceptibility and development of multidrug-resistant S. maltophilia strains

Bacteriophages of Mycobacterium tuberculosis, their diversity, and potential therapeutic uses: a review

Tuberculosis (TB) caused by Mycobacterium tuberculosis (M. tuberculosis) is a highly infectious disease and worldwide health problem. Based on the WHO TB report, 9 million active TB cases are emerging, leading to 2 million deaths each year. The recent emergence of multidrug-resistant tuberculosis (MDR-TB) and extensively drug-resistant tuberculosis (XDR-TB) strains emphasizes the necessity to improve novel therapeutic plans. Among the various developing antibacterial approaches, phage therapy is thought to be a precise hopeful resolution. Mycobacteriophages are viruses that infect bacteria such as Mycobacterium spp., containing the M. tuberculosis complex. Phages and phage-derived proteins can act as promising antimicrobial agents. Also, phage cocktails can broaden the spectrum of lysis activity against bacteria. Recent researches have also shown the efective combination of antibiotics and phages to defeat the infective bacteria. There are limitations and concerns about phage therapy. For example, human immune response to phage therapy, transferring antibiotic resistance genes, emerging resistance to phages, and safety issues. So, in the present study, we introduced mycobacteriophages, their use as therapeutic agents, and their advantages and limitations as therapeutic applications