title: Rapid diagnostic method for detection of mumps virus genome by loop-mediated isothermal amplification.
abstract: Most mumps patients are clinically diagnosed without any virological examinations, but some diagnosed cases of mumps may be caused by other pathogens or secondary vaccine failure (SVF). To clarify these issues, a sensitive, specific, and rapid diagnostic method is required. We obtained 60 salivary swabs from 34 patients with natural infection during the course of the illness, 10 samples from patients with vaccine-associated parotitis, and 5 samples from patients with SVF. Total RNA was extracted and subjected to reverse transcription-PCR (RT-PCR) and loop-mediated isothermal amplification (LAMP) for genome amplification. We detected mumps virus RNA corresponding to 0.1 PFU by LAMP within 60 min after RNA extraction, with the same sensitivity as RT-nested PCR. Mumps virus was isolated in 30 of 33 samples within day 2, and mumps virus genome was amplified by LAMP in 32 of them. The quantity of virus titer was calculated by monitoring the time to reach the threshold of turbidity. The viral load decreased after day 3 and was lower in patients serologically diagnosed as having SVF with milder illness. Accuracy of LAMP for the detection of mumps virus genome was confirmed; furthermore, it is of benefit for calculating the viral load, which reflects disease pathogenesis.
authors.affiliation: Okafuji Pediatric Clinic, Himeji, Hyougo, Japan.
authors.forename: Takao
authors.initials: T
authors.lastname: Okafuji
authors.forename: Naoko
authors.initials: N
authors.lastname: Yoshida
authors.forename: Motoko
authors.initials: M
authors.lastname: Fujino
authors.forename: Yoshie
authors.initials: Y
authors.lastname: Motegi
authors.forename: Toshiaki
authors.initials: T
authors.lastname: Ihara
authors.forename: Yoshinori
authors.initials: Y
authors.lastname: Ota
authors.forename: Tsugunori
authors.initials: T
authors.lastname: Notomi
authors.forename: Tetsuo
authors.initials: T
authors.lastname: Nakayama
mesh_terms: D000818:Animals; D002648:Child; D002522:Chlorocebus aethiops; D016679:Genome, Viral; D006801:Humans; D009107:Mumps / Q000175:diagnosis* / Q000503:physiopathology / Q000821:virology; D009108:Mumps Vaccine / Q000009:adverse effects; D009109:Mumps virus / Q000235:genetics / Q000302:isolation & purification*; D021141:Nucleic Acid Amplification Techniques / Q000379:methods*; D010309:Parotitis / Q000175:diagnosis / Q000821:virology; D012367:RNA, Viral / Q000032:analysis; D012463:Saliva / Q000821:virology; D013048:Specimen Handling / Q000379:methods; D013997:Time Factors; D014709:Vero Cells; D019562:Viral Load
chemical_list: D009108:Mumps Vaccine; D012367:RNA, Viral