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