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Comparison of virus isolation using the Vero E6 cell line with real-time RT-PCR assay for the detection of human metapneumovirus.

Matsuzaki Y, Mizuta K, Takashita E, Okamoto M, Itagaki T, Katsushima F, Katsushima Y, Nagai Y, Nishimura H - BMC Infect. Dis. (2010)

Bottom Line: Between December 2007 and July 2008, we obtained 224 nasopharyngeal swab specimens from patients with acute respiratory infection and tested by the two methods.Among specimens collected within 3 days after symptom onset, all of the real-time RT-PCR positive specimens having a viral load of more than 1.25x105 copies/ml were found positive by cell culture.Thus, this method is a useful method for epidemiological and virological research even in facilities with minimal laboratory resources.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Microbiology, Yamagata Prefectural Institute of Public Health, Tokamachi 1-6-6, Yamagata 990-0031, Japan.

ABSTRACT

Background: The use of cell culture for the diagnosis of human metapneumovirus (hMPV) infection is uncommon at present and molecular method such as reverse-transcription PCR (RT-PCR) has been widely and most commonly used as the preferred test. We aimed to compare the results of virus isolation using Vero E6 cells with real-time RT-PCR for the detection of hMPV, since such a comparison data is not available.

Methods: Between December 2007 and July 2008, we obtained 224 nasopharyngeal swab specimens from patients with acute respiratory infection and tested by the two methods.

Results: Forty-three (19.2%) were found positive by cell culture and 62 (27.7%) by real-time RT-PCR. Cell cultures were positive for 42 of 62 specimens found positive by real-time RT-PCR (67.7% sensitivity) and for 1 of 162 specimens found negative by real-time RT-PCR (99.4% specificity), respectively. The sensitivity of the cell culture was 76.2-87.5% (mean 81.8%) when specimens were collected within 3 days after the onset of symptoms, and the sensitivity decreased to 50% or less thereafter. Among specimens collected within 3 days after symptom onset, all of the real-time RT-PCR positive specimens having a viral load of more than 1.25x105 copies/ml were found positive by cell culture.

Conclusions: Cell culture using Vero E6 cell line has 81.8% sensitivity compared with the real-time RT-PCR method, when specimens are collected within 3 days after the onset of symptoms. Thus, this method is a useful method for epidemiological and virological research even in facilities with minimal laboratory resources.

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Related in: MedlinePlus

Viral loads of real-time RT-PCR-positive specimens and the number of days from onset of fever to specimen collection. Solid triangles indicate the 62 specimens that tested positive for hMPV by real-time RT-PCR assay. Specimens testing negative by cell culture are marked with daggers.
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Figure 1: Viral loads of real-time RT-PCR-positive specimens and the number of days from onset of fever to specimen collection. Solid triangles indicate the 62 specimens that tested positive for hMPV by real-time RT-PCR assay. Specimens testing negative by cell culture are marked with daggers.

Mentions: Viral loads of the 62 specimens tested positive by real-time RT-PCR were distributed between 1.99 × 102 and 3.85 ×106. The results of the real-time RT-PCR are shown in Figure 1. All of the real-time RT-PCR-positive specimens having more than 1.25×105 hMPV copies/ml were also found positive by cell culture, though two specimens with 8.79×105 and 1.36×106 copies/ml that were collected on Day 4 and 5 after the onset of symptoms, respectively, were negative. The sensitivity of virus isolation for specimens obtained at 1-3 days after the onset of symptoms was 76.2-87.5% (mean 81.8%), whereas it was less than 25-50% (mean 33.3%) for 4-7 days (Table 1). The sensitivity of virus isolation for specimens obtained within 3 days after the onset of symptoms was statistically higher than that for specimens obtained more than 3 days after the onset of symptoms (P = .0006).


Comparison of virus isolation using the Vero E6 cell line with real-time RT-PCR assay for the detection of human metapneumovirus.

Matsuzaki Y, Mizuta K, Takashita E, Okamoto M, Itagaki T, Katsushima F, Katsushima Y, Nagai Y, Nishimura H - BMC Infect. Dis. (2010)

Viral loads of real-time RT-PCR-positive specimens and the number of days from onset of fever to specimen collection. Solid triangles indicate the 62 specimens that tested positive for hMPV by real-time RT-PCR assay. Specimens testing negative by cell culture are marked with daggers.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2902479&req=5

Figure 1: Viral loads of real-time RT-PCR-positive specimens and the number of days from onset of fever to specimen collection. Solid triangles indicate the 62 specimens that tested positive for hMPV by real-time RT-PCR assay. Specimens testing negative by cell culture are marked with daggers.
Mentions: Viral loads of the 62 specimens tested positive by real-time RT-PCR were distributed between 1.99 × 102 and 3.85 ×106. The results of the real-time RT-PCR are shown in Figure 1. All of the real-time RT-PCR-positive specimens having more than 1.25×105 hMPV copies/ml were also found positive by cell culture, though two specimens with 8.79×105 and 1.36×106 copies/ml that were collected on Day 4 and 5 after the onset of symptoms, respectively, were negative. The sensitivity of virus isolation for specimens obtained at 1-3 days after the onset of symptoms was 76.2-87.5% (mean 81.8%), whereas it was less than 25-50% (mean 33.3%) for 4-7 days (Table 1). The sensitivity of virus isolation for specimens obtained within 3 days after the onset of symptoms was statistically higher than that for specimens obtained more than 3 days after the onset of symptoms (P = .0006).

Bottom Line: Between December 2007 and July 2008, we obtained 224 nasopharyngeal swab specimens from patients with acute respiratory infection and tested by the two methods.Among specimens collected within 3 days after symptom onset, all of the real-time RT-PCR positive specimens having a viral load of more than 1.25x105 copies/ml were found positive by cell culture.Thus, this method is a useful method for epidemiological and virological research even in facilities with minimal laboratory resources.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Microbiology, Yamagata Prefectural Institute of Public Health, Tokamachi 1-6-6, Yamagata 990-0031, Japan.

ABSTRACT

Background: The use of cell culture for the diagnosis of human metapneumovirus (hMPV) infection is uncommon at present and molecular method such as reverse-transcription PCR (RT-PCR) has been widely and most commonly used as the preferred test. We aimed to compare the results of virus isolation using Vero E6 cells with real-time RT-PCR for the detection of hMPV, since such a comparison data is not available.

Methods: Between December 2007 and July 2008, we obtained 224 nasopharyngeal swab specimens from patients with acute respiratory infection and tested by the two methods.

Results: Forty-three (19.2%) were found positive by cell culture and 62 (27.7%) by real-time RT-PCR. Cell cultures were positive for 42 of 62 specimens found positive by real-time RT-PCR (67.7% sensitivity) and for 1 of 162 specimens found negative by real-time RT-PCR (99.4% specificity), respectively. The sensitivity of the cell culture was 76.2-87.5% (mean 81.8%) when specimens were collected within 3 days after the onset of symptoms, and the sensitivity decreased to 50% or less thereafter. Among specimens collected within 3 days after symptom onset, all of the real-time RT-PCR positive specimens having a viral load of more than 1.25x105 copies/ml were found positive by cell culture.

Conclusions: Cell culture using Vero E6 cell line has 81.8% sensitivity compared with the real-time RT-PCR method, when specimens are collected within 3 days after the onset of symptoms. Thus, this method is a useful method for epidemiological and virological research even in facilities with minimal laboratory resources.

Show MeSH
Related in: MedlinePlus