Influenza A viral loads in respiratory samples collected from patients infected with pandemic H1N1, seasonal H1N1 and H3N2 viruses.
BACKGROUND
Nasopharyngeal aspirate (NPA), nasal swab (NS), and throat swab (TS) are common specimens used for diagnosis of respiratory virus infections based on the detection of viral genomes, viral antigens and viral isolation. However, there is no documented data regarding the type of specimen that yields the best result of viral detection. In this study, quantitative real time RT-PCR specific for M gene was used to determine influenza A viral loads present in NS, NPA and TS samples collected from patients infected with the 2009 pandemic H1N1, seasonal H1N1 and H3N2 viruses. Various copy numbers of RNA transcripts derived from recombinant plasmids containing complete M gene insert of each virus strain were assayed by RT-PCR. A standard curve for viral RNA quantification was constructed by plotting each Ct value against the log quantity of each standard RNA copy number.


RESULTS
Copy numbers of M gene were obtained through the extrapolation of Ct values of the test samples against the corresponding standard curve. Among a total of 29 patients with severe influenza enrolled in this study (12 cases of the 2009 pandemic influenza, 5 cases of seasonal H1N1 and 12 cases of seasonal H3N2 virus), NPA was found to contain significantly highest amount of viral loads and followed in order by NS and TS specimen. Viral loads among patients infected with those viruses were comparable regarding type of specimen analyzed.


CONCLUSIONS
Based on M gene copy numbers, we conclude that NPA is the best specimen for detection of influenza A viruses, and followed in order by NS and TS.
Ngaosuwankul
Nathamon
N
Department of Microbiology, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, Thailand.
Noisumdaeng
Pirom
P
Komolsiri
Pisut
P
Pooruk
Phisanu
P
Chokephaibulkit
Kulkanya
K
Chotpitayasunondh
Tawee
T
Sangsajja
Chariya
C
Chuchottaworn
Charoen
C
Farrar
Jeremy
J
Puthavathana
Pilaipan
P
D000293:Adolescent; D000328:Adult; D000368:Aged; D002648:Child; D002675:Child, Preschool; D006801:Humans; D007223:Infant; D053118:Influenza A Virus, H1N1 Subtype / Q000302:isolation & purification*; D053122:Influenza A Virus, H3N2 Subtype / Q000302:isolation & purification*; D007251:Influenza, Human / Q000821:virology*; D008875:Middle Aged; D009297:Nasal Mucosa / Q000821:virology*; D009305:Nasopharynx / Q000821:virology*; D010614:Pharynx / Q000821:virology*; D012367:RNA, Viral / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D019562:Viral Load; D014763:Viral Matrix Proteins / Q000235:genetics; D055815:Young Adult
C031540:M-protein, influenza virus; D012367:RNA, Viral; D014763:Viral Matrix Proteins
Nature. 2009 Jun 18;459(7249):931-9
19525932
J Infect Dis. 2009 Aug 15;200(4):485-8
19591578
Lancet. 2004 Feb 21;363(9409):617-9
14987888
Nat Med. 2006 Oct;12(10):1203-7
16964257
J Infect Dis. 2009 Aug 15;200(4):492-500
19591575
Arch Virol. 2001 Dec;146(12):2275-89
11811679
Immunol Rev. 2008 Oct;225:68-84
18837776
Wkly Epidemiol Rec. 2009 May 29;84(22):197-202
19480086
J Infect Dev Ctries. 2009 Jun 01;3(5):331-4
19759501
Nature. 2009 Aug 20;460(7258):1021-5
19672242
J Med Virol. 2010 Jan;82(1):1-7
19950247
J Clin Virol. 2004 Mar;29(3):179-88
14962787