Molecular identification of Pseudomonas aeruginosa recovered from cystic fibrosis patients.

OBJECTIVE: Precise identification of various morphotypes of Pseduomonas aeruginosa which developed during cystic fibrosis (CF) is of prime importance. We aimed to identify the isolates of P. aeruginosa recovered from CF patients at the genus and species level through primers targeting oprI and oprL genes via PCR. METHODS: Sputum samples or throat swabs were taken from 100 CF patients and plated on cetrimide agar. All suspected colonies were primarily screened for P. aeruginosa by a combination of phenotypic tests. Molecular identification of colonies was performed using specific primers for oprI and oprL genes. RESULTS: Based on phenotypic tests, P. aeruginosa isolates were recovered from 40% of CF patients. Forty isolates yielded amplicon of oprI gene using genus-specific primers confirming the identity of fluorescent pseudomonads. However, 37 of 40 isolates yielded amplicon of oprL gene using species-specific primers, verifying the identity of P. aeruginosa. CONCLUSION: This study showed that the species-specific PCR targeting oprL gene can be used as accurate test for identification of highly adaptable P. aeruginosa in CF patients. This procedure may provide a simple and reliable method for identification of various morphotypes.

Identification and molecular epidemiology of nosocomial outbreaks due to Burkholderia cepacia in cystic fibrosis patients of Masih Daneshvary Hospital, Iran.

INTRODUCTION: B. cepacia complex have emerged as an important opportunistic pathogen in hospitalized and immunocompromised patients. Small hospital outbreaks are frequent and are usually due to a single contaminated environmental source. In this study we were going to investigate the role of B.cepacia complex in those patients suspected to involve with cystic fibrosis and evaluate responsible types in Masih Daneshvary Hospital. METHODS: One hundred specimens were collected from all admitted patients who were suspected to cystic fibrosis to Masih Daneshvary hospital during one year April 2011 till end of March 2012. All were culture and identified standard procedure. All samples were checked by API system (API20NE) and by specific PCR method for genus Bulkhorderia and Bcc as well. Identified strains were finally tested by PFGE system to identifying specific involving pulse-types. RESULT: . Isolation and identification methods revealed 5 specimens were B.cepasia, The frequency of the cystic fibrosis detected at this study was lower than other similar study previously reported. All these isolates showed similar pattern by PFGE standard protocol that may have spread from a single source and could not be attributed to cross infections from patient to patients. DISCUSSION: Application of PFGE and identification of pulse-type is a potential tool to enhance the investigation of apparent nosocomial outbreaks of B.cepacia. However it needs to be adjusted with environmental findings. Implementation of educational programs and adherence to infection control policies are obviously the main element for complete elimination of an outbreak.

Clonal groups of high-level gentamicin-resistant Enterococcus faecium isolated from municipal wastewater and clinical samples in Tehran, Iran.

AIMS: Clonality among high-level gentamicin-resistant Enterococcus faecium (HLGR-EF) isolates obtained from clinical and sewage treatment plants (STP) were investigated using PhePlate system (PhP), ribotyping and pulsed-field gel electrophoresis (PFGE). METHODS AND RESULTS: During 1 year study (September 2005-2006), a total of 106 HLGR-EF isolates were collected from clinical (n = 48) and STP (n = 58) samples in Tehran, Iran. Biochemical fingerprinting of these isolates using the PhP showed the presence of 21 PhP types (diversity index, Di = 0.97) among the clinical and 21 PhP types (Di = 0.91) among the STP isolates. Representative isolates of each PhP type (n = 42) were further characterized by the ribotyping method. Sixteen ribotypes were identified among the isolates with five types shared between the clinical and STP isolates. PFGE recognized 24 clonal types among these isolates with three pulsotypes shared between the clinical and STP isolates. Combination of the two techniques (PFGE and ribotyping) resulted in 24 (Di = 0.96) and 16 (Di = 0.93) types among the strains isolated from clinical and STP samples, respectively. CONCLUSIONS: We concluded that the combination of PhP typing, ribotyping and PFGE could be extremely discriminatory when examining HLGR-EF isolates. SIGNIFICANCE AND IMPACT OF THE STUDY: The emergence of highly diverse HLGR-EF population in Iran is of serious concern especially because of their multi-resistances.