Browsing by Subject "qPCR"

Parvoviruses are among the smallest known viruses with a genome of ~ 5 kilobases. To date, six parvoviruses have been identified in human samples, with parvovirus B19 and human bocavirus being the only two known human pathogens in this family. Bufavirus, tusavirus and cutavirus are the most recently discovered parvoviruses, all belonging to the Protoparvovirus genus. Bufavirus was predominately discovered in fecal samples of children with diarrhea in Africa, Asia, and Europe. Cutavirus was detected in fecal samples of children with diarrhea in South America and Africa but has also been found in skin biopsies of patients with cutaneous T cell carcinoma and malignant melanoma. Tusavirus was discovered in a stool specimen of one child with diarrhea in Tunisia. At the moment, there are too little data to determine the identity of tusavirus as a human virus. More data and evidence are required to assess the association of the three parvoviruses with human diseases. In this study, a multiplex real-time PCR method was established to facilitate the detection and quantification of the three human-associated protoparvoviruses for further epidemiology and pathobiology study. Differentiation of different viruses was achieved by using three uniquely labeled probes. The multiplex assay was able to detect ≤ 10 copies/μL of bufavirus, tusavirus, and cutavirus plasmid templates simultaneously, with an average efficiency from 100.54% to 103.76%. The assay was applied to assess the prevalence of the three viruses in skin tissues of 93 non-immunosuppressed individuals with contact dermatitis and 137 immunosuppressed transplant recipients. Bufavirus and tusavirus DNA was detected in neither of the cohorts, which might indicate the rarity of the two viruses in skin tissues. Cutavirus DNA was detected in four (2.92%) transplant recipients, but all samples from the non-immunosuppressed group were negative. Among the four cutavirus positive patients, one was diagnosed with squamous cell carcinoma. These findings further support previous discoveries of cutavirus DNA in skin tissues and serve as evidence for the identity of cutavirus as a human virus. However, its association with cancer remains to be further investigated.