title: Field evaluation of an improved cell line for the detection of human adenoviruses in environmental samples.
abstract: Human enteric adenoviruses (HAdVs) are commonly detected in waters contaminated with human fecal material and persistent in the environment. Detecting infectious enteric HAdVs is limited by the difficulty of growing them in cell cultures. Recently, an improved cell line (293 CMV) has been described, which enhanced the propagation of enteric HAdVs (Kim et al., 2010. Appl. Environ. Microbiol. 76, 2509-2516). The present study evaluated the transactivated 293 CMV cell line for detecting enteric HAdVs from field samples, which is an important step in demonstrating the usefulness of the improved cell line for water monitoring programs. Field samples consisted of the following: concentrated sewage samples (from 1L) collected from three different wastewater treatment plants (WWTPs) and concentrated raw source water samples (from 20L) collected from six water treatment plants (WTPs). Infectious HAdVs were detected using a combined cell culture/mRNA RT-PCR assay. Concentrated samples were assayed, in parallel, using the standard (STD) G293 and 293 CMV cell lines. Viral replication was determined by measuring viral mRNA and viral DNA levels during infection. Infectious HAdVs were successfully detected from environmental samples using the new transactivated and standard cell lines. Infectivity assays of concentrated sewage samples demonstrated higher viral mRNA expression (p=0.02) and viral DNA concentrations (p=0.02) in the transactivated 293 CMV than in the G293 cell line. Although not statistically significant, infectious HAdVs were detected in more raw water samples using the 293 CMV cells (8 of 18) than in the STD G293 cells (4 of 18). However, when results of the source water samples were pooled, the number of flasks positive using the 293 CMV cells was significantly greater than those using the G293 cells (p=0.01). Overall, the results of the present study demonstrate the effectiveness of the new transactivated 293 CMV cell line for improved propagation and detection of HAdVs from environmental samples.
authors.affiliation: University of North Carolina at Chapel Hill, Department of Environmental Sciences and Engineering Gillings School of Global Public Health, Chapel Hill, NC, USA.
authors.forename: Patsy M
authors.initials: PM
authors.lastname: Polston
authors.affiliation: University of North Carolina at Chapel Hill, Department of Environmental Sciences and Engineering Gillings School of Global Public Health, Chapel Hill, NC, USA. Electronic address: roberto.a.rodriguez@uth.tmc.edu.
authors.forename: Roberto A
authors.initials: RA
authors.lastname: Rodríguez
authors.affiliation: Seoul National University, Center for Human and Environmental Microbiome, Graduate School of Public Health, Seoul, South Korea.
authors.forename: KyungJin
authors.initials: K
authors.lastname: Seo
authors.affiliation: Seoul National University, Center for Human and Environmental Microbiome, Graduate School of Public Health, Seoul, South Korea.
authors.forename: Misoon
authors.initials: M
authors.lastname: Kim
authors.affiliation: Seoul National University, Center for Human and Environmental Microbiome, Graduate School of Public Health, Seoul, South Korea; Seoul National University, Bio-MAX Institute, Seoul, South Korea.
authors.forename: GwangPyo
authors.initials: G
authors.lastname: Ko
authors.affiliation: University of North Carolina at Chapel Hill, Department of Environmental Sciences and Engineering Gillings School of Global Public Health, Chapel Hill, NC, USA.
authors.forename: Mark D
authors.initials: MD
authors.lastname: Sobsey
mesh_terms: D000258:Adenovirus Infections, Human / Q000821:virology*; D000260:Adenoviruses, Human / Q000235:genetics / Q000302:isolation & purification* / Q000502:physiology; D018929:Cell Culture Techniques; D002460:Cell Line; D004279:DNA, Viral / Q000032:analysis; D005240:Feasibility Studies; D005243:Feces / Q000821:virology; D006801:Humans; D012333:RNA, Messenger / Q000032:analysis; D060888:Real-Time Polymerase Chain Reaction; D012722:Sewage / Q000821:virology*; D014779:Virus Replication; D062065:Wastewater / Q000821:virology; D014871:Water Microbiology
chemical_list: D004279:DNA, Viral; D012333:RNA, Messenger; D012722:Sewage; D062065:Wastewater
keywords: CC/mRNA; CMV 293; Detection; Enteric adenovirus; Real-time PCR; Transactivated cell line