This study carried out to detect a adhesion genes (icaA and ica D) among orthopedic patients showing prosthetic joint infection in Helmia Armed Forces Hospital.
The hip and knee aspirate samples (49) were collected from (patients showing infected prosthesis). Understand conditions and transfer rapidly to the lab.
Notes: (5) sample were negative control means total (54) samples.
The aim of this study to detect biofilm formation and also detect some genes (icaA, icaD) which are responsible for biofilm formation. and gene expression
The material and methods
The samples were cultured on blood Agar medium then the isolates were cultured on Conog Red. Agar medium to detect biofilm formation.
Molecular detection carried out: firstly DNA was extracted from bacterial isolates, RNA was also extracted and was used as template for real time PCR (qRCR) by light cycler (Roch) PCR, syber green as fluorescent dye.
Reverse transcriptase was done, specific primers were used for icaA and icaD to transfere RNA into cDNA genes 16rSRNA was used as reference control.
The results
The results showed that 67% of the samples (hip and knee aspirates) were biofilm formation (black colonies +ve biofilm) red colonies (-ve) biofilm formation. The results showed that in hip aspirate samples) (4) out of (28) 14.8% were positive for both (icaA, icaD) and positive for biofilm formation. In knee aspirate sample (5) out of (21) 28.5% were positive for both (icaA and icaD) and positive for biofilm formation, in current study.
The phenotype (biofilm formation) don't correlate with the presence of icaA, icaD.
The gene expression was evaluated with the presence of mRNA (icaA, icaD) for example, in current study (hip aspirate samples) (4) out of (28) 14.2% and negative for biofilm formation (phenotype).
In knee isolates samples (3) out of (21) were negative for icaA and icaD but positive phenotype.
Conclusion: biofilm bacteria is difficult to eradicate and it causes catastrophic problem for prosthetic infected orthopedic patients.
There are some factors lead to biofilm formation genes basically and the environment, the surface of biomaterial. The presence, the concentration of some substances like: glucose, iron, phenols, limited oxygen. The genes expression of icaA, icaD is proportional with amount of mRNA of these genes,
Key words: Intercellular adhesion genes, icaA and icaD, prosthetic joint in infection, Total hip and total knee arthroplasty (THA and TKA), real time PCR, biofilm formation.
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