Development of a polymerase chain reaction applicable to rapid and sensitive detection ofClonorchis sinensiseggs in human stool samples

Abstract

Microscopic examination of eggs of parasitic helminths in stool samples has been the most widely used classical diagnostic method for infections, but tiny and low numbers of eggs in stool samples often hamper diagnosis of helminthic infections with classical microscopic examination. Moreover, it is also difficult to differentiate parasite eggs by the classical method, if they have similar morphological characteristics. In this study, we developed a rapid and sensitive polymerase chain reaction (PCR)-based molecular diagnostic method for detection of Clonorchis sinensis eggs in stool samples. Nine primers were designed based on the long-terminal repeat (LTR) of C. sinensis retrotransposon1 (CsRn1) gene, and seven PCR primer sets were paired. Polymerase chain reaction with each primer pair produced specific amplicons for C. sinensis, but not for other trematodes including Metagonimus yokogawai and Paragonimus westermani. Particularly, three primer sets were able to detect 10 C. sinensis eggs and were applicable to amplify specific amplicons from DNA samples purified from stool of C. sinensis-infected patients. This PCR method could be useful for diagnosis of C. sinensis infections in human stool samples with a high level of specificity and sensitivity.

Accuracy of the Kato-Katz method and formalin-ether concentration technique for the diagnosis of Clonorchis sinensis, and implication for assessing drug efficacy

“…Although collection of more than one stool sample causes logistic inconvenience and might lower patient compliance, we recommend that at least two stool samples should be obtained in order to achieve an accurate diagnosis at treatment follow-up. FECT cannot be recommended for the diagnosis of clonorchiasis unless it is combined with other techniques, such as polymerase chain reaction (PCR), a highly sensitive approach, which is gaining importance for helminth diagnosis [ 42 - 44 ], but might not be available in resource-constrained settings.…”

Serodiagnostic antigens of Clonorchis sinensis identified and evaluated by high-throughput proteogenomics

“…Diagnosing C. sinensis infection classically involves microscopic detection of the parasite eggs from stool samples. To detect specific target genes [30,31], the PCR methods are sensitive but complicated in procedures, hindering field applications in the endemic areas. Serodiagnostic methods are useful for screening patients infected with C. sinensis and used to complement the stool microscopy.…”

A co-infection case report of Taenia saginata in a patient with subclinical clonorchiasis confirmed by the combination of diagnostic tools

“…Human taeniasis is usually diagnosed by observing ova or gravid proglottids in the patient’s stool. From diagnoses conducted after 1993, the Taenia tapeworms infecting humans in Korea were identified as T. solium, T. saginata , and T. asiatica [5, 17, 18]. However, additional differential modalities may be required to clearly distinguish among these species, as morphological characteristics, such as the presence of an unarmed rostellum on the scolex of adult, the large number of uterine twigs, and the presence of a posterior protuberance, can be difficult to observe in individual strobili [17, 18].…”

“…Of food-borne parasites, Clonorchis sinensis showed the highest prevalence in Korea showing 2.4% of egg-positive rate in general population [3]. The parasite is more prevalent in populations living by rivers, with an egg-positive rate of 11.4% [4], as humans usually become infected after ingesting raw or undercooked freshwater fish harboring C. sinensis metacercariae [5]. Human taeniasis, another food-borne parasitic infection, is a zoonotic disease, because it involves pigs ( Taenia solium and Taenia asiatica ) or cattle ( Taenia saginata ) as an intermediate host and humans as the definitive host [6, 7].…”