Molecular study of some virulence genes of Pseudomonas aeruginosa isolated from different infections in hospitals of Baghdad
Authors: Ali Ali Muhsin , Al-Kenanei Khaled Aziz , Hussein Sana’a Noori , Bdaiwi Qasim Obaid 
Published: October 2019

One hundred isolates ofPseudomonas aeruginosawere obtained from patients admitted to Baghdad hospitals, Iraq during the period between May 2018 until July 2018. These isolates were distributed as 15 isolates from blood, 25 isolates from urinary tract infections, 10 isolates from sputum, 12 isolates from wounds, 15 isolates from ear infections, 15 isolates from bronchial wash of patients suffering from respiratory tract infections in addition to 8 isolates from cystic fibrosis patients. The isolates were initially identified by culturing on MacConkey agar, blood agar andP. aeruginosaagar then diagnosed by performing some morphological and biochemical tests. The second diagnosis was done by API 20E system followed by Vitek 2 compact system. Antibiotic sensitivity test was carried out towards 15 antibiotics using disc diffusion method (Kirby–Bauer method). The results of sensitivity test showed thatP. aeruginosaisolates possessed high resistance towards most antibiotics under study, the most antibiotic resistance was towards Gentamicin 87 (87%), whereas the lowest resistance was towards Imipenem 10 (10%). In this study, two types of methods were used in the detection of biofilm formation: the first one was Congo red agar method and the second one was microtiter plate method. In the first method, results showed that biofilm formed by 57/100 (57%) according to black color production on media, whereas in the second method was 69/100 (69%) produce strong adherence according to OD in ELISA reader. Genotypic detection of many virulence factors related toP. aeruginosawas performed using conventional PCR. These included: gene coded for exoenzyme S (exoS), exoenzyme U (exoU), exotoxin A (toxA), two phospholipases C encoded by (plcH) and (plcN), alginate (algD), (lasB), rpsl,proteaseIV, and Neuraminidase (nan1). The results revealed that the most frequent gene wasexoSas it was detected in 87/100 (87%) isolates, whereas the least frequent gene wasnan1as it was detected in only 9/100 (9%). The frequency of detection of other genes were as follows:toxAiin 55/100 (55%);plcHin 45/100 (45%);exoUin 42/100 (42%);plcNin 33/100 (33%);proteaseIVin 31/100 (31%),algDin 29/100 (29%);lasBin 28/100 (28%), andrpslin 25/100 (25%). Phylogenetic analysis by Enterobacterial Repetitive Intergenic Consensus (ERIC-PCR), ERIC-DNA Fingerprinting revealed the diversity of all isolates in Baghdad by using Dice coefficient and the unweighted pair group method with arthmetic average (group method) of phylogenetic analysis. The percentage level of similarity clearly showed that the isolates examined by species were divided into two distinct cluster numbers, in addition to three single isolates (clone), that clustered at a similarity level of (93%). According to the statistical analysis, it was found that the correlation coefficient of ERIC genotyping method with virulence genes in this study and antibiotics sensitivity test was significant atP < 0.05 (two-tailed), whereas correlation with biofilm was not significant.