Evaluation of real‐time PCR assay for the detection of Ascaris suum contamination in meat and organ meats

Abstract

Ascarid larva migrans syndrome (ascarid LMS) is caused by ascarid roundworms including Ascaris suum. In East Asia, ascarid LMS has been considered as a food-borne disease in adulthood who has the dietary habitat of consuming raw or lightly cooked meat and organ meats. To evaluate the potential risk of A. suum infection from these foods, Ascaris specific real-time PCR was developed.The assay could constantly detect A. suum DNA up to 10 fg. Its specificity was confirmed by nonamplification with Toxocara canis and Toxocara cati DNA. A. suum DNA could be amplified from not only a single larva but also mouse liver spiked with a larva. Moreover, the assay could detect A. suum DNA in experimentally infected mouse liver, and showed higher sensitivity than a conventional digestion method. This real-time PCR assay would be useful for detecting the A. suum larval contamination in meat and organ meats. Practical applicationsIt has been considered that one of the most important risk factors for ascarid LMS in humans is the consumption of raw or undercooked meat or organ meats of domestic animals infected with Ascaris suum. Thus, we developed the novel real-time PCR with the high sensitivity and specificity in order to specifically detect A. suum DNA from animal tissues. This assay can be applied to the meat inspection procedure for the identification of parasite larval contamination that may adversely impact on public health as well as on animal health and welfare.

Comparison of real‐time PCR and cultural method for detection of bacterial load in pasteurized milk

“…In the aforementioned study, a poor agreement between the quantitative output of the real‐time PCR and the bacterial culture method was observed (Hein et al, ). Yen Thi reported that the development of sensitive and specific TaqMan real‐time PCR assay allowing rapid and reliable identification of A. suum larvae in meat and organs meat (Hoang et al, ).…”

Application of a real-time PCR assay for the detection of <i>Ascaris suum</i> DNA in the liver of experimentally infected chickens

“…A TaqMan probe with the sequence of 5'-FAM-CGTGAGCCACATAGTAAATTGCACACAAAATG-TAMRA-3' was designed in the sequence of the amplified product. The real-time PCR condition was described previously[9].Statistical analysis of the differences in the positive rate of A. suum DNA detection between 50 mg lesion and 500 mg of the remaining samples in each group was done using Fisher's exact test with a significance level of 5%.…”

“…Ascaris suum , an intestinal nematode, is known to be distributed worldwide. Besides parasitizing in pigs as a definitive host, this roundworm can infect other animals, such as chickens and cattle, and is considered a paratenic host [ 7 , 8 , 9 , 16 ]. In addition, this roundworm is a pathogenic agent causing ascarid larva migrans syndrome (ascarid LMS) in humans [ 5 , 6 , 12 , 13 , 14 ].…”

“…However, this method cannot always identify the presence of larval contamination in meat/offal because serological tests reflect not only present infections, but also past ones. In a previous study, we evaluated a real-time PCR assay with high sensitivity and specificity in order to directly identify the existence of A. suum in meat and internal organs [ 9 ]. This study contained a limitation due to the processing sample amount: the maximum weight of applicable tissue for DNA sample preparation was 0.5 g. To apply this method for detecting A. suum DNA in domestic animals, such as chickens and cattle, a large size of liver should be subsampled for testing.…”

“…Worms were washed several times in saline (0.85%) and the lower part of the uterus was then removed to collect the fertilized eggs. The procedure of fertilized egg embryonation was described in detail in a previous study [ 9 ]. Embryonated eggs were then used for the experimental infection of chickens.…”