Background: Trichinellosis is caused by Trichinella spiralis muscle larvae (ML), which in swine is the main source of transmission. Trichinellosis is detected by enzyme-linked immunosorbent assay (ELISA) using excretory–secretory antigens (ESAg). Preparation of ESAg is difficult, and it produces a low content. Furthermore, the sensitivity and specificity of the test depends on the quality of the antigen that is produced in each batch. Crude somatic antigens (CSAg) of T. spiralis ML at molecular weights (MWs) of 101, 79, and 43 kDa were previously positive for swine trichinellosis sera, with no cross-reaction observed for normal sera, except that 79 and 43 kDa each reacted with one of the coccidiosis and with mixed infections containing trichuriasis and coccidiosis. Therefore, the current study aimed to obtain eluted antigens of T. spiralis ML CSAg, which were analyzed by IgG-ELISA for detecting swine trichinellosis and identification by two-dimensional polyacrylamide gel electrophoresis (2-DE) and liquid chromatography-tandem mass spectrometry (LC MS-MS).Methods: In this study, T. spiralis larvae CSAg at 101, 79, and 43 kDa (TsCSAg-101, TsCSAg-79, and TsCSAg-43, respectively) were eluted from sodium dodecyl sulfate–polyacrylamide gel electrophoresis. The eluted antigens were analyzed by IgG-ELISA for sensitivities and specificities. In addition, three specific antigens were identified by 2-DE and LC MS-MS.Results: Sensitivity of IgG-ELISA using three eluted antigens was persistent at 100%. Specificities were 90.63%, 95.54%, and 97.77% following TsCSAg-101, TsCSAg-79, and TsCSAg-43, respectively. The LC MS-MS results showed that 18/20 spots of the antigens were identified for 11 different proteins. One protein has several isoforms; for example, a serine proteinase and the phosphoenolpyruvate carboxykinase protein.Conclusions: TsCSAg-43 showed the highest specificity compared to the other two eluted antigens, which indicated that these specific proteins (a 45k antigen–trichina [fragment], a DNA topoisomerase 2-alpha, an antigen targeted by protective antibodies, and a conserved hypothetical protein [gi339234223]) should be developed and produced in large volumes in further study.

Background: Angiostrongylus cantonensis is a well-known pathogen causing human angiostrongyliasis eosinophilic meningitis. Humans, as accidental hosts, are infected by eating undercooked snails containing third-stage larvae. A. malaysiensis is closely related to A. cantonensis and has been described as a potential human pathogen. Recently, the two species have been reported to have overlapping distributions in the same endemic area, particularly in the Indochina region. Because of their similar morphological characteristics, misidentification often occurs, particularly of the third-stage larva in the snail intermediate host. Methods: We designed species-specific primers to mitochondrial cytochrome b, which was used as a genetic marker. SYBR-green quantitative real-time PCR (qPCR) was employed to quantitatively detect and identify the third-stage larvae and tissue debris in the cerebrospinal fluid (CSF) of a patient, and to quantify third-stage larvae in the snail Achatina fulica collected from the field.Results: The newly designed primers were highly specific and sensitive, even when using conventional PCR. SYBR green qPCR quantitatively detected around 10−4 ng of genomic DNA from one larva and facilitated the specific detection and identification of parasitic genetic material from the CSF of a patient with angiostrongyliasis. The method also estimated the number of larvae in A. fulica and revealed that the primary source of Angiostrongylus infection in the King Rama IX public park study area was A. malaysiensis; although, the two Angiostrongylus species each infected 10% of the snails. Conclusions: Our SYBR green qPCR method is a useful and inexpensive technique for parasite identification and has sufficient sensitivity and specificity to detect a single larva and simultaneously discriminate between A. cantonensis and A. malaysiensis. The number of larvae infecting or co-infecting the snail intermediate host can also be estimated. In future research, this qPCR method could be employed in a molecular survey of A. cantonensis and A. malaysiensis occurrence within intermediate and definitive hosts. The technique should also be applied in a study analyzing CSF specimens from patients with eosinophilic meningitis to assess the usefulness of the method for clinical diagnosis.