Polymerase chain reaction (PCR) and sequencing are useful for species identification of Trichinella spp., especially when their morphological characteristics useful for identifying taxa are lacking. In the present study, nine Trichinella isolates from different provinces in mainland China were identified by the PCR-based method using the 5S ribosomal DNA intergene spacer region (5S ISR) and the mitochondrial large subunit ribosomal DNA genes as molecular markers. The results indicated that eight isolates originating from domestic pigs and one isolate originating from civet cat (Paguma larvata) showed identical DNA banding pattern to Trichinella spiralis. Sequence analysis of the 5S ISR gene further confirmed that the nine Trichinella isolates were T. spiralis and revealed the intraspecies genetic variation within T. spiralis.