Establishment of rats model of Pneumocystic carinii infection and study on the sensitivity of ITS-nested PCR assay

Abstract

Objective To establish the rats model of Pneumocystis carinii infection and study on the sensitivity of ITS1-5.8S rDNA-ITS2 nested PCR. Methods SD rats were divided as experiment group and control group at random. Each experiment rat was rendered immunedepressant by subcutaneous injection of dex-amethasone from the 1 st week with 1 mg per rat twice a week for 8 weeks, and the control rats with normal feeding. P. carinii of experiment group was examined microscopically in bronchoalveolar lavage (BAL) on lung impression smears and lung tissue smears stained with Gomorismethenamine silver (GMS) at the 2nd, 4th, 6th and 8th week after injection, respectively. Those of control group was examined at the beginning of experi-ment and the 10th week. The DNAs of P. carinii were extracted beth from the BAL and lung tissues of rats. ITS1-5.8S rDNA -ITS2 of P. carinii were amplified by nested PCR. The sensitivity was compared between GMS and nested PCR. Results P. carinii could be observed from the 6th week in the experiment group and at the 8th week with 100% (10/10) positive in lung impression smears and lung tissue smears, 80% (8/10) in BAL smear. The control group was negative. Nested PCR detected the DNAs of P. carinii both from BAL and lung tissue at the 8th week with 100% positive, and all negative in control group. Comparing the detect efficiency of P. carinii among BAL, lung impression smears and lung tissue smears by GMS method, BAL was the lowest, lung tissue smears was the highest. 20% specimens of BAL couldnt be detected by GMS method, while nested PCR worked. Conclusions The rats model of P. carinii infection was successfully established by injection ofdexamethasone. ITS1-5.8S rDNA-ITS2 nested PCR is a method with high sensitivity and specificity, whichcould be potentially used to detect Pneumocystis pneumonia in clinic. Key words: Pncumocystis; Rats model; Internal transcribed spacer (ITS) ; Nested PCR; Sensitivity; Specificity