Evaluation of the therapeutic effect of a novel bacteriophage in the healing process of infected wounds with Klebsiella pneumoniae in mice.

OBJECTIVE: Bacterial wound infections have recently become a threat to public health. The emergence of multidrug-resistant (MDR) strains of Klebsiella pneumoniae highlights the need for a new treatment method. The effectiveness of bacteriophages has been observed for several infections in animal models and human trials. In this study, we assessed the effectiveness of bacteriophages in the treatment of wound infections associated with MDR and biofilm-producing K. pneumoniae and compared its effectiveness with that of gentamicin. METHODS: A lytic phage against MDR K. pneumoniae was isolated and identified. The effectiveness of phages in the treatment of wound infection in mice was investigated and its effectiveness was compared with gentamicin. RESULTS: The results showed that the isolated phage belonged to the Drexlerviridae family. This phage acts like gentamicin and effectively eliminates bacteria from wounds. In addition, mice in the phage therapy group were in better physical condition. CONCLUSION: Our results demonstrated the success of phage therapy in the treatment of mice wounds infected with K. pneumoniae. These results indicate the feasibility of topical phage therapy for the safe treatment of wound infections.

Evaluation of ethanol and EDTA concentrations in the expression of biofilm-producing smf-1, rpfF genes in XDR clinical isolates of Stenotrophomonas maltophilia.

BACKGROUND: Stenotrophomonas maltophilia is able to cause infections in immunocompromised patients, and the treatment of this opportunistic pathogen is complicated due to its virulence factors, antibiotic resistance, and the ability of the bacteria to produce biofilm. The main goals of this study were to assess the susceptibility of extensively drug-resistant (XDR) isolates to ethanol and EDTA, and evaluating the synergistic effect of these disinfectants, and also survey the effect of exposure to sub-inhibitory concentrations of ethanol and EDTA on the expression of biofilm-producing smf-1, rpfF genes. RESULTS: The results showed that EDTA significantly increased the effectiveness of the ethanol and have a synergistic effect. All of the 10 XDR isolates included in the current study harbored smf-1 and rpfF genes and produced biofilm. After exposure to MIC, sub-MIC, synergism, and sub-synergism of ethanol and EDTA, the expression of smf-1 and rpfF genes was repressed significantly. CONCLUSION: In the current study, it was indicated that the expression of biofilm-producing genes was repressed when bacteria are exposed to different concentrations of ethanol and EDTA. Future studies should include more complex microbial communities residing in the hospitals, and more disinfectants use in hospitals. Expression of other virulence genes in different conditions is suggested.

Characterization of novel bacteriophage PSKP16 and its therapeutic potential against ß-lactamase and biofilm producer strain of K2-Hypervirulent Klebsiella pneumoniae pneumonia infection in mice model.

BACKGROUND: Severe infections caused by ß- lactamase producers, hypervirulent Klebsiella pneumoniae (BhvKp) with K2 serotype, highlight emergency need for new therapeutic strategies against this pathogen. We aimed to assess the efficacy of a novel phage, PSKP16, in the treating of pneumonia induced by BhvKp in mice models. METHOD: Genome sequences of PSKP16 were analyzed, and associated information can be found in NCBI. We applied treatment in two ways: by using mice for immediate and delayed treatments. Moreover, acute pneumonia obtained by BhvKp with intranasal method, was characterized in terms of histopathology of pulmonary lesions, biomarkers of inflammation level, leukocytes cells infiltration extent in mice, and was assessed treatment of them with PSKP16 multiplicity of infection (MOI: 10), either individually or in combination with gentamicin. Assessment of the ability of PSKP16 to inhibit BhvKp biofilm was studied. RESULTS: PSKP16 was associated with the Drexlerviridae family, and had a genome size of 46,712 bp, and 67 predicted ORFs. Herein, prompt phage administration's efficacy to decrease bacterial load and improve the survival rate in pneumonia models was faster than the synergism model with delay, but both almost displayed similar endpoints. The distribution of BhvKp strains in the lung was consistent with the histopathological findings, simultaneous inflammation, and level of serum tumor necrosis factor-α (TNF α). The phage treatment presented a lack of severe lesions and alveolar edema, reduction of inflammatory cell infiltration, which not only was it not associated with an over-inflammation but also provided a faster correction of blood cell count abnormalities compared to gentamicin. Phage with a high concentration in in vitro model effectively eliminated biofilms. CONCLUSION: It is essential to raise clinical awareness and management of BhvKp infections, signaled as the next superbug in waiting. The results of our study underscore the importance of PSKP16 as a phage with promising therapeutic potential in treating BhvKp-induced pneumonia.

Biofilm formation, antimicrobial resistance genes, and genetic diversity of Salmonella enterica subspecies enterica serotype Enteritidis isolated from food and animal sources in Iran.

OBJECTIVES: Salmonella enterica serovar Entritidis is an important pathogen in foodborne diseases and causes gastroenteritis. Several studies have investigated the genetic diversity of the strains of this bacterium. However, our knowledge of the discriminatory power of the molecular methods is limited. METHODS: In total, 34 strains of S. enteritidis were isolated from food related to animals. Antibiotic resistance of the strains, antibiotic resistance genes, and biofilm formation capacity of the strains were evaluated. For the genetic analysis of the strains, PFGE was performed using AvrII restriction enzyme. RESULTS: Among the tested antibiotics, cefuroxime, nalidixic acid, and ciprofloxacin showed the highest resistance rates (79.4%, 47%, and 44.2%, respectively). Only three antibiotic-resistance genes were identified in these strains (blaTEM: 67.6%, tetA: 9%, and sul2: 3%). In total, 91% of the strains were biofilm producers. Clustering of strains using AvrII for 26 samples with the same XbaI PFGE profile showed that these strains were in one clone and had high homogeneity. CONCLUSIONS: In conclusion, it is better to use a combination of several typing methods for typing strains that are genetically very close so that the results are reliable.

Isolation of Sphingomonas paucimobilis from an ocular infection and identification using ribosomal RNA gene: First case report from Iran.

Key Clinical Message: Sphingomonas paucimobilis can cause infection in healthy people. As this bacterium is slow-growing, special attention should be paid to the timely diagnosis and control of its antibiotic resistance to prevent the spread of resistant strains. Abstract: This study reports a case of ocular infection caused by Sphingomonas paucimobilis and its treatment with various antibiotics. A middle-aged woman with prolonged purulent eye discharge was admitted to an ophthalmology clinic in Qazvin, Iran. A strain of S. paucimobilis was isolated from the patient. The sample was identified by Sanger sequencing of the 16s rRNA gene, and an antibiogram test was performed to determine its resistance profile. The patient was treated with ceftazidime and levofloxacin eye drops. The bacterial culture was negative 18 days after starting ceftazidime and levofloxacin treatment. The antibiogram results showed that the isolated bacterium was resistant to aminoglycosides and colistin. This study highlights that S. paucimobilis can cause disease even in immunocompetent individuals. Due to the different resistance profiles of this bacterium, treatment should be based on antibiogram results.

Status of Crimean-Congo haemorrhagic fever virus in ticks in Iran: A systematic review with meta-analysis.

Crimean-Congo haemorrhagic fever (CCHF) is a re-emerging viral haemorrhagic fever causing outbreaks in Iran in the last 15 years. In this systematic review and meta-analysis, the status of Crimean-Congo haemorrhagic fever virus (CCHFV) in ticks would be evaluated. PubMed, Google Scholar and Web of Science were searched for peer-reviewed original papers published between 2000 and 1 July 2022. We included papers that evaluated the prevalence of CCHFV in individual ticks using reverse transcription polymerase chain reaction (RT-PCR). The pooled prevalence of CCHFV was 6.0% (95% confidence interval [CI]: 4.5-7.9%), with heterogeneity (I2 = 82.706; P < 0.0001). The prevalence of CCHFV was higher related to regions with above sea level of 1001-1500m (6.4%; 95% CI: 4.3-9.5%), an average temperature of ≤15 °C (8.3%; 95% CI: 5.6-12.0%), latitude of ≥36° (8.1%; 95% CI: 5.2-12.3%), an annual rainfall of 101-300 mm (9.8%; 95% CI: 6.1-15.4%) and humidity of ≥61% (10.2%; 95% CI: 5.1-19.3%). Due to the importance of CCHF, it is better to do new epidemiologic studies on ticks by related organizations and adjacent regions of some provinces in which human cases have been previously reported.

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.

In Vitro Activity of Fosfomycin on Multidrug-Resistant Strains of Klebsiella pneumoniae and Klebsiella oxytoca Causing Urinary Tract Infection.

With the emergence of multi-drug resistant strains among Klebsiella isolates, the use of old drugs such as fosfomycin has been considered. In this context, we investigated the effect of fosfomycin on biofilm-producing Klebsiella pneumoniae and Klebsiella oxytoca strains isolated from ICU patients. A total of 90 isolates of Klebsiella pneumoniae and 30 isolates of Klebsiella oxytoca were collected from the ICU ward. All isolates were confirmed by biochemical and genotypic methods. Antibiotic susceptibility testing was performed by disc diffusion method and for fosfomycin and colistin, minimum inhibitory concentration (MIC) was done using micro broth dilution. The presence of the beta-lactamase encoding genes, biofilm-related genes, and fosfomycin resistance-related genes was detected by PCR. Finally, for fosfomycin-resistant isolates, we determined the sequence type by the MLST method. Sensitivity rate to fosfomycin in Klebsiella pneumoniae and Klebsiella oxytoca isolates was 92.2% and 100%, respectively. Fosfomycin was the most active antimicrobial agent with 96% sensitivity among all tested antibiotics. All tested isolates could produce biofilm. The frequency of biofilm-related genes for Klebsiella pneumoniae was as follows: 95.5% fimH, 86.6% mrkD, 77.7% mrkA, and 50% wcaG. The frequency of these genes for Klebsiella oxytoca was as follows: 56.6% fimA, 46.6% mrkA, 93.3% matB, and 90% pilQ. Only 4.4% of Klebsiella pneumoniae isolates showed resistance to fosfomycin, and the fosA gene was detected in all of them. Our results showed that fosfomycin effectively inhibits multidrug-resistant (MDR) strains of Klebsiella pneumoniae and Klebsiella oxytoca.

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 effective 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.

Evaluation of antibacterial activity of five biocides and the synergistic effect of biocide/EDTA combinations on biofilm-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 sub-inhibitory 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.

Activity of meropenem-vaborbactam against different beta-lactamase producing Klebsiella pneumoniae and Escherichia coli isolates in Iran.

We evaluated the activity of meropenem-vaborbactam against different beta-lactamase producing Klebsiella pneumoniae and Escherichia coli isolates. In our study antibiotic susceptibility testing, double disk synergy test, modified Hodge test were applied. Detection of ESBL, AmpC, and carbapenemase genes was performed by PCR. Multilocus sequence typing (MLST) analysis was done on OXA-48 producing K. pneumoniae strains. Our results showed that among E. coli and K. pneumoniae isolates, 41.1% and 40% of strains produced ESBL, respectively. Additionally, the prevalence of AmpC producing K. pneumoniae and E. coli was 4% and 45.5%, respectively. Altogether 64.2% of K. pneumoniae strains and one E. coli isolate produced carbapenemase. Among OXA-48 producing K. pneumoniae strains ST3500 and ST2528 were detected by MLST. Based on the phenotypic results of this study, vaborbactam was an effective inhibitor on the third-generation cephalosporin-resistant isolates (P < 0.0001). Meropenem-vaborbactam combination had the highest efficacy on KPC producing strains, and it had limited activity on isolates producing OXA-48 type beta-lactamases, whereas no effect was observed on NDM-1 producing isolates. Our study provided valuable information regarding the vaborbactam inhibitory effect on ß-lactamase-producing strains.

Evaluation of mechanisms of colistin resistance in Klebsiella pneumoniae strains isolated from patients with urinary tract infection in ICU.

Background and Objectives: One of the major causes of urinary tract infections is Klebsiella pneumoniae. Currently, few studies investigated the mechanisms of resistance to colistin in Iran. The current study aimed to determine the prevalence of plasmid and chromosome-mediated resistance to colistin in K. pneumoniae isolates. Materials and Methods: 177 urine samples were collected from patients with urinary tract infections hospitalized in the intensive care unit (ICU) of hospitals in the city of Qazvin. K. pneumoniae isolates were identified by standard biochemical methods, resistance to colistin among K. pneumoniae isolates were tested by disk diffusion and microbroth dilution methods. The chromosomal mutation and presence of the mcr genes in colistin-resistant K. pneumoniae were evaluated by PCR. Results: Out of 177 samples, 65 K. pneumoniae were obtained from patients in the ICU. Six colistin-resistant isolates were isolated with MIC values ≥4 µg/mL, none of them was positive for mcr1-5. In 4 isolates, missense mutation in mgrB gene resulted in amino acid substitutions and in one isolate of mgrB gene was found intact mgrB gene. Conclusion: The results suggest that mgrB mutation was the main mutation among colistin-resistant isolates and plasmid-borne colistin resistance was not expanded among strains.

Phenotype and genetic determination of resistance to common disinfectants among biofilm-producing and non-producing Pseudomonas aeruginosa strains from clinical specimens in Iran.

BACKGROUND: Pseudomonas aeruginosa is a common pathogen in Hospitalized patients, and its various resistance mechanisms contribute to patient morbidity and mortality. The main aims of the present study were to assess the susceptibility of biofilm-producing and non-producing P. aeruginosa isolates to the five commonly used Hospital disinfectants, to evaluate the synergistic effect of selected disinfectants and Ethylene-diamine-tetra acetic acid (EDTA), and the effect of exposure to sub-inhibitory concentrations of Sodium hypochlorite on antimicrobial susceptibility test. RESULTS: The results showed that sodium hypochlorite 5% and Ethanol 70% were the most and least effective disinfectants against P. aeruginosa, respectively. The addition of EDTA significantly increased the effectiveness of the selected disinfectants. The changes in the antibiotic-resistance profiles after exposure to sub-inhibitory concentrations of disinfectants were observed for different classes of antibiotics (Carbapenems, Aminoglycosides, Cephalosporins, Fluoroquinolones). As well as near the all isolates harbored efflux pump genes and 117 (97.5%) of isolates produced biofilm. CONCLUSION: In the current study, the mixture of disinfectant and EDTA were the most suitable selection to disinfect Hospital surfaces and instruments. Also, it was clear that exposure to sub-inhibitory concentrations of Sodium hypochlorite results in resistance to some antibiotics in P. aeruginosa species. Strong and intermediate biofilm formers belonged to MDR/XDR strains. Future studies should include more complex microbial communities residing in the Hospitals, and more disinfectants use in Hospitals.

Diagnostic Accuracy of Rapid Antigen Tests for COVID-19 Detection: A Systematic Review With Meta-analysis.

Introduction: Reverse transcription-polymerase chain reaction (RT-PCR) to detect SARS-CoV-2 is time-consuming and sometimes not feasible in developing nations. Rapid antigen test (RAT) could decrease the load of diagnosis. However, the efficacy of RAT is yet to be investigated comprehensively. Thus, the current systematic review and meta-analysis were conducted to evaluate the diagnostic accuracy of RAT against RT-PCR methods as the reference standard. Methods: We searched the MEDLINE/Pubmed and Embase databases for the relevant records. The QUADAS-2 tool was used to assess the quality of the studies. Diagnostic accuracy measures [i.e., sensitivity, specificity, diagnostic odds ratio (DOR), positive likelihood ratios (PLR), negative likelihood ratios (NLR), and the area under the curve (AUC)] were pooled with a random-effects model. All statistical analyses were performed with Meta-DiSc (Version 1.4, Cochrane Colloquium, Barcelona, Spain). Results: After reviewing retrieved records, we identified 60 studies that met the inclusion criteria. The pooled sensitivity and specificity of the rapid antigen tests against the reference test (the real-time PCR) were 69% (95% CI: 68-70) and 99% (95% CI: 99-99). The PLR, NLR, DOR and the AUC estimates were found to be 72 (95% CI: 44-119), 0.30 (95% CI: 0.26-0.36), 316 (95% CI: 167-590) and 97%, respectively. Conclusion: The present study indicated that using RAT kits is primarily recommended for the early detection of patients suspected of having COVID-19, particularly in countries with limited resources and laboratory equipment. However, the negative RAT samples may need to be confirmed using molecular tests, mainly when the symptoms of COVID-19 are present.

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.

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.

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 water.

Amniotic Membrane Transplantation for Persistent Epithelial Defects and Ulceration due to Pseudomonas Keratitis in a Rabbit Model.

PURPOSE: The use of amniotic membrane has been suggested in the treatment of infectious keratitis for its intrinsic anti-infective properties probably mediated by its anti-inflammatory effects. The aim of this study was to investigate the effect of amniotic membrane transplantation (AMT) along with ciprofloxacin to cure the primary stages of Pseudomonas keratitis. METHODS: In total, 28 rabbits were selected and divided in four groups as follows: group 1 as control, group 2 with amniotic membrane, group 3 with ciprofloxacin, and group 4 with amniotic membrane combined with ciprofloxacin. About 0.05 cc suspension of Pseudomonas aeruginosa, 27853 ATCC was injected into corneal stroma. RESULTS: The results showed groups of AMT, AMT + ciprofloxacin, and ciprofloxacin had 0% perforation while the control group had 85.6%. Average infiltration of 5.5 mm was observed in ciprofloxacin group, 5 mm in AMT + ciprofloxacin group, 24 mm in AMT group, and finally 23.75 mm for control. Amniotic membrane showed to be effective in prevention of cornea perforation as well as remission of Pseudomonas keratitis. There was no significant difference between ciprofloxacin groups in comparison with ciprofloxacin + AMT group. However, regarding the anti-inflammatory effect, the process of improvement of inflammation in ciprofloxacin + AMT group was faster. CONCLUSION: Transplantation of amniotic membrane in the primary stages of Pseudomonas keratitis treatment remarkably prevents the disease and it can be used to control its process.

Use of a new multiplex quantitative polymerase chain reaction based assay for simultaneous detection of Neisseria meningitidis, Escherichia coli K1 , Streptococcus agalactiae, and Streptococcus pneumoniae.

BACKGROUND AND OBJECTIVES: Neisseria meningitidis, Escherichia coli K1, Streptococcus agalactiae, and Streptococcus pneumoniae cause 90% of bacterial meningitis. Almost all infected people die or have irreversible neurological complications. Therefore, it is essential to have a diagnostic kit with the ability to quickly detect these fatal infections. MATERIALS AND METHODS: The project involved 212 patients from whom cerebrospinal fluid samples were obtained. After total genome extraction and performing multiplex quantitative polymerase chain reaction (qPCR), the presence or absence of each infectious factor was determined by comparing with standard strains. RESULTS: The specificity, sensitivity, positive predictive value, and negative predictive value calculated were 100%, 92.9%, 50%, and 100%, respectively. So, due to the high specificity and sensitivity of the designed primers, they can be used instead of bacterial culture that takes at least 24 to 48 hours. CONCLUSION: The remarkable benefit of this method is associated with the speed (up to 3 hours) at which the procedure could be completed. It is also worth noting that this method can reduce the personnel unintentional errors which may occur in the laboratory. On the other hand, as this method simultaneously identifies four common factors that cause bacterial meningitis, it could be used as an auxiliary method diagnostic technique in laboratories particularly in cases of emergency medicine.

Phenotypic Identification and Genotypic Characterization of Plasmid-Mediated AmpC ß-Lactamase-Producing Escherichia coli and Klebsiella pneumoniae Isolates in Iran.

One of the mechanisms of Klebsiella pneumoniae and Escherichia coli resistance to ß-lactam antibiotics is the production of ß-lactamase enzymes. Among these are the AmpC ß-lactamases, which confer resistance to a class of antibiotics. However, little is known about the AmpC ß-lactamases of K. pneumoniae and E. coli clinical isolates in Qazvin, Iran. This study was designed to assess the AmpC ß­lactamases-producing strains and also identify the prevalence of AmpC ß­lactamases genes. Antimicrobial susceptibility tests were performed on 435 K. pneumoniae and E. coli isolates using disk diffusion technique. Plasmid-mediated AmpC genes were studied using a multiplex PCR assay. The AmpC ß-lactamase-producer isolates were studied by employing cefoxitin disk diffusion test, AmpC induction test, AmpC cefoxitin-EDTA test, and boronic acid disk test. Our results showed that of 46 (18.4%) cefoxitin-insensitive E. coli isolates, 10 (21.7%) were positive for AmpC ß-lactamase genes, among them 4 (8.69%) isolates were positive for blaDHA genes and 6 (13%) for blaCIT genes. Of 57 (30.4%) cefoxitin-insensitive K. pneumoniae isolates, 10 (17.5%) were positive for AmpC gene with 4 (6.34%) and 6 (9.5%) isolates positive for blaDHA and blaCIT genes, respectively. However, no MOX, ACC, FOX, or EBC genes were detected in the isolates. Considering the results of different confirmatory phenotypic tests, the AmpC cefoxitin-EDTA test showed a higher discriminatory power for detecting AmpC ß-lactamase-producing strains. The specificity and sensitivity of AmpC cefoxitin-EDTA were 77%, 100% for K. pneumonia and 70%, 90% for E. coli higher than the other two tests, respectively. Also, the authors demonstrated high prevalence rate for resistance to certain antibiotics, such as cefuroxime, trimethoprim-sulfamethoxazole, ampicillin, and cefotaxime. In conclusion, our study provided valuable information regarding the plasmid-mediated AmpC ß-lactamase gene content, antibiotic resistance, and confirmatory phenotypic tests for AmpC ß-lactamases in E. coli and K. pneumoniae isolates from clinical sources.