Molecular Phylogeny Of Pneumocystis Based On Mt LSU RRNA And ITS1-5.8SrRNA-ITS2 Gene Sequence

Objective: To study the phylogeny of Pneumocystis from human and mammalianhosts based on 2 molecular labels, mt LSU rRNA and ITS1-5.8SrRNA-ITS2 genesequences. Methods: Experimental animals were immunosuppressed by a quantitativesubcutaneous injection of dexamethasone according to the animal's body weight.Bronchalveolar lavage fluid (BALF) and lung tissue from different animals and sputumspecimens from clinical suspicious PCP patients were collected. All collected specimenswere stained by Wright-Giemsa's compound stain to determine the severity of infection.A DNA extraction protocol was followed. Primers amplifying mt LSU rRNA andITSl-5.8SrRNA-ITS2 genes were designed and synthesized. The specificity andsensitivity of the primers were detected. The PCR products or positive clones weresequenced to obtain the target genes data. Softwares were used to analyse and comparethe identity and genetic distance of Pneumocystis from different hosts. The sequenceswere aligned with related sequences in GenBank and used together to constructphylogenetic tree. Resultes: Induced by dexamethasone the PCP animal model wasestablished. Two target genes were specifically amplified by PCR. The mt LSU rRNAgene could be sensitively amplified from even 0.6pg DNA. Five sequences of mt LSUrRNA genes of Pneumocystis from 4 hosts were obtained, among them those fromMeriones unguiculatus and human were new ones. Four sequences of ITS 1-5.8SrRNA-ITS2 genes of Pneumocystis from 4 hosts were obtained. Of them those from Merionesunguiculatus and New Zealand White Rabbit were new ones. By analyzing the identityand genetic distance of these gene sequences, it was demonstrated that the Pneumocystisspecies isolated from different hosts possess genetic diversity and host specificity.Phylogenetic trees showed the possible coevolution between Pneumocystis and their...