流感病毒核酸测定分析前变量和生物危害暴露风险评估
|
摘要
背景：
基于PCR的核酸测定技术已成为流感病毒学监测和诊断的首选方法。然而除了核酸测定方法、测定试剂、使用的引物和探针外,核酸测定分析前的样品储存、转运以及核酸分离纯化方法等变量也对核酸测定结果产生重要影响。此外,核酸测定过程中潜在的生物危害也是核酸测定技术实际应用必须考虑的因素。
目的：
1)为提高核酸检测样本的制备效率,将基于磁珠技术的自动核酸提取系统Maxwell 16 System应用于流感病毒核酸提取并分析该方法对实时RT-PCR的影响,以评估该自动系统用于流感病毒核酸检测的适宜性。
2)了解不同核酸提取试剂的裂解缓冲液稳定流感病毒RNA的能力,为将裂解缓冲液用作流感病毒核酸检测样品的储存和转运介质奠定基础。
3)测定常用的病毒核酸提取方法的裂解缓冲液灭活流感病毒的能力,以评估流感病毒核酸提取过程中的生物危害暴露风险。
方法：
1) Maxwell 16 System提取系列稀释的流感病毒培养液以及咽拭子、支气管肺泡灌洗液(bronchoalveolar lavage fluid, BALF)、人和禽粪便标本中的流感病毒核酸,以CDC开发的实时RT-PCR (real-time RT-PCR, rRT-PCR)方法检测提取物,所获结果与经典的基于硅胶柱层析的手工核酸提取试剂QIAamp Viral RNA Mini Kit (QIAamp Kit)提取所获得的结果相比较,分析Maxwell 16 System提取流感病毒核酸的敏感性、线性、重复性以及从多种临床和环境样本中提取流感病毒核酸的效率。
2)分别以QIAamp Kit中的Buffer AVL和Maxwell 16 System中的Lysis Solution两种裂解缓冲液处理流感病毒溶液,处理后的裂解缓冲液/病毒混合物分别保存于室温、4℃和-20℃或者进行多次冻融,以rRT-PCR分析病毒RNA量的变化,以扩增流感病毒基因组RNA第7节段全长的传统RT-PCR (conventional RT-PCR, cRT-PCR)评估病毒RNA的完整性。
3)以Maxwell 16 System的Lysis Solution QIAamp Kit的Buffer AVL和改良的异硫氰酸胍法试剂的RNA提取试剂A液(RNA Extraction Reagent A, ERA)等三种常用的病毒RNA提取方法的裂解缓冲液处理高滴度的流感病毒,系列对数稀释病毒处理液后接种于MDCK细胞进行分离培养,以倒置显微镜观察细胞生长状态,血凝实验确定流感病毒存在与否。
结果：
1)与QIAamp Kit比较,Maxwell 16 System的标准程序(Maxwell 16-S)有良好的线性、重复性以及更高的分析敏感性；Maxwell 16-S程序从咽拭子和BALF标本中可分离到更多的病毒RNA和/或更少的PCR抑制物,然而在粪便标本中的效率刚好相反,不能获得满意的效果；通过改良Maxwell 16程序,Maxwell 16 System清除粪便基质中PCR抑制物的能力得到改善,提取禽粪便标本病毒核酸的效率与QIAamp方法可比,而提取人粪便标本中病毒核酸的效率仍不及QIAamp方法。
2) Lysis Solution和Buffer AVL两种裂解缓冲液处理的病毒样品,-20℃放置160天和冻融5次均仍能收获量没有明显减少的完整的病毒基因组RNA；室温条件下,病毒RNA降解明显,两缓冲液处理后放置3天均不能确定收获到完整的流感病毒RNA分子；4℃条件下保存一定时间能收获到全长的流感病毒RNA分子；而在4℃和室温条件下,Buffer AVL处理较Lysis Solution处理后样品中的病毒RNA降解速度更快。
3) Lysis Solution、Buffer AVL和ERA三种裂解缓冲液处理病毒所获得的裂解溶液/病毒混合物,通过对数稀释结果发现,在低稀释度时和试剂对照一样,接种24h内引起MDCK细胞的完全死亡；在高稀释度时,三种裂解溶液/病毒混合物没有引起MDCK细胞病变,且培养物血凝实验阴性,而同时设置的病毒对照出现了明显的细胞病变,且培养物血凝实验阳性。
结论：
1) Maxwell 16 System结合rRT-PCR可以用于流感病毒的定量和定性测定；其清除不同样本基质中的PCR抑制物的能力不同,从而对rRT-PCR检测不同样本的敏感性产生影响,使其适用于咽拭子、BALF和禽粪标本中流感病毒RNA的提取,而不适于人粪便标本病毒核酸的提取；对于一些复杂的样本基质,适当地减少样本输入量可以提高检出。
2)不同的核酸提取试剂的裂解缓冲液用作流感病毒样本的保存剂都有助于病毒RNA的稳定性,可抵抗多次冻融对病毒RNA的损伤,冷藏或冷冻的低温条件下(特别是-20℃或更低温度)较长时间仍可收获到完整的流感病毒基因组RNA；但不同方法学的裂解缓冲液稳定病毒RNA的能力不尽相同,实际应用之前需进行验证。
3) Lysis Solution、Buffer AVL和ERA三种裂解溶液均可完全灭活流感病毒,经裂解缓冲液初步裂解后的核酸提取操作的生物危害风险大大降低。这对于将流感病毒核酸的自动提取仪器移出生物安全柜,或者在低防护水平区域或实验室进行核酸提取的初始裂解后操作,以及对于将裂解缓冲液用作流感病毒样品的稳定剂都有指导意义。
Background PCR-based nucleic acid (NA) testing is increasingly the first-choice method for virological surveillence and diagnosis of influenza. However, not only used NA assay protocols, reagents, primers and probes, but also pre-analytical variables, including the storage and transport of samples and NA purification methods, have major effects on the performance of NA assay. And, potential biohazard exposure to personnel during NA assay must be taken into consideration and would affect practical use of NA assay.
Objective
1) To apply Maxwell 16 System, an automated NA extraction platform based-on magnetic bead technology, to automated extraction of influenza virus (flu-v) RNA for diagnosis of flu using PCR-based testing and evaluated its performance and suitability for detection of flu-v from various samples.
2) To assess the effects of lysis buffers of different NA extraction methods on the stability of flu-v RNA in order to lay a foundation for the utilization of lysis buffer as preservative of flu-v samples.
3) To evaluate the biohazard risk during extraction of flu-v RNA by examining the ability of lysis buffers of frequently used NA extraction methods to inactivate flu-v.
Methods
1) Following extraction by Maxwell 16 System and QIAamp Viral RNA Mini Kit (QIAamp Kit) from flu-v stock dilution series, throat swabs, bronchoalveolar lavage fluid (BALF) and fecal samples of human and poultry origin, the extracted RNA was assayed by the CDC-developed real-time RT-PCR (rRT-PCR) protocols. The performance of Maxwell 16 System, including sensitivity, linearity, precision and extraction efficiency of flu-v RNA from the various clinical and field samples, was evaluated by comparison with QIAamp Kit based on silica gel column chromatography, a common used method for extraction of flu-v RNA.
2) Influenza virus stocks were treated respectively with two lysis buffers, Lysis Solution (Lysis Buffer+Proteinase K) from Maxwell 16 System and Buffer AVL with carrier RNA from QIAamp Kit, and the lysis buffer-virus mixtures were kept at 22℃,4℃,—20℃or underwent cyclic freeze-thaw. The relative quantity and quality of viral RNA after storage or freeze-thaw were evaluated with an rRT-PCR assay and a conventional RT-PCR assay targeting full-length matrix gene of flu-v.
3) Three lysis buffers, Lysis Solution, Buffer AVL and RNA Extraction Reagent A (ERA) from a modified guanidinium isothiocyanate kit, which based on three different methods commonly used in viral RNA extraction, were selected to process high-titer flu-v stocks (105.7TCID50/ml) according to manufacturer's instructions and preliminary research of ours. The lysis buffer-virus combinations were then serially tenfold diluted using virus growth medium and the dilutions were inoculated onto MDCK cells for virus isolation. Cell growth was observed daily under an inverted microscope. Cultures were harvested and hemagglutination tests were done with the supernatant when cytopathic effect (CPE) of cultured cells developed more than 75%.
Results
1) Extraction with Maxwell 16 System standard procedure (Maxwell 16-S) resulted in good linearity across a wide range, low inter-and intra-run variation and higher analysis sensitivity compared to those with the QIAamp extraction. Maxwell 16-S extraction yielded more RNA and/or fewer PCR inhibitors from throat swabs and BALF samples than QIAamp extraction but they had opposite performance for extraction from fecal samples. When extraction with a modified procedure (with reduced input and increased output) of Maxwell 16 System (Maxwell 16-M), the improved capacity to remove PCR inhibitors contributed to an enhanced sensitivity for detection of flu-v in poultry fecal samples although still a relative low sensitivity in human fecal samples compared to those with QIAamp Kit.
2) Entire matrix gene was amplified from samples treated with either of the lysis buffers after storage of 160 days at -20℃or five times of freeze-thaw, and the relative intensity of amplification products did not show to decline. Viral RNA in Buffer AVL degraded more rapidly than in Lysis Solution when kept at 22℃and 4℃, while the integrity of viral RNA could be maintain for certain time intervals at 4℃in both of the lysis buffers.
3) Massive cell death occurred within 24 hours after inoculation of lysis buffer-virus mixtures at low dilutions as well as the corresponding reagent controls at low dilutions. Cytopathic effect did not develop even after three blind passages of all the lysis buffer-virus mixtures at high dilutions and no viable virus in the cultures was detected by the hemagglutination tests, while infectious virus was isolated in the virus controls diluted at the same concentrations.
Conclusion
1) Qualitative and quantitative detection of flu-v is feasible using Maxwell 16 System in combination with rRT-PCR. But it differs in capacities of removing PCR inhibitors from different sample matrices, thereby having impacts on the sensitivity of rRT-PCR assay. A properly reduction of input volume of sample extracted with Maxwell 16 System for challenging matrices may improve detection.
2) The Maxwell 16-S procedure is suitable for extraction of flu-v RNA from throat swab and BALF samples, and the Maxwell 16-M procedure is suitable for extraction from poultry fecal samples, while a more optimized Maxwell 16 procedure is needed to develop for extraction from human fecal matrix.
3) Intact and non-decreased influenza virus RNA could obtained from samples preserved in lysis buffer underwent cyclic freeze-thaw or a long period of storage at refrigeration and frozen temperatures. However, lysis buffers different in methodology have not always the same ability to stabilize viral RNA.
4) The lysis buffers of the three nucleic acid extraction methods could render high-titer influenza virus non-infectious according to the procedures recommended by the manufacturers. Thus the risk of biohazard exposure during NA extraction was significantly reduced after initial lysis of flu-v samples, which has implications for moving processing of flu-v samples within an automated extractor outside of a biosafety cabinet or under low containment laboratories, and for application of lysis buffer as stabilizing agent of flu-v samples during storage and transport.
基于PCR的核酸测定技术已成为流感病毒学监测和诊断的首选方法。然而除了核酸测定方法、测定试剂、使用的引物和探针外,核酸测定分析前的样品储存、转运以及核酸分离纯化方法等变量也对核酸测定结果产生重要影响。此外,核酸测定过程中潜在的生物危害也是核酸测定技术实际应用必须考虑的因素。
目的：
1)为提高核酸检测样本的制备效率,将基于磁珠技术的自动核酸提取系统Maxwell 16 System应用于流感病毒核酸提取并分析该方法对实时RT-PCR的影响,以评估该自动系统用于流感病毒核酸检测的适宜性。
2)了解不同核酸提取试剂的裂解缓冲液稳定流感病毒RNA的能力,为将裂解缓冲液用作流感病毒核酸检测样品的储存和转运介质奠定基础。
3)测定常用的病毒核酸提取方法的裂解缓冲液灭活流感病毒的能力,以评估流感病毒核酸提取过程中的生物危害暴露风险。
方法：
1) Maxwell 16 System提取系列稀释的流感病毒培养液以及咽拭子、支气管肺泡灌洗液(bronchoalveolar lavage fluid, BALF)、人和禽粪便标本中的流感病毒核酸,以CDC开发的实时RT-PCR (real-time RT-PCR, rRT-PCR)方法检测提取物,所获结果与经典的基于硅胶柱层析的手工核酸提取试剂QIAamp Viral RNA Mini Kit (QIAamp Kit)提取所获得的结果相比较,分析Maxwell 16 System提取流感病毒核酸的敏感性、线性、重复性以及从多种临床和环境样本中提取流感病毒核酸的效率。
2)分别以QIAamp Kit中的Buffer AVL和Maxwell 16 System中的Lysis Solution两种裂解缓冲液处理流感病毒溶液,处理后的裂解缓冲液/病毒混合物分别保存于室温、4℃和-20℃或者进行多次冻融,以rRT-PCR分析病毒RNA量的变化,以扩增流感病毒基因组RNA第7节段全长的传统RT-PCR (conventional RT-PCR, cRT-PCR)评估病毒RNA的完整性。
3)以Maxwell 16 System的Lysis Solution QIAamp Kit的Buffer AVL和改良的异硫氰酸胍法试剂的RNA提取试剂A液(RNA Extraction Reagent A, ERA)等三种常用的病毒RNA提取方法的裂解缓冲液处理高滴度的流感病毒,系列对数稀释病毒处理液后接种于MDCK细胞进行分离培养,以倒置显微镜观察细胞生长状态,血凝实验确定流感病毒存在与否。
结果：
1)与QIAamp Kit比较,Maxwell 16 System的标准程序(Maxwell 16-S)有良好的线性、重复性以及更高的分析敏感性；Maxwell 16-S程序从咽拭子和BALF标本中可分离到更多的病毒RNA和/或更少的PCR抑制物,然而在粪便标本中的效率刚好相反,不能获得满意的效果；通过改良Maxwell 16程序,Maxwell 16 System清除粪便基质中PCR抑制物的能力得到改善,提取禽粪便标本病毒核酸的效率与QIAamp方法可比,而提取人粪便标本中病毒核酸的效率仍不及QIAamp方法。
2) Lysis Solution和Buffer AVL两种裂解缓冲液处理的病毒样品,-20℃放置160天和冻融5次均仍能收获量没有明显减少的完整的病毒基因组RNA；室温条件下,病毒RNA降解明显,两缓冲液处理后放置3天均不能确定收获到完整的流感病毒RNA分子；4℃条件下保存一定时间能收获到全长的流感病毒RNA分子；而在4℃和室温条件下,Buffer AVL处理较Lysis Solution处理后样品中的病毒RNA降解速度更快。
3) Lysis Solution、Buffer AVL和ERA三种裂解缓冲液处理病毒所获得的裂解溶液/病毒混合物,通过对数稀释结果发现,在低稀释度时和试剂对照一样,接种24h内引起MDCK细胞的完全死亡；在高稀释度时,三种裂解溶液/病毒混合物没有引起MDCK细胞病变,且培养物血凝实验阴性,而同时设置的病毒对照出现了明显的细胞病变,且培养物血凝实验阳性。
结论：
1) Maxwell 16 System结合rRT-PCR可以用于流感病毒的定量和定性测定；其清除不同样本基质中的PCR抑制物的能力不同,从而对rRT-PCR检测不同样本的敏感性产生影响,使其适用于咽拭子、BALF和禽粪标本中流感病毒RNA的提取,而不适于人粪便标本病毒核酸的提取；对于一些复杂的样本基质,适当地减少样本输入量可以提高检出。
2)不同的核酸提取试剂的裂解缓冲液用作流感病毒样本的保存剂都有助于病毒RNA的稳定性,可抵抗多次冻融对病毒RNA的损伤,冷藏或冷冻的低温条件下(特别是-20℃或更低温度)较长时间仍可收获到完整的流感病毒基因组RNA；但不同方法学的裂解缓冲液稳定病毒RNA的能力不尽相同,实际应用之前需进行验证。
3) Lysis Solution、Buffer AVL和ERA三种裂解溶液均可完全灭活流感病毒,经裂解缓冲液初步裂解后的核酸提取操作的生物危害风险大大降低。这对于将流感病毒核酸的自动提取仪器移出生物安全柜,或者在低防护水平区域或实验室进行核酸提取的初始裂解后操作,以及对于将裂解缓冲液用作流感病毒样品的稳定剂都有指导意义。
Background PCR-based nucleic acid (NA) testing is increasingly the first-choice method for virological surveillence and diagnosis of influenza. However, not only used NA assay protocols, reagents, primers and probes, but also pre-analytical variables, including the storage and transport of samples and NA purification methods, have major effects on the performance of NA assay. And, potential biohazard exposure to personnel during NA assay must be taken into consideration and would affect practical use of NA assay.
Objective
1) To apply Maxwell 16 System, an automated NA extraction platform based-on magnetic bead technology, to automated extraction of influenza virus (flu-v) RNA for diagnosis of flu using PCR-based testing and evaluated its performance and suitability for detection of flu-v from various samples.
2) To assess the effects of lysis buffers of different NA extraction methods on the stability of flu-v RNA in order to lay a foundation for the utilization of lysis buffer as preservative of flu-v samples.
3) To evaluate the biohazard risk during extraction of flu-v RNA by examining the ability of lysis buffers of frequently used NA extraction methods to inactivate flu-v.
Methods
1) Following extraction by Maxwell 16 System and QIAamp Viral RNA Mini Kit (QIAamp Kit) from flu-v stock dilution series, throat swabs, bronchoalveolar lavage fluid (BALF) and fecal samples of human and poultry origin, the extracted RNA was assayed by the CDC-developed real-time RT-PCR (rRT-PCR) protocols. The performance of Maxwell 16 System, including sensitivity, linearity, precision and extraction efficiency of flu-v RNA from the various clinical and field samples, was evaluated by comparison with QIAamp Kit based on silica gel column chromatography, a common used method for extraction of flu-v RNA.
2) Influenza virus stocks were treated respectively with two lysis buffers, Lysis Solution (Lysis Buffer+Proteinase K) from Maxwell 16 System and Buffer AVL with carrier RNA from QIAamp Kit, and the lysis buffer-virus mixtures were kept at 22℃,4℃,—20℃or underwent cyclic freeze-thaw. The relative quantity and quality of viral RNA after storage or freeze-thaw were evaluated with an rRT-PCR assay and a conventional RT-PCR assay targeting full-length matrix gene of flu-v.
3) Three lysis buffers, Lysis Solution, Buffer AVL and RNA Extraction Reagent A (ERA) from a modified guanidinium isothiocyanate kit, which based on three different methods commonly used in viral RNA extraction, were selected to process high-titer flu-v stocks (105.7TCID50/ml) according to manufacturer's instructions and preliminary research of ours. The lysis buffer-virus combinations were then serially tenfold diluted using virus growth medium and the dilutions were inoculated onto MDCK cells for virus isolation. Cell growth was observed daily under an inverted microscope. Cultures were harvested and hemagglutination tests were done with the supernatant when cytopathic effect (CPE) of cultured cells developed more than 75%.
Results
1) Extraction with Maxwell 16 System standard procedure (Maxwell 16-S) resulted in good linearity across a wide range, low inter-and intra-run variation and higher analysis sensitivity compared to those with the QIAamp extraction. Maxwell 16-S extraction yielded more RNA and/or fewer PCR inhibitors from throat swabs and BALF samples than QIAamp extraction but they had opposite performance for extraction from fecal samples. When extraction with a modified procedure (with reduced input and increased output) of Maxwell 16 System (Maxwell 16-M), the improved capacity to remove PCR inhibitors contributed to an enhanced sensitivity for detection of flu-v in poultry fecal samples although still a relative low sensitivity in human fecal samples compared to those with QIAamp Kit.
2) Entire matrix gene was amplified from samples treated with either of the lysis buffers after storage of 160 days at -20℃or five times of freeze-thaw, and the relative intensity of amplification products did not show to decline. Viral RNA in Buffer AVL degraded more rapidly than in Lysis Solution when kept at 22℃and 4℃, while the integrity of viral RNA could be maintain for certain time intervals at 4℃in both of the lysis buffers.
3) Massive cell death occurred within 24 hours after inoculation of lysis buffer-virus mixtures at low dilutions as well as the corresponding reagent controls at low dilutions. Cytopathic effect did not develop even after three blind passages of all the lysis buffer-virus mixtures at high dilutions and no viable virus in the cultures was detected by the hemagglutination tests, while infectious virus was isolated in the virus controls diluted at the same concentrations.
Conclusion
1) Qualitative and quantitative detection of flu-v is feasible using Maxwell 16 System in combination with rRT-PCR. But it differs in capacities of removing PCR inhibitors from different sample matrices, thereby having impacts on the sensitivity of rRT-PCR assay. A properly reduction of input volume of sample extracted with Maxwell 16 System for challenging matrices may improve detection.
2) The Maxwell 16-S procedure is suitable for extraction of flu-v RNA from throat swab and BALF samples, and the Maxwell 16-M procedure is suitable for extraction from poultry fecal samples, while a more optimized Maxwell 16 procedure is needed to develop for extraction from human fecal matrix.
3) Intact and non-decreased influenza virus RNA could obtained from samples preserved in lysis buffer underwent cyclic freeze-thaw or a long period of storage at refrigeration and frozen temperatures. However, lysis buffers different in methodology have not always the same ability to stabilize viral RNA.
4) The lysis buffers of the three nucleic acid extraction methods could render high-titer influenza virus non-infectious according to the procedures recommended by the manufacturers. Thus the risk of biohazard exposure during NA extraction was significantly reduced after initial lysis of flu-v samples, which has implications for moving processing of flu-v samples within an automated extractor outside of a biosafety cabinet or under low containment laboratories, and for application of lysis buffer as stabilizing agent of flu-v samples during storage and transport.
引文
1. Alexander, D. J.2007. An overview of the epidemiology of avian influenza. Vaccine 25:5637-44.
2. Amendola, A., A. Ranghiero, A. Zanetti, and E. Pariani.2011. Is avian influenza virus A(H5N1) a real threat to human health? J Prev Med Hyg 52:107-10.
3. Anwar, A., G Wan, K. B. Chua, J. T. August, and H. P. Too.2009. Evaluation of pre-analytical variables in the quantification of dengue virus by real-time polymerase chain reaction. J Mol Diagn 11:537-42.
4. Bartlett, J. G, and F. G Hayden.2005. Influenza A (H5N1):will it be the next pandemic influenza? Are we ready? Ann Intern Med 143:460-2.
5. Blow, J. A., D. J. Dohm, D. L. Negley, and C. N. Mores.2004. Virus inactivation by nucleic acid extraction reagents. J Virol Methods 119:195-8.
6. Blow, J. A., C. N. Mores, J. Dyer, and D. J. Dohm.2008. Viral nucleic acid stabilization by RNA extraction reagent. J Virol Methods 150:41-4.
7. Boggild, A. K., and A. J. McGeer.2010. Laboratory diagnosis of 2009 H1N1 influenza A virus. Crit Care Med 38:e38-42.
8. Brown, I. H. Summary of avian influenza activity in Europe, Asia, and Africa, 2006-2009. Avian Dis 54:187-93.
9. Das, A., E. Spackman, D. Senne, J. Pedersen, and D. L. Suarez.2006. Development of an internal positive control for rapid diagnosis of avian influenza virus infections by real-time reverse transcription-PCR with lyophilized reagents. J Clin Microbiol 44:3065-73.
10. Dwyer, D. E., D. W. Smith, M. G. Catton, and I. G Barr.2006. Laboratory diagnosis of human seasonal and pandemic influenza virus infection. Med J Aust 185:S48-53.
11. Espy, M. J., J. R. Uhl, L. M. Sloan, S. P. Buckwalter, M. F. Jones, E. A. Vetter, J. D. Yao, N. L. Wengenack, J. E. Rosenblatt, F. R. Cockerill,3rd, and T. F. Smith.2006. Real-time PCR in clinical microbiology:applications for routine laboratory testing. Clin Microbiol Rev 19:165-256.
12. Evers, D. L., R. D. Slemons, and J. K. Taubenberger.2007. Effect of preservative on recoverable RT-PCR amplicon length from influenza A virus in bird feces. Avian Dis 51:965-8.
13. Forster, J. L., V. B. Harkin, D. A. Graham, and S. J. McCullough.2008. The effect of sample type, temperature and RNAlater on the stability of avian influenza virus RNA. J Virol Methods 149:190-4.
14. Kimman, T. G, E. Smit, and M. R. Klein.2008. Evidence-based biosafety:a review of the principles and effectiveness of microbiological containment measures. Clin Microbiol Rev 21:403-25.
15. Krafft, A. E., K. L. Russell, A. W. Hawksworth, S. McCall, M. Irvine, L. T. Daum, J. L. Connoly, A. H. Reid, J. C. Gaydos, and J. K. Taubenberger. 2005. Evaluation of PCR testing of ethanol-fixed nasal swab specimens as an augmented surveillance strategy for influenza virus and adenovirus identification. J Clin Microbiol 43:1768-75.
16. Layne, S. P.2006. Human influenza surveillance:the demand to expand. Emerg Infect Dis 12:562-8.
17. Luinstra, K., A. Petrich, S. Castriciano, M. Ackerman, S. Chong, S. Carruthers, B. Ammons, J. B. Mahony, and M. Smieja.2011. Evaluation and clinical validation of an alcohol-based transport medium for preservation and inactivation of respiratory viruses. J Clin Microbiol 49:2138-42.
18. Peiris, J. S., M. D. de Jong, and Y. Guan.2007. Avian influenza virus (H5N1):a threat to human health. Clin Microbiol Rev 20:243-67.
19. Peiris, J. S., L. L. Poon, and Y. Guan.2009. Emergence of a novel swine-origin influenza A virus (S-OIV) H1N1 virus in humans. J Clin Virol 45:169-73.
20. Petrich, A., J. Mahony, S. Chong, G Broukhanski, F. Gharabaghi, G. Johnson, L. Louie, K. Luinstra, B. Willey, P. Akhaven, L. Chui, F. Jamieson, M. Louie, T. Mazzulli, R. Tellier, M. Smieja, W. Cai, M. Chernesky, and S. E. Richardson.2006. Multicenter comparison of nucleic acid extraction methods for detection of severe acute respiratory syndrome coronavirus RNA in stool specimens. J Clin Microbiol 44:2681-8.
21. Tania, N. QPCR:Target Preparation [M]//R. Mueller, S. Bustin. Real-time PCR in Microbiology-From Diagnosis to Characterization. Norfolk:Caister Academic Press,2007:71-99.
22. Rabenau, H. F., H. H. Kessler, M. Kortenbusch, A. Steinhorst, R. B. Raggam, and A. Berger.2007. Verification and validation of diagnostic laboratory tests in clinical virology. J Clin Virol 40:93-8.
23. Suarez, D. L., A. Das, and E. Ellis.2007. Review of rapid molecular diagnostic tools for avian influenza virus. Avian Dis 51:201-8.
24. Uyeki, T. M.2008. Global epidemiology of human infections with highly pathogenic avian influenza A (H5N1) viruses. Respirology 13 Suppl 1:S2-9.
25. Wang, R., and J. K. Taubenberger.2010. Methods for molecular surveillance of influenza. Expert Rev Anti Infect Ther 8:517-27.
26. WHO.2004. Laboratory Biosafety Manual-Third Edition [EB/OL]. [2012-04-15]. http://www.who.int/csr/resources/publications/biosafety/Biosafety7.pdf.
27. WHO.2011. The use of PCR in the surveillance and diagnosis of influenza [EB/OL]. [2012-04-16]. http://www.who.int/influenza/resources/documents/final_who_pcr_meeting_r eport_aug_2011_en.pdf.
28. Yang, G, D. E. Erdman, M. Kodani, J. Kools, M. D. Bowen, and B. S. Fields.2010. Comparison of commercial systems for extraction of nucleic acids from DNA/RNA respiratory pathogens. J Virol Methods 171:195-9.
1. Baumeister, A. K., M. Runge, M. Ganter, A. A. Feenstra, F. Delbeck, and H. Kirchhoff.1998. Detection of Mycoplasma hyopneumoniae in bronchoalveolar lavage fluids of pigs by PCR. J Clin Microbiol 36:1984-8.
2. Beck, E. T., and K. J. Henrickson.2010. Molecular diagnosis of respiratory viruses. Future Microbiol 5:901-16.
3. Chiu, R. W., Y. Jin, G. T. Chung, W. B. Lui, A. T. Chan, W. Lim, and Y. M. Dennis Lo.2006. Automated extraction protocol for quantification of SARS-coronavirus RNA in serum:an evaluation study. BMC Infect Dis 6:20.
4. Das, A., E. Spackman, M. J. Pantin-Jackwood, and D. L. Suarez.2009. Removal of real-time reverse transcription polymerase chain reaction (RT-PCR) inhibitors associated with cloacal swab samples and tissues for improved diagnosis of Avian influenza virus by RT-PCR. J Vet Diagn Invest 21:771-8.
5. Das, A., E. Spackman, D. Senne, J. Pedersen, and D. L. Suarez.2006. Development of an internal positive control for rapid diagnosis of avian influenza virus infections by real-time reverse transcription-PCR with lyophilized reagents. J Clin Microbiol 44:3065-73.
6. Espy, M. J., J. R. Uhl, L. M. Sloan, S. P. Buckwalter, M. F. Jones, E. A. Vetter, J. D. Yao, N. L. Wengenack, J. E. Rosenblatt, F. R. Cockerill,3rd, and T. F. Smith.2006. Real-time PCR in clinical microbiology:applications for routine laboratory testing. Clin Microbiol Rev 19:165-256.
7. Gobbers, E., T. A. Oosterlaken, M. J. van Bussel, R. Melsert, A. C. Kroes, and E. C. Claas.2001. Efficient extraction of virus DNA by NucliSens Extractor allows sensitive detection of hepatitis B virus by PCR. J Clin Microbiol 39:4339-43.
8. Goldstein, R. A., P. K. Rohatgi, E. H. Bergofsky, E. R. Block, R. P. Daniele, D. R. Dantzker, G. S. Davis, G. W. Hunninghake, T. E. King, Jr., W. J. Metzger, and et al.1990. Clinical role of bronchoalveolar lavage in adults with pulmonary disease. Am Rev Respir Dis 142:481-6.
9. Hale, A. D., J. Green, and D. W. Brown.1996. Comparison of four RNA extraction methods for the detection of small round structured viruses in faecal specimens. J Virol Methods 57:195-201.
10. Hourfar, M. K., U. Michelsen, M. Schmidt, A. Berger, E. Seifried, and W. K. Roth.2005. High-throughput purification of viral RNA based on novel aqueous chemistry for nucleic acid isolation. Clin Chem 51:1217-22.
11. Kou, Z., Y. Li, Z. Yin, S. Guo, M. Wang, X. Gao, P. Li, L. Tang, P. Jiang, Z. Luo, Z. Xin, C. Ding, Y. He, Z. Ren, P. Cui, H. Zhao, Z. Zhang, S. Tang, B. Yan, F. Lei, and T. Li.2009. The survey of H5N1 flu virus in wild birds in 14 Provinces of China from 2004 to 2007. PLoS One 4:e6926.
12. Lee, N., P. K. Chan, D. S. Hui, T. H. Rainer, E. Wong, K. W. Choi, G. C. Lui, B. C. Wong, R. Y. Wong, W. Y. Lam, I. M. Chu, R. W. Lai, C. S. Cockram, and J. J. Sung.2009. Viral loads and duration of viral shedding in adult patients hospitalized with influenza. J Infect Dis 200:492-500.
13. Li, I. W., I. F. Hung, K. K. To, K. H. Chan, S. S. Wong, J. F. Chan, V. C. Cheng, O. T. Tsang, S. T. Lai, Y. L. Lau, and K. Y. Yuen.2010. The natural viral load profile of patients with pandemic 2009 influenza A(H1N1) and the effect of oseltamivir treatment. Chest 137:759-68.
14. Loens, K., K. Bergs, D. Ursi, H. Goossens, and M. Ieven.2007. Evaluation of NucliSens easyMAG for automated nucleic acid extraction from various clinical specimens. J Clin Microbiol 45:421-5.
15. Lu, H., M. M. Ismail, O. A. Khan, Y. Al Hammad, S. S. Abdel Rhman, and M. H. Al-Blowi.2010. Epidemic outbreaks, diagnostics, and control measures of the H5N1 highly pathogenic avian influenza in the Kingdom of Saudi Arabia,2007-08. Avian Dis 54:350-6.
16. Mulrennan, S., S. S. Tempone, I. T. Ling, S. H. Williams, G. C. Gan, R. J. Murray, and D. J. Speers.2010. Pandemic influenza (H1N1) 2009 pneumonia:CURB-65 score for predicting severity and nasopharyngeal sampling for diagnosis are unreliable. PLoS One 5:el2849.
17. Perandin, F., P. C. Pollara, F. Gargiulo, C. Bonfanti, and N. Manca.2009. Performance evaluation of the automated NucliSens easyMAG nucleic acid extraction platform in comparison with QIAamp Mini kit from clinical specimens. Diagn Microbiol Infect Dis 64:158-65.
18. Petrich, A., J. Mahony, S. Chong, G. Broukhanski, F. Gharabaghi, G. Johnson, L. Louie, K. Luinstra, B. Willey, P. Akhaven, L. Chui, F. Jamieson, M. Louie, T. Mazzulli, R. Tellier, M. Smieja, W. Cai, M. Chernesky, and S. E. Richardson.2006. Multicenter comparison of nucleic acid extraction methods for detection of severe acute respiratory syndrome coronavirus RNA in stool specimens. J Clin Microbiol 44:2681-8.
19. Poon, L. L., K. H. Chan, O. K.Wong, W. C. Yam, K. Y. Yuen, Y. Guan, Y. M. Lo, and J. S. Peiris.2003. Early diagnosis of SARS coronavirus infection by real time RT-PCR. J Clin Virol 28:233-8.
20. Tania, N. QPCR:Target Preparation [M]//R. Mueller, S. Bustin. Real-time PCR in Microbiology-From Diagnosis to Characterization. Norfolk:Caister Academic Press,2007:71-99.
21. Promega Corporation.2009. Maxwell 16 Viral Total Nucleic Acid Purification System [EB/OL]. [2012-03-12]. http://www.promega.eom/-/media/Files/Resources/Protocols/Technical%20B ulletins/101/Maxwell%2016%20Viral%20Total%20Nucleic%20Acid%20Puri fication%20System%20 AS 1155%20Protocol.pdf.
22. Radstrom, P., R. Knutsson, P. Wolffs, M. Lovenklev, and C. Lofstrom. 2004. Pre-PCR processing:strategies to generate PCR-compatible samples. Mol Biotechnol 26:133-46.
23. Reed., L. J., and H. Muench.1938. A simple method for estimating fifty percent endpoints. Am J Hyg 27:493-497.
24. Reznikov, M., T. K. Blackmore, J. J. Finlay-Jones, and D. L. Gordon. 1995. Comparison of nasopharyngeal aspirates and throat swab specimens in a polymerase chain reaction-based test for Mycoplasma pneumoniae. Eur J Clin Microbiol Infect Dis 14:58-61.
25. Selvaraju, S. B., and R. Selvarangan.2010. Evaluation of three influenza A and B real-time reverse transcription-PCR assays and a new 2009 H1N1 assay for detection of influenza viruses. J Clin Microbiol 48:3870-5.
26. Suarez, D. L., A. Das, and E. Ellis.2007. Review of rapid molecular diagnostic tools for avian influenza virus. Avian Dis 51:201-8.
27. Tan, S. C., and B. C. Yiap.2009. DNA, RNA, and protein extraction:the past and the present. J Biomed Biotechnol 2009:574398.
28. Tewari, D., C. Zellers, H. Acland, and J. C. Pedersen.2007. Automated extraction of avian influenza virus for rapid detection using real-time RT-PCR. J Clin Virol 40:142-5.
29. Thorpe, C., and S. Darcy.2009. A/H1N1 flu pandemic. Consider bronchoalveolar lavage. BMJ 339:b5094.
30. Uyeki, T. M.2008. Global epidemiology of human infections with highly pathogenic avian influenza A (H5N1) viruses. Respirology 13 Suppl 1:S2-9.
31. Wang, H., Y. Mao, L. Ju, J. Zhang, Z. Liu, X. Zhou, Q. Li, Y. Wang, S. Kim, and L. Zhang.2004. Detection and monitoring of SARS coronavirus in the plasma and peripheral blood lymphocytes of patients with severe acute respiratory syndrome. Clin Chem 50:1237-40.
32. Wang, R., and J. K. Taubenberger.2010. Methods for molecular surveillance of influenza. Expert Rev Anti Infect Ther 8:517-27.
33. WHO.2009. CDC protocol of real time RTPCR for influenza A (H1N1) [EB/OL]. (2009-10-06) [2012-02-25]. http://www.who.int/csr/resources/publications/swineflu/CDCRealtimeRTPCR_SwineHlAssay-2009_20090430.pdf.
34. WHO.2006. Collecting, preserving and shipping specimens for the diagnosis of avian influenza A(H5N1) virus infection [EB/OL]. [2012-2-18]. http://www.who.int/csr/resources/publications/surveillance/CDS_EPR_ARO_2006_1.pdf.
35. WHO.2011. The use of PCR in the surveillance and diagnosis of influenza [EB/OL]. [2012-04-16]. http://www.who.int/influenza/resources/documents/final_who_pcr__meeting_r eport_aug_2011_en.pdf.
36. Wilson, I. G.1997. Inhibition and facilitation of nucleic acid amplification. Appl Environ Microbiol 63:3741-51.
37. Yoo, S. J., S. J. Moon, E. Y. Kuak, H. M. Yoo, C. K Kim, M. J. Chey, and B. M. Shin.2010. Frequent detection of pandemic (H1N1) 2009 virus in stools of hospitalized patients. J Clin Microbiol 48:2314-5.
38. Zaman, R. U., A. S. Alamgir, M. Rahman, E. Azziz-Baumgartner, E. S. Gurley, M. A. Sharker, W. A. Brooks, T. Azim, A. M. Fry, S. Lindstrom, L. V. Gubareva, X. Xu, R. J. Garten, M. J. Hossain, S. U. Khan, L.I. Faruque, S. S. Ameer, A. I. Klimov, and S. P. Luby.2009. Influenza in outpatient ILI case-patients in national hospital-based surveillance, Bangladesh,2007-2008. PLoS One 4:e8452.
39. 中国国家流感中心.2007.国家流感中心标准操作规程(修订版)[EB/OL].(2007-03-19) [2012-04-08]. http://www.cnic.org.cn/uploadfile/2009/1028/20091028104522533.rar.
1. Anwar, A., G Wan, K. B. Chua, J. T. August, and H. P. Too.2009. Evaluation of pre-analytical variables in the quantification of dengue virus by real-time polymerase chain reaction. J Mol Diagn 11:537-42.
2. Blacksell, S. D., S. Khounsy, and H. A. Westbury.2004. The effect of sample degradation and RNA stabilization on classical swine fever virus RT-PCR and ELISA methods. J Virol Methods 118:33-7.
3. Blow, J. A., D. J. Dohm, D. L. Negley, and C. N. Mores.2004. Virus inactivation by nucleic acid extraction reagents. J Virol Methods 119:195-8.
4. Blow, J. A., C. N. Mores, J. Dyer, and D. J. Dohm.2008. Viral nucleic acid stabilization by RNA extraction reagent. J Virol Methods 150:41-4.
5. Bruisten, S. M., P. Oudshoorn, P. van Swieten, B. Boeser-Nunnink, P. van Aarle, S. P. Tondreau, and H. T. Cuypers.1997. Stability of HIV-1 RNA in blood during specimen handling and storage prior to amplification by NASBA-QT. J Virol Methods 67:199-207.
6. Bustin, S. A., and T. Nolan.2004. Pitfalls of quantitative real-time reverse-transcription polymerase chain reaction. J Biomol Tech 15:155-66.
7. Evers, D. L., R. D. Slemons, and J. K. Taubenberger.2007. Effect of preservative on recoverable RT-PCR amplicon length from influenza A virus in bird feces. Avian Dis 51:965-8.
8. Fleige, S., and M. W. Pfaffl.2006. RNA integrity and the effect on the real-time qRT-PCR performance. Mol Aspects Med 27:126-39.
9. Forster, J. L., V. B. Harkin, D. A. Graham, and S. J. McCullough.2008. The effect of sample type, temperature and RNAlater on the stability of avian influenza virus RNA. J Virol Methods 149:190-4.
10. Frisbie, B., Y. W. Tang, M. Griffin, K. Poehling, P. F. Wright, K. Holland, and K. M. Edwards.2004. Surveillance of childhood influenza virus infection:what is the best diagnostic method to use for archival samples? J Clin Microbiol 42:1181-4.
11. Ghedin, E., J. Laplante, J. DePasse, D. E. Wentworth, R. P. Santos, M. L. Lepow, J. Porter, K. Stellrecht, X. Lin, D. Operario, S. Griesemer, A. Fitch, R. A. Halpin, T. B. Stockwell, D. J. Spiro, E. C. Holmes, and K. St George. 2011. Deep sequencing reveals mixed infection with 2009 pandemic influenza A (H1N1) virus strains and the emergence of oseltamivir resistance. J Infect Dis 203:168-74.
12. Inoue, E., X. Wang, Y. Osawa, and K. Okazaki.2010. Full genomic amplification and subtyping of influenza A virus using a single set of universal primers. Microbiol Immunol 54:129-34.
13. Kasahara, T., T. Miyazaki, H. Nitta, A. Ono, T. Miyagishima, T. Nagao, and T. Urushidani.2006. Evaluation of methods for duration of preservation of RNA quality in rat liver used for transcriptome analysis. J Toxicol Sci 31:509-19.
14. Krafft, A. E., K. L. Russell, A. W. Hawksworth, S. McCall, M. Irvine, L. T. Daum, J. L. Connoly, A. H. Reid, J. C. Gaydos, and J. K. Taubenberger. 2005. Evaluation of PCR testing of ethanol-fixed nasal swab specimens as an augmented surveillance strategy for influenza virus and adenovirus identification. J Clin Microbiol 43:1768-75.
15. Lee, D. H., L. Li, L. Andrus, and A. M. Prince.2002. Stabilized viral nucleic acids in plasma as an alternative shipping method for NAT. Transfusion 42:409-13.
16. Luinstra, K., A. Petrich, S. Castriciano, M. Ackerman, S. Chong, S. Carruthers, B. Ammons, J. B. Mahony, and M. Smieja.2011. Evaluation and clinical validation of an alcohol-based transport medium for preservation and inactivation of respiratory viruses. J Clin Microbiol 49:2138-42.
17. McCauley, J. W., and B. W. Mahy.1983. Structure and function of the influenza virus genome. Biochem J 211:281-94.
18. Mehlmann, M., M. B. Townsend, R. L. Stears, R. D. Kuchta, and K. L. Rowlen.2005. Optimization of fragmentation conditions for microarray analysis of viral RNA. Anal Biochem 347:316-23.
19. Nelson, M. I., Y. Tan, E. Ghedin, D. E. Wentworth, K. St George, L. Edelman, E. T. Beck, J. Fan, T. T. Lam, S. Kumar, D. J. Spiro, L. Simonsen, C. Viboud, E. C. Holmes, K. J. Henrickson, and J. M. Musser. 2010. Phylogeography of the spring and fall waves of the H1N1/09 pandemic influenza virus in the United States. J Virol 85:828-34.
20. QIAGEN Inc.2010. QIAamp Viral RNA Mini Handbook [EB/OL].[2012-04-23]. http://www.qiagen.com/literature/render.aspx?id=200375.
21. Rabenau, H. F., H. H. Kessler, M. Kortenbusch, A. Steinhorst, R. B. Raggam, and A. Berger.2007. Verification and validation of diagnostic laboratory tests in clinical virology. J Clin Virol 40:93-8.
22. Tania, N. QPCR:Target Preparation [M]//R. Mueller, S. Bustin. Real-time PCR in Microbiology-From Diagnosis to Characterization. Norfolk:Caister Academic Press,2007:71-99.
23. Runstadler, J. A., G M. Happ, R. D. Slemons, Z. M. Sheng, N. Gundlach, M. Petrula, D. Senne, J. Nolting, D. L. Evers, A. Modrell, H. Huson, S. Hills, T. Rothe, T. Marr, and J. K. Taubenberger.2007. Using RRT-PCR analysis and virus isolation to determine the prevalence of avian influenza virus infections in ducks at Minto Flats State Game Refuge, Alaska, during August 2005. Arch Virol 152:1901-10.
24. Sayan, M., M. Meric, S. Celebi, and A. Willke.2009. [Elimination of PCR inhibitors in routine diagnostic real-time PCR assay and results of internal amplification control]. Mikrobiyol Bul 43:179-81.
25. Schoor, O., T. Weinschenk, J. Hennenlotter, S. Corvin, A. Stenzl, H. G Rammensee, and S. Stevanovic.2003. Moderate degradation does not preclude microarray analysis of small amounts of RNA. Biotechniques 35:1192-6,1198-201.
26. Uhlenhaut, C., and M. Kracht.2005. Viral infectivity is maintained by an RNA protection buffer. J Virol Methods 128:189-91.
27. Wang, R., L. Soll, V. Dugan, J. Runstadler, G Happ, R. D. Slemons, and J. K. Taubenberger.2008. Examining the hemagglutinin subtype diversity among wild duck-origin influenza A viruses using ethanol-fixed cloacal swabs and a novel RT-PCR method. Virology 375:182-9.
28. Wang, R., and J. K. Taubenberger.2010. Methods for molecular surveillance of influenza. Expert Rev Anti Infect Ther 8:517-27.
29. Ward, C. L., M. H. Dempsey, C. J. Ring, R. E. Kempson, L. Zhang, D. Gor, B. W. Snowden, and M. Tisdale.2004. Design and performance testing of quantitative real time PCR assays for influenza A and B viral load measurement. J Clin Virol 29:179-88.
30. Whittier, C. A., W. Horne, B. Slenning, M. Loomis, and M. K. Stoskopf. 2004. Comparison of storage methods for reverse-transcriptase PCR amplification of rotavirus RNA from gorilla (Gorilla g. gorilla) fecal samples. J Virol Methods 116:11-7.
31. WHO.2009. CDC protocol of real time RTPCR for influenza A (H1N1) [EB/OL]. (2009-10-06) [2012-02-25]. http://www.who.int/csr/resources/publications/swineflu/CDCRealtimeRTPCR_SwineH1Assay-2009_20090430.pdf.
32. WHO.2011. Manual for the laboratory diagnosis and virological surveillance of influenza [EB/OL]. [2012-04-03]. http://whqlibdoc.who.int/publications/2011/9789241548090_eng.pdf.
33. WHO.2011. The use of PCR in the surveillance and diagnosis of influenza [EB/OL]. [2012-04-16]. http://www.who.int/influenza/resources/documents/final_who_pcr__meeting_r eport_aug_2011_en.pdf.
34. WHO.2009. WHO information for laboratory diagnosis of pandemic (H1N1) 2009 virus in humans-revised [EB/OL]. (2009-12-23) [2012-03-22]. http://www.who.int/csr/resources/publications/swineflu/WHO_Diagnostic_Re commendationsH IN 1_20090521.pdf.
35. Wilson, I. G.1997. Inhibition and facilitation of nucleic acid amplification. Appl Environ Microbiol 63:3741-51.
36. Zaman, R. U., A. S. Alamgir, M. Rahman, E. Azziz-Baumgartner, E. S. Gurley, M. A. Sharker, W. A. Brooks, T. Azim, A. M. Fry, S. Lindstrom, L. V. Gubareva, X. Xu, R. J. Garten, M. J. Hossain, S. U. Khan, L. I. Faruque, S. S. Ameer, A. I. Klimov, and S. P. Luby.2009. Influenza in outpatient ILI case-patients in national hospital-based surveillance, Bangladesh,2007-2008. PLoS One 4:e8452.
1. Bartlett, J. G., and F. G. Hayden.2005. Influenza A (H5N1):will it be the next pandemic influenza? Are we ready? Ann Intern Med 143:460-2.
2. Beck, E. T., and K. J. Henrickson.2010. Molecular diagnosis of respiratory viruses. Future Microbiol 5:901-16.
3. Blow, J. A., D. J. Dohm, D. L. Negley, and C. N. Mores.2004. Virus inactivation by nucleic acid extraction reagents. J Virol Methods 119:195-8.
4. Bruisten, S. M., P. Oudshoorn, P. van Swieten, B. Boeser-Nunnink, P. van Aarle, S. P. Tondreau, and H. T. Cuypers.1997. Stability of HIV-1 RNA in blood during specimen handling and storage prior to amplification by NASBA-QT. J Virol Methods 67:199-207.
5. Chirgwin, J. M., A. E. Przybyla, R. J. MacDonald, and W. J. Rutter.1979. Isolation of biologically active ribonucleic acid from sources enriched in ribonuclease. Biochemistry 18:5294-9.
6. Dang, J. L., K. Heroux, J. Kearney, A. Arasteh, M. Gostomski, and P. A. Emanuel.2001. Bacillus spore inactivation methods affect detection assays. Appl Environ Microbiol 67:3665-70.
7. Drosten, C., H. W. Doerr, W. Lim, K. Stohr, and M. Niedrig.2004. SARS molecular detection external quality assurance. Emerg Infect Dis 10:2200-3.
8. Dwyer, D. E., D. W. Smith, M. G. Catton, and I. G. Barr.2006. Laboratory diagnosis of human seasonal and pandemic influenza virus infection. Med J Aust 185:S48-53.
9. Espy, M. J., J. R. Uhl, L. M. Sloan, S. P. Buckwalter, M. F. Jones, E. A. Vetter, J. D. Yao, N. L. Wengenack, J. E. Rosenblatt, F. R. Cockerill,3rd, and T. F. Smith.2006. Real-time PCR in clinical microbiology:applications for routine laboratory testing. Clin Microbiol Rev 19:165-256.
10. Espy, M. J., J. R. Uhl, L. M. Sloan, J. E. Rosenblatt, F. R. Cockerill,3rd, and T. F. Smith.2002. Detection of vaccinia virus, herpes simplex virus, varicella-zoster virus, and Bacillus anthracis DNA by LightCycler polymerase chain reaction after autoclaving:implications for biosafety of bioterrorism agents. Mayo Clin Proc 77:624-8.
11. Kimman, T. G, E. Smit, and M. R. Klein.2008. Evidence-based biosafety:a review of the principles and effectiveness of microbiological containment measures. Clin Microbiol Rev 21:403-25.
12. Luinstra, K., A. Petrich, S. Castriciano, M. Ackerman, S. Chong, S. Carruthers, B. Ammons, J. B. Mahony, and M. Smieja.2011. Evaluation and clinical validation of an alcohol-based transport medium for preservation and inactivation of respiratory viruses. J Clin Microbiol 49:2138-42.
13. Luna, V. A., D. King, C. Davis, T. Rycerz, M. Ewert, A. Cannons, P. Amuso, and J. Cattani.2003. Novel sample preparation method for safe and rapid detection of Bacillus anthracis spores in environmental powders and nasal swabs. J Clin Microbiol 41:1252-5.
14. P.J, S., F. P.C, H. F.I, and Tellez I.2010. What Have We Learned from the Novel Influenza A (H1N1) Pandemic in 2009 for Strengthening Pandemic Influenza Preparedness?. Arch Med Res 40:673-676.
15. Ross, L., C. L. Vavro, S. L. Kehne, D. R. McClernon, and M. St Clair. 2001. Substitution of a commercially available, RNA extraction procedure in an HIV-1 genotyping system improves sensitivity and allows reduced sample volume. J Virol Methods 96:1-4.
16. Spackman, E., and D. L. Suarez.2005. Use of a novel virus inactivation method for a multicenter avian influenza real-time reverse transcnptase-polymerase chain reaction proficiency study. J Vet Diagn Invest 17:76-80.
17. Suarez, D. L., E. Spackman, D. A. Senne, L. Bulaga, A. C. Welsch, and K. Froberg.2003. The effect of various disinfectants on detection of avian influenza virus by real time RT-PCR. Avian Dis 47:1091-5.
18. Uyeki, T. M.2008. Global epidemiology of human infections with highly pathogenic avian influenza A (H5N1) viruses. Respirology 13 Suppl 1:S2-9.
19. Vonhippel, P. H., and K. Y. Wong.1964. Neutral Salts:The Generality of Their Effects on the Stability of Macromolecular Conformations. Science 145:577-80.
20. WHO.2010. H1N1 in post-pandemic period [EB/OL]. (2010-08-10) [2012-01-19] http://www.who.int/mediacentre/news/statements/2010/hlnl_vpc_20100810/e n/.
21. WHO.2004. Laboratory Biosafety Manual-Third Edition [EB/OL]. [2012-04-15]. http://www.who.int/csr/resources/publications/biosafety/Biosafety7.pdf.
22. WHO.2011. Manual for the laboratory diagnosis and virological surveillance of influenza [EB/OL]. [2012-04-13]. http://whqlibdoc.who.int/publications/2011/9789241548090_eng.pdf.
23. WHO.2005. WHO laboratory biosafety guidelines for handling specimens suspected of containing avian influenza A virus [EB/OL]. (2005-01-12) [2012-03-28] http://www.who.int/influenza/resources/documents/guidelines_handling_speci mens/en/.
24. 刘洪波,吴衍恒,朱远峰等.病毒RNA提取试剂对甲型流感病毒的灭活效果[J].中国热带医学,2011,11(7)：812-814.
1. Sambrook J, Russel D. Molecular Cloning:A Laboratory Manual [M].3rd ed. New York:Cold Spring Harbor Laboratory Press,2001.
2. Chomczynski P, Sacchi N. The single-step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction:twenty-something years on [J]. Nat Protoc,2006, 1(2):581-585.
3. Birnboim HC, Doly J. A rapid alkaline extraction procedure for screening recombinant plasmid DNA [J]. Nucleic Acids Res,1979,7(6):1513-1523.
4. Wink M. An Introduction to Molecular Biotechnology:Molecular Fundamentals, Methods and Application in Modern Biotechnology [M], Weinheim:Wiley-VCH, 2006.
5. Gjerde DT, Hoang L, Hornby D. RNA Purification and Analysis:Sample Preparation, Extraction, Chromatography [M],1st ed. Weinheim:Wiley-VCH, 2009.
6. Smith CE, Holmes DL, Simpson DJ, et al. Groseh. Mixed-bed solid phase and its use in the isolation of nucleic acids:US,6376194[P/OL]. 2002-04-23 [2010-12-02]. http://patft.uspto.gov/netacgi/nph-Parser?Sectl=PTO2&Sect2=HITOFF&p=1&u =%2Fnetahtml%2FPTO%2Fsearch-bool.html&r=1&f=G&1=50&co1=AND&d= PTXT&sl=%22Mixed-bed+solid+phase+isolation+nucleic+acids%22.TI.&OS= TTL/%22Mixed-bed+solid+phase+and+its+use+in+the+isolation+of+nucleic+aci ds%22&RS=TTL/%22Mixed-bed+solid+phase+and+its+use+in+the+isolation+o f+nucleic+acids%22.
7. Esser KH, Marx WH, Lisowsky T. MaxXbond:first regeneration system for DNA binding silica matrices [J]. Nat Methods,2006,3(1):1-2.
8. Dederich DA, Okwuonu G, Garner T, et al. Glass bead purification of plasmid template DNA for high throughtput sequencing of mammalian genomes [J]. Nucleic Acids Res,2002,30(7):e32.
9. Padhye W, York C, Burkiewiez A. Nucleic acid purification on silica gel and glass mixtures:US,5658548 [P/OL].1997-08-19[2010-12-02]. http://patft.uspto.gov/netacgi/nph-Parser?Sect1=PTO1&Sect2=HITOFF&d=PAL L&p=1&u=%2Fnetahtml%2FPTO%2Fsrchnum.htm&r=1&f=G&1=50&s1=5658 548.PN.&OS=PN/5658548&RS=PN/5658548
10. Little MC. Process for the purification of DNA on diatomaceous earth:US, 5075430 [P/OL].1991-12-24[2010-12-02]. http://patft.uspto.gov/netacgi/nph-Parser?Sect1=PTO2&Sect2=HITOFF&p=1&u =%2Fnetahtml%2FPTO%2Fsearch-bool.html&r=1&f=G&1=50&co1=AND&d= PTXT&s1=%22Process+purification+DNA+diatomaceous+earth%22.TI.&OS=T TL/
11. Berensmeier S. Magnetic particles for the separation and purification of nucleic acids [J]. Appl Microbiol Biotechnol,2006,73(3):495-504.
12. Nargessi RD. Magnetic isolation and purification of nucleic acids:US,6855499 [P/OL].2005-02-15[2010-12-02]. http://patft.uspto.gov/netacgi/nph-Parser?Sect1=PTO2&Sect2=HITOFF&p=1&u =%2Fnetahtml%2FPTO%2Fsearch-bool.html&r=1&f=G&1=50&co1=AND&d= PTXT&s1=%22Magnetic+isolation+purification+nucleic+acids%22.TI.&OS=TT L/.
13. Bio-Nobile. QuickPickTM kits for magnetic particle purifications [EB/OL]. [2010-10-22]. http://www.bionobile.com/Quick_kits.html.
14. QIAGEN Inc. QAsymphony DNA Handbook [EB/OL]. [2010-10-15]. http://www.qiagen.com/products/qiasymphonydnakit.aspx#Tabs=t1.
15. Applied Biosystems, MagMAXTM Total Nucleic Acid Isolation Kit [EB/OL]. [2010-11-18].https://products.appliedbiosystems.com/ab/en/US/adirect/ab?cmd=c atNavigate2&catID=604554&tab=DetailInfo.
16. Beckman Coulter Inc. Agencourt AMPure XP [EB/OL]. [2010-11-05]. http://www.beckmangenomics.com/products/dna_purification_and_cleanup/agen court_ampure_xp.html
17. QIAGEN Inc. QIAGEN Genomic DNA Handbook [EB/OL]. [2010-11-20]. http://www.qiagen.com/products/genomicdnastabilizationpurification/qiagengeno mictipsystem/qiagengenomic-tip 100g.aspx#Tabs=t2.
18. QIAGEN Inc. AllPrep DNA/RNA/Protein Mini Handbook [EB/OL]. [2010-12-10]. http://www.qiagen.com/products/rnastabilizationpurification/allprepdnarnaprotein minikit. aspx#Tabs=t2.
19. Terra-Ju Group. DeRiPRO DNA, RNA and Proteins Extraction Technology [EB/OL]. [2010-11-16]. http://terra-ju.com/TLS.htm.
20. Boyd J. Robotic laboratory automation [J]. Science,2002,295(5554):517-518.
21. Promega Corporation. Maxwell(?)16:Personal AutomationTM for the Forensic Lab [EB/OL]. [2010-11-25]. http://www.promega.com/maxwell16/forensic/default.htm.
22. Analytik Jena AG. InnuPure C12 Extraction System [EB/OL]. [2010-11-25]. http://www.analytik-jena.com/en/bio-solutions/Products-/Instruments/Autornatic-Purfication 4046/.
2. Amendola, A., A. Ranghiero, A. Zanetti, and E. Pariani.2011. Is avian influenza virus A(H5N1) a real threat to human health? J Prev Med Hyg 52:107-10.
3. Anwar, A., G Wan, K. B. Chua, J. T. August, and H. P. Too.2009. Evaluation of pre-analytical variables in the quantification of dengue virus by real-time polymerase chain reaction. J Mol Diagn 11:537-42.
4. Bartlett, J. G, and F. G Hayden.2005. Influenza A (H5N1):will it be the next pandemic influenza? Are we ready? Ann Intern Med 143:460-2.
5. Blow, J. A., D. J. Dohm, D. L. Negley, and C. N. Mores.2004. Virus inactivation by nucleic acid extraction reagents. J Virol Methods 119:195-8.
6. Blow, J. A., C. N. Mores, J. Dyer, and D. J. Dohm.2008. Viral nucleic acid stabilization by RNA extraction reagent. J Virol Methods 150:41-4.
7. Boggild, A. K., and A. J. McGeer.2010. Laboratory diagnosis of 2009 H1N1 influenza A virus. Crit Care Med 38:e38-42.
8. Brown, I. H. Summary of avian influenza activity in Europe, Asia, and Africa, 2006-2009. Avian Dis 54:187-93.
9. Das, A., E. Spackman, D. Senne, J. Pedersen, and D. L. Suarez.2006. Development of an internal positive control for rapid diagnosis of avian influenza virus infections by real-time reverse transcription-PCR with lyophilized reagents. J Clin Microbiol 44:3065-73.
10. Dwyer, D. E., D. W. Smith, M. G. Catton, and I. G Barr.2006. Laboratory diagnosis of human seasonal and pandemic influenza virus infection. Med J Aust 185:S48-53.
11. Espy, M. J., J. R. Uhl, L. M. Sloan, S. P. Buckwalter, M. F. Jones, E. A. Vetter, J. D. Yao, N. L. Wengenack, J. E. Rosenblatt, F. R. Cockerill,3rd, and T. F. Smith.2006. Real-time PCR in clinical microbiology:applications for routine laboratory testing. Clin Microbiol Rev 19:165-256.
12. Evers, D. L., R. D. Slemons, and J. K. Taubenberger.2007. Effect of preservative on recoverable RT-PCR amplicon length from influenza A virus in bird feces. Avian Dis 51:965-8.
13. Forster, J. L., V. B. Harkin, D. A. Graham, and S. J. McCullough.2008. The effect of sample type, temperature and RNAlater on the stability of avian influenza virus RNA. J Virol Methods 149:190-4.
14. Kimman, T. G, E. Smit, and M. R. Klein.2008. Evidence-based biosafety:a review of the principles and effectiveness of microbiological containment measures. Clin Microbiol Rev 21:403-25.
15. Krafft, A. E., K. L. Russell, A. W. Hawksworth, S. McCall, M. Irvine, L. T. Daum, J. L. Connoly, A. H. Reid, J. C. Gaydos, and J. K. Taubenberger. 2005. Evaluation of PCR testing of ethanol-fixed nasal swab specimens as an augmented surveillance strategy for influenza virus and adenovirus identification. J Clin Microbiol 43:1768-75.
16. Layne, S. P.2006. Human influenza surveillance:the demand to expand. Emerg Infect Dis 12:562-8.
17. Luinstra, K., A. Petrich, S. Castriciano, M. Ackerman, S. Chong, S. Carruthers, B. Ammons, J. B. Mahony, and M. Smieja.2011. Evaluation and clinical validation of an alcohol-based transport medium for preservation and inactivation of respiratory viruses. J Clin Microbiol 49:2138-42.
18. Peiris, J. S., M. D. de Jong, and Y. Guan.2007. Avian influenza virus (H5N1):a threat to human health. Clin Microbiol Rev 20:243-67.
19. Peiris, J. S., L. L. Poon, and Y. Guan.2009. Emergence of a novel swine-origin influenza A virus (S-OIV) H1N1 virus in humans. J Clin Virol 45:169-73.
20. Petrich, A., J. Mahony, S. Chong, G Broukhanski, F. Gharabaghi, G. Johnson, L. Louie, K. Luinstra, B. Willey, P. Akhaven, L. Chui, F. Jamieson, M. Louie, T. Mazzulli, R. Tellier, M. Smieja, W. Cai, M. Chernesky, and S. E. Richardson.2006. Multicenter comparison of nucleic acid extraction methods for detection of severe acute respiratory syndrome coronavirus RNA in stool specimens. J Clin Microbiol 44:2681-8.
21. Tania, N. QPCR:Target Preparation [M]//R. Mueller, S. Bustin. Real-time PCR in Microbiology-From Diagnosis to Characterization. Norfolk:Caister Academic Press,2007:71-99.
22. Rabenau, H. F., H. H. Kessler, M. Kortenbusch, A. Steinhorst, R. B. Raggam, and A. Berger.2007. Verification and validation of diagnostic laboratory tests in clinical virology. J Clin Virol 40:93-8.
23. Suarez, D. L., A. Das, and E. Ellis.2007. Review of rapid molecular diagnostic tools for avian influenza virus. Avian Dis 51:201-8.
24. Uyeki, T. M.2008. Global epidemiology of human infections with highly pathogenic avian influenza A (H5N1) viruses. Respirology 13 Suppl 1:S2-9.
25. Wang, R., and J. K. Taubenberger.2010. Methods for molecular surveillance of influenza. Expert Rev Anti Infect Ther 8:517-27.
26. WHO.2004. Laboratory Biosafety Manual-Third Edition [EB/OL]. [2012-04-15]. http://www.who.int/csr/resources/publications/biosafety/Biosafety7.pdf.
27. WHO.2011. The use of PCR in the surveillance and diagnosis of influenza [EB/OL]. [2012-04-16]. http://www.who.int/influenza/resources/documents/final_who_pcr_meeting_r eport_aug_2011_en.pdf.
28. Yang, G, D. E. Erdman, M. Kodani, J. Kools, M. D. Bowen, and B. S. Fields.2010. Comparison of commercial systems for extraction of nucleic acids from DNA/RNA respiratory pathogens. J Virol Methods 171:195-9.
1. Baumeister, A. K., M. Runge, M. Ganter, A. A. Feenstra, F. Delbeck, and H. Kirchhoff.1998. Detection of Mycoplasma hyopneumoniae in bronchoalveolar lavage fluids of pigs by PCR. J Clin Microbiol 36:1984-8.
2. Beck, E. T., and K. J. Henrickson.2010. Molecular diagnosis of respiratory viruses. Future Microbiol 5:901-16.
3. Chiu, R. W., Y. Jin, G. T. Chung, W. B. Lui, A. T. Chan, W. Lim, and Y. M. Dennis Lo.2006. Automated extraction protocol for quantification of SARS-coronavirus RNA in serum:an evaluation study. BMC Infect Dis 6:20.
4. Das, A., E. Spackman, M. J. Pantin-Jackwood, and D. L. Suarez.2009. Removal of real-time reverse transcription polymerase chain reaction (RT-PCR) inhibitors associated with cloacal swab samples and tissues for improved diagnosis of Avian influenza virus by RT-PCR. J Vet Diagn Invest 21:771-8.
5. Das, A., E. Spackman, D. Senne, J. Pedersen, and D. L. Suarez.2006. Development of an internal positive control for rapid diagnosis of avian influenza virus infections by real-time reverse transcription-PCR with lyophilized reagents. J Clin Microbiol 44:3065-73.
6. Espy, M. J., J. R. Uhl, L. M. Sloan, S. P. Buckwalter, M. F. Jones, E. A. Vetter, J. D. Yao, N. L. Wengenack, J. E. Rosenblatt, F. R. Cockerill,3rd, and T. F. Smith.2006. Real-time PCR in clinical microbiology:applications for routine laboratory testing. Clin Microbiol Rev 19:165-256.
7. Gobbers, E., T. A. Oosterlaken, M. J. van Bussel, R. Melsert, A. C. Kroes, and E. C. Claas.2001. Efficient extraction of virus DNA by NucliSens Extractor allows sensitive detection of hepatitis B virus by PCR. J Clin Microbiol 39:4339-43.
8. Goldstein, R. A., P. K. Rohatgi, E. H. Bergofsky, E. R. Block, R. P. Daniele, D. R. Dantzker, G. S. Davis, G. W. Hunninghake, T. E. King, Jr., W. J. Metzger, and et al.1990. Clinical role of bronchoalveolar lavage in adults with pulmonary disease. Am Rev Respir Dis 142:481-6.
9. Hale, A. D., J. Green, and D. W. Brown.1996. Comparison of four RNA extraction methods for the detection of small round structured viruses in faecal specimens. J Virol Methods 57:195-201.
10. Hourfar, M. K., U. Michelsen, M. Schmidt, A. Berger, E. Seifried, and W. K. Roth.2005. High-throughput purification of viral RNA based on novel aqueous chemistry for nucleic acid isolation. Clin Chem 51:1217-22.
11. Kou, Z., Y. Li, Z. Yin, S. Guo, M. Wang, X. Gao, P. Li, L. Tang, P. Jiang, Z. Luo, Z. Xin, C. Ding, Y. He, Z. Ren, P. Cui, H. Zhao, Z. Zhang, S. Tang, B. Yan, F. Lei, and T. Li.2009. The survey of H5N1 flu virus in wild birds in 14 Provinces of China from 2004 to 2007. PLoS One 4:e6926.
12. Lee, N., P. K. Chan, D. S. Hui, T. H. Rainer, E. Wong, K. W. Choi, G. C. Lui, B. C. Wong, R. Y. Wong, W. Y. Lam, I. M. Chu, R. W. Lai, C. S. Cockram, and J. J. Sung.2009. Viral loads and duration of viral shedding in adult patients hospitalized with influenza. J Infect Dis 200:492-500.
13. Li, I. W., I. F. Hung, K. K. To, K. H. Chan, S. S. Wong, J. F. Chan, V. C. Cheng, O. T. Tsang, S. T. Lai, Y. L. Lau, and K. Y. Yuen.2010. The natural viral load profile of patients with pandemic 2009 influenza A(H1N1) and the effect of oseltamivir treatment. Chest 137:759-68.
14. Loens, K., K. Bergs, D. Ursi, H. Goossens, and M. Ieven.2007. Evaluation of NucliSens easyMAG for automated nucleic acid extraction from various clinical specimens. J Clin Microbiol 45:421-5.
15. Lu, H., M. M. Ismail, O. A. Khan, Y. Al Hammad, S. S. Abdel Rhman, and M. H. Al-Blowi.2010. Epidemic outbreaks, diagnostics, and control measures of the H5N1 highly pathogenic avian influenza in the Kingdom of Saudi Arabia,2007-08. Avian Dis 54:350-6.
16. Mulrennan, S., S. S. Tempone, I. T. Ling, S. H. Williams, G. C. Gan, R. J. Murray, and D. J. Speers.2010. Pandemic influenza (H1N1) 2009 pneumonia:CURB-65 score for predicting severity and nasopharyngeal sampling for diagnosis are unreliable. PLoS One 5:el2849.
17. Perandin, F., P. C. Pollara, F. Gargiulo, C. Bonfanti, and N. Manca.2009. Performance evaluation of the automated NucliSens easyMAG nucleic acid extraction platform in comparison with QIAamp Mini kit from clinical specimens. Diagn Microbiol Infect Dis 64:158-65.
18. Petrich, A., J. Mahony, S. Chong, G. Broukhanski, F. Gharabaghi, G. Johnson, L. Louie, K. Luinstra, B. Willey, P. Akhaven, L. Chui, F. Jamieson, M. Louie, T. Mazzulli, R. Tellier, M. Smieja, W. Cai, M. Chernesky, and S. E. Richardson.2006. Multicenter comparison of nucleic acid extraction methods for detection of severe acute respiratory syndrome coronavirus RNA in stool specimens. J Clin Microbiol 44:2681-8.
19. Poon, L. L., K. H. Chan, O. K.Wong, W. C. Yam, K. Y. Yuen, Y. Guan, Y. M. Lo, and J. S. Peiris.2003. Early diagnosis of SARS coronavirus infection by real time RT-PCR. J Clin Virol 28:233-8.
20. Tania, N. QPCR:Target Preparation [M]//R. Mueller, S. Bustin. Real-time PCR in Microbiology-From Diagnosis to Characterization. Norfolk:Caister Academic Press,2007:71-99.
21. Promega Corporation.2009. Maxwell 16 Viral Total Nucleic Acid Purification System [EB/OL]. [2012-03-12]. http://www.promega.eom/-/media/Files/Resources/Protocols/Technical%20B ulletins/101/Maxwell%2016%20Viral%20Total%20Nucleic%20Acid%20Puri fication%20System%20 AS 1155%20Protocol.pdf.
22. Radstrom, P., R. Knutsson, P. Wolffs, M. Lovenklev, and C. Lofstrom. 2004. Pre-PCR processing:strategies to generate PCR-compatible samples. Mol Biotechnol 26:133-46.
23. Reed., L. J., and H. Muench.1938. A simple method for estimating fifty percent endpoints. Am J Hyg 27:493-497.
24. Reznikov, M., T. K. Blackmore, J. J. Finlay-Jones, and D. L. Gordon. 1995. Comparison of nasopharyngeal aspirates and throat swab specimens in a polymerase chain reaction-based test for Mycoplasma pneumoniae. Eur J Clin Microbiol Infect Dis 14:58-61.
25. Selvaraju, S. B., and R. Selvarangan.2010. Evaluation of three influenza A and B real-time reverse transcription-PCR assays and a new 2009 H1N1 assay for detection of influenza viruses. J Clin Microbiol 48:3870-5.
26. Suarez, D. L., A. Das, and E. Ellis.2007. Review of rapid molecular diagnostic tools for avian influenza virus. Avian Dis 51:201-8.
27. Tan, S. C., and B. C. Yiap.2009. DNA, RNA, and protein extraction:the past and the present. J Biomed Biotechnol 2009:574398.
28. Tewari, D., C. Zellers, H. Acland, and J. C. Pedersen.2007. Automated extraction of avian influenza virus for rapid detection using real-time RT-PCR. J Clin Virol 40:142-5.
29. Thorpe, C., and S. Darcy.2009. A/H1N1 flu pandemic. Consider bronchoalveolar lavage. BMJ 339:b5094.
30. Uyeki, T. M.2008. Global epidemiology of human infections with highly pathogenic avian influenza A (H5N1) viruses. Respirology 13 Suppl 1:S2-9.
31. Wang, H., Y. Mao, L. Ju, J. Zhang, Z. Liu, X. Zhou, Q. Li, Y. Wang, S. Kim, and L. Zhang.2004. Detection and monitoring of SARS coronavirus in the plasma and peripheral blood lymphocytes of patients with severe acute respiratory syndrome. Clin Chem 50:1237-40.
32. Wang, R., and J. K. Taubenberger.2010. Methods for molecular surveillance of influenza. Expert Rev Anti Infect Ther 8:517-27.
33. WHO.2009. CDC protocol of real time RTPCR for influenza A (H1N1) [EB/OL]. (2009-10-06) [2012-02-25]. http://www.who.int/csr/resources/publications/swineflu/CDCRealtimeRTPCR_SwineHlAssay-2009_20090430.pdf.
34. WHO.2006. Collecting, preserving and shipping specimens for the diagnosis of avian influenza A(H5N1) virus infection [EB/OL]. [2012-2-18]. http://www.who.int/csr/resources/publications/surveillance/CDS_EPR_ARO_2006_1.pdf.
35. WHO.2011. The use of PCR in the surveillance and diagnosis of influenza [EB/OL]. [2012-04-16]. http://www.who.int/influenza/resources/documents/final_who_pcr__meeting_r eport_aug_2011_en.pdf.
36. Wilson, I. G.1997. Inhibition and facilitation of nucleic acid amplification. Appl Environ Microbiol 63:3741-51.
37. Yoo, S. J., S. J. Moon, E. Y. Kuak, H. M. Yoo, C. K Kim, M. J. Chey, and B. M. Shin.2010. Frequent detection of pandemic (H1N1) 2009 virus in stools of hospitalized patients. J Clin Microbiol 48:2314-5.
38. Zaman, R. U., A. S. Alamgir, M. Rahman, E. Azziz-Baumgartner, E. S. Gurley, M. A. Sharker, W. A. Brooks, T. Azim, A. M. Fry, S. Lindstrom, L. V. Gubareva, X. Xu, R. J. Garten, M. J. Hossain, S. U. Khan, L.I. Faruque, S. S. Ameer, A. I. Klimov, and S. P. Luby.2009. Influenza in outpatient ILI case-patients in national hospital-based surveillance, Bangladesh,2007-2008. PLoS One 4:e8452.
39. 中国国家流感中心.2007.国家流感中心标准操作规程(修订版)[EB/OL].(2007-03-19) [2012-04-08]. http://www.cnic.org.cn/uploadfile/2009/1028/20091028104522533.rar.
1. Anwar, A., G Wan, K. B. Chua, J. T. August, and H. P. Too.2009. Evaluation of pre-analytical variables in the quantification of dengue virus by real-time polymerase chain reaction. J Mol Diagn 11:537-42.
2. Blacksell, S. D., S. Khounsy, and H. A. Westbury.2004. The effect of sample degradation and RNA stabilization on classical swine fever virus RT-PCR and ELISA methods. J Virol Methods 118:33-7.
3. Blow, J. A., D. J. Dohm, D. L. Negley, and C. N. Mores.2004. Virus inactivation by nucleic acid extraction reagents. J Virol Methods 119:195-8.
4. Blow, J. A., C. N. Mores, J. Dyer, and D. J. Dohm.2008. Viral nucleic acid stabilization by RNA extraction reagent. J Virol Methods 150:41-4.
5. Bruisten, S. M., P. Oudshoorn, P. van Swieten, B. Boeser-Nunnink, P. van Aarle, S. P. Tondreau, and H. T. Cuypers.1997. Stability of HIV-1 RNA in blood during specimen handling and storage prior to amplification by NASBA-QT. J Virol Methods 67:199-207.
6. Bustin, S. A., and T. Nolan.2004. Pitfalls of quantitative real-time reverse-transcription polymerase chain reaction. J Biomol Tech 15:155-66.
7. Evers, D. L., R. D. Slemons, and J. K. Taubenberger.2007. Effect of preservative on recoverable RT-PCR amplicon length from influenza A virus in bird feces. Avian Dis 51:965-8.
8. Fleige, S., and M. W. Pfaffl.2006. RNA integrity and the effect on the real-time qRT-PCR performance. Mol Aspects Med 27:126-39.
9. Forster, J. L., V. B. Harkin, D. A. Graham, and S. J. McCullough.2008. The effect of sample type, temperature and RNAlater on the stability of avian influenza virus RNA. J Virol Methods 149:190-4.
10. Frisbie, B., Y. W. Tang, M. Griffin, K. Poehling, P. F. Wright, K. Holland, and K. M. Edwards.2004. Surveillance of childhood influenza virus infection:what is the best diagnostic method to use for archival samples? J Clin Microbiol 42:1181-4.
11. Ghedin, E., J. Laplante, J. DePasse, D. E. Wentworth, R. P. Santos, M. L. Lepow, J. Porter, K. Stellrecht, X. Lin, D. Operario, S. Griesemer, A. Fitch, R. A. Halpin, T. B. Stockwell, D. J. Spiro, E. C. Holmes, and K. St George. 2011. Deep sequencing reveals mixed infection with 2009 pandemic influenza A (H1N1) virus strains and the emergence of oseltamivir resistance. J Infect Dis 203:168-74.
12. Inoue, E., X. Wang, Y. Osawa, and K. Okazaki.2010. Full genomic amplification and subtyping of influenza A virus using a single set of universal primers. Microbiol Immunol 54:129-34.
13. Kasahara, T., T. Miyazaki, H. Nitta, A. Ono, T. Miyagishima, T. Nagao, and T. Urushidani.2006. Evaluation of methods for duration of preservation of RNA quality in rat liver used for transcriptome analysis. J Toxicol Sci 31:509-19.
14. Krafft, A. E., K. L. Russell, A. W. Hawksworth, S. McCall, M. Irvine, L. T. Daum, J. L. Connoly, A. H. Reid, J. C. Gaydos, and J. K. Taubenberger. 2005. Evaluation of PCR testing of ethanol-fixed nasal swab specimens as an augmented surveillance strategy for influenza virus and adenovirus identification. J Clin Microbiol 43:1768-75.
15. Lee, D. H., L. Li, L. Andrus, and A. M. Prince.2002. Stabilized viral nucleic acids in plasma as an alternative shipping method for NAT. Transfusion 42:409-13.
16. Luinstra, K., A. Petrich, S. Castriciano, M. Ackerman, S. Chong, S. Carruthers, B. Ammons, J. B. Mahony, and M. Smieja.2011. Evaluation and clinical validation of an alcohol-based transport medium for preservation and inactivation of respiratory viruses. J Clin Microbiol 49:2138-42.
17. McCauley, J. W., and B. W. Mahy.1983. Structure and function of the influenza virus genome. Biochem J 211:281-94.
18. Mehlmann, M., M. B. Townsend, R. L. Stears, R. D. Kuchta, and K. L. Rowlen.2005. Optimization of fragmentation conditions for microarray analysis of viral RNA. Anal Biochem 347:316-23.
19. Nelson, M. I., Y. Tan, E. Ghedin, D. E. Wentworth, K. St George, L. Edelman, E. T. Beck, J. Fan, T. T. Lam, S. Kumar, D. J. Spiro, L. Simonsen, C. Viboud, E. C. Holmes, K. J. Henrickson, and J. M. Musser. 2010. Phylogeography of the spring and fall waves of the H1N1/09 pandemic influenza virus in the United States. J Virol 85:828-34.
20. QIAGEN Inc.2010. QIAamp Viral RNA Mini Handbook [EB/OL].[2012-04-23]. http://www.qiagen.com/literature/render.aspx?id=200375.
21. Rabenau, H. F., H. H. Kessler, M. Kortenbusch, A. Steinhorst, R. B. Raggam, and A. Berger.2007. Verification and validation of diagnostic laboratory tests in clinical virology. J Clin Virol 40:93-8.
22. Tania, N. QPCR:Target Preparation [M]//R. Mueller, S. Bustin. Real-time PCR in Microbiology-From Diagnosis to Characterization. Norfolk:Caister Academic Press,2007:71-99.
23. Runstadler, J. A., G M. Happ, R. D. Slemons, Z. M. Sheng, N. Gundlach, M. Petrula, D. Senne, J. Nolting, D. L. Evers, A. Modrell, H. Huson, S. Hills, T. Rothe, T. Marr, and J. K. Taubenberger.2007. Using RRT-PCR analysis and virus isolation to determine the prevalence of avian influenza virus infections in ducks at Minto Flats State Game Refuge, Alaska, during August 2005. Arch Virol 152:1901-10.
24. Sayan, M., M. Meric, S. Celebi, and A. Willke.2009. [Elimination of PCR inhibitors in routine diagnostic real-time PCR assay and results of internal amplification control]. Mikrobiyol Bul 43:179-81.
25. Schoor, O., T. Weinschenk, J. Hennenlotter, S. Corvin, A. Stenzl, H. G Rammensee, and S. Stevanovic.2003. Moderate degradation does not preclude microarray analysis of small amounts of RNA. Biotechniques 35:1192-6,1198-201.
26. Uhlenhaut, C., and M. Kracht.2005. Viral infectivity is maintained by an RNA protection buffer. J Virol Methods 128:189-91.
27. Wang, R., L. Soll, V. Dugan, J. Runstadler, G Happ, R. D. Slemons, and J. K. Taubenberger.2008. Examining the hemagglutinin subtype diversity among wild duck-origin influenza A viruses using ethanol-fixed cloacal swabs and a novel RT-PCR method. Virology 375:182-9.
28. Wang, R., and J. K. Taubenberger.2010. Methods for molecular surveillance of influenza. Expert Rev Anti Infect Ther 8:517-27.
29. Ward, C. L., M. H. Dempsey, C. J. Ring, R. E. Kempson, L. Zhang, D. Gor, B. W. Snowden, and M. Tisdale.2004. Design and performance testing of quantitative real time PCR assays for influenza A and B viral load measurement. J Clin Virol 29:179-88.
30. Whittier, C. A., W. Horne, B. Slenning, M. Loomis, and M. K. Stoskopf. 2004. Comparison of storage methods for reverse-transcriptase PCR amplification of rotavirus RNA from gorilla (Gorilla g. gorilla) fecal samples. J Virol Methods 116:11-7.
31. WHO.2009. CDC protocol of real time RTPCR for influenza A (H1N1) [EB/OL]. (2009-10-06) [2012-02-25]. http://www.who.int/csr/resources/publications/swineflu/CDCRealtimeRTPCR_SwineH1Assay-2009_20090430.pdf.
32. WHO.2011. Manual for the laboratory diagnosis and virological surveillance of influenza [EB/OL]. [2012-04-03]. http://whqlibdoc.who.int/publications/2011/9789241548090_eng.pdf.
33. WHO.2011. The use of PCR in the surveillance and diagnosis of influenza [EB/OL]. [2012-04-16]. http://www.who.int/influenza/resources/documents/final_who_pcr__meeting_r eport_aug_2011_en.pdf.
34. WHO.2009. WHO information for laboratory diagnosis of pandemic (H1N1) 2009 virus in humans-revised [EB/OL]. (2009-12-23) [2012-03-22]. http://www.who.int/csr/resources/publications/swineflu/WHO_Diagnostic_Re commendationsH IN 1_20090521.pdf.
35. Wilson, I. G.1997. Inhibition and facilitation of nucleic acid amplification. Appl Environ Microbiol 63:3741-51.
36. Zaman, R. U., A. S. Alamgir, M. Rahman, E. Azziz-Baumgartner, E. S. Gurley, M. A. Sharker, W. A. Brooks, T. Azim, A. M. Fry, S. Lindstrom, L. V. Gubareva, X. Xu, R. J. Garten, M. J. Hossain, S. U. Khan, L. I. Faruque, S. S. Ameer, A. I. Klimov, and S. P. Luby.2009. Influenza in outpatient ILI case-patients in national hospital-based surveillance, Bangladesh,2007-2008. PLoS One 4:e8452.
1. Bartlett, J. G., and F. G. Hayden.2005. Influenza A (H5N1):will it be the next pandemic influenza? Are we ready? Ann Intern Med 143:460-2.
2. Beck, E. T., and K. J. Henrickson.2010. Molecular diagnosis of respiratory viruses. Future Microbiol 5:901-16.
3. Blow, J. A., D. J. Dohm, D. L. Negley, and C. N. Mores.2004. Virus inactivation by nucleic acid extraction reagents. J Virol Methods 119:195-8.
4. Bruisten, S. M., P. Oudshoorn, P. van Swieten, B. Boeser-Nunnink, P. van Aarle, S. P. Tondreau, and H. T. Cuypers.1997. Stability of HIV-1 RNA in blood during specimen handling and storage prior to amplification by NASBA-QT. J Virol Methods 67:199-207.
5. Chirgwin, J. M., A. E. Przybyla, R. J. MacDonald, and W. J. Rutter.1979. Isolation of biologically active ribonucleic acid from sources enriched in ribonuclease. Biochemistry 18:5294-9.
6. Dang, J. L., K. Heroux, J. Kearney, A. Arasteh, M. Gostomski, and P. A. Emanuel.2001. Bacillus spore inactivation methods affect detection assays. Appl Environ Microbiol 67:3665-70.
7. Drosten, C., H. W. Doerr, W. Lim, K. Stohr, and M. Niedrig.2004. SARS molecular detection external quality assurance. Emerg Infect Dis 10:2200-3.
8. Dwyer, D. E., D. W. Smith, M. G. Catton, and I. G. Barr.2006. Laboratory diagnosis of human seasonal and pandemic influenza virus infection. Med J Aust 185:S48-53.
9. Espy, M. J., J. R. Uhl, L. M. Sloan, S. P. Buckwalter, M. F. Jones, E. A. Vetter, J. D. Yao, N. L. Wengenack, J. E. Rosenblatt, F. R. Cockerill,3rd, and T. F. Smith.2006. Real-time PCR in clinical microbiology:applications for routine laboratory testing. Clin Microbiol Rev 19:165-256.
10. Espy, M. J., J. R. Uhl, L. M. Sloan, J. E. Rosenblatt, F. R. Cockerill,3rd, and T. F. Smith.2002. Detection of vaccinia virus, herpes simplex virus, varicella-zoster virus, and Bacillus anthracis DNA by LightCycler polymerase chain reaction after autoclaving:implications for biosafety of bioterrorism agents. Mayo Clin Proc 77:624-8.
11. Kimman, T. G, E. Smit, and M. R. Klein.2008. Evidence-based biosafety:a review of the principles and effectiveness of microbiological containment measures. Clin Microbiol Rev 21:403-25.
12. Luinstra, K., A. Petrich, S. Castriciano, M. Ackerman, S. Chong, S. Carruthers, B. Ammons, J. B. Mahony, and M. Smieja.2011. Evaluation and clinical validation of an alcohol-based transport medium for preservation and inactivation of respiratory viruses. J Clin Microbiol 49:2138-42.
13. Luna, V. A., D. King, C. Davis, T. Rycerz, M. Ewert, A. Cannons, P. Amuso, and J. Cattani.2003. Novel sample preparation method for safe and rapid detection of Bacillus anthracis spores in environmental powders and nasal swabs. J Clin Microbiol 41:1252-5.
14. P.J, S., F. P.C, H. F.I, and Tellez I.2010. What Have We Learned from the Novel Influenza A (H1N1) Pandemic in 2009 for Strengthening Pandemic Influenza Preparedness?. Arch Med Res 40:673-676.
15. Ross, L., C. L. Vavro, S. L. Kehne, D. R. McClernon, and M. St Clair. 2001. Substitution of a commercially available, RNA extraction procedure in an HIV-1 genotyping system improves sensitivity and allows reduced sample volume. J Virol Methods 96:1-4.
16. Spackman, E., and D. L. Suarez.2005. Use of a novel virus inactivation method for a multicenter avian influenza real-time reverse transcnptase-polymerase chain reaction proficiency study. J Vet Diagn Invest 17:76-80.
17. Suarez, D. L., E. Spackman, D. A. Senne, L. Bulaga, A. C. Welsch, and K. Froberg.2003. The effect of various disinfectants on detection of avian influenza virus by real time RT-PCR. Avian Dis 47:1091-5.
18. Uyeki, T. M.2008. Global epidemiology of human infections with highly pathogenic avian influenza A (H5N1) viruses. Respirology 13 Suppl 1:S2-9.
19. Vonhippel, P. H., and K. Y. Wong.1964. Neutral Salts:The Generality of Their Effects on the Stability of Macromolecular Conformations. Science 145:577-80.
20. WHO.2010. H1N1 in post-pandemic period [EB/OL]. (2010-08-10) [2012-01-19] http://www.who.int/mediacentre/news/statements/2010/hlnl_vpc_20100810/e n/.
21. WHO.2004. Laboratory Biosafety Manual-Third Edition [EB/OL]. [2012-04-15]. http://www.who.int/csr/resources/publications/biosafety/Biosafety7.pdf.
22. WHO.2011. Manual for the laboratory diagnosis and virological surveillance of influenza [EB/OL]. [2012-04-13]. http://whqlibdoc.who.int/publications/2011/9789241548090_eng.pdf.
23. WHO.2005. WHO laboratory biosafety guidelines for handling specimens suspected of containing avian influenza A virus [EB/OL]. (2005-01-12) [2012-03-28] http://www.who.int/influenza/resources/documents/guidelines_handling_speci mens/en/.
24. 刘洪波,吴衍恒,朱远峰等.病毒RNA提取试剂对甲型流感病毒的灭活效果[J].中国热带医学,2011,11(7)：812-814.
1. Sambrook J, Russel D. Molecular Cloning:A Laboratory Manual [M].3rd ed. New York:Cold Spring Harbor Laboratory Press,2001.
2. Chomczynski P, Sacchi N. The single-step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction:twenty-something years on [J]. Nat Protoc,2006, 1(2):581-585.
3. Birnboim HC, Doly J. A rapid alkaline extraction procedure for screening recombinant plasmid DNA [J]. Nucleic Acids Res,1979,7(6):1513-1523.
4. Wink M. An Introduction to Molecular Biotechnology:Molecular Fundamentals, Methods and Application in Modern Biotechnology [M], Weinheim:Wiley-VCH, 2006.
5. Gjerde DT, Hoang L, Hornby D. RNA Purification and Analysis:Sample Preparation, Extraction, Chromatography [M],1st ed. Weinheim:Wiley-VCH, 2009.
6. Smith CE, Holmes DL, Simpson DJ, et al. Groseh. Mixed-bed solid phase and its use in the isolation of nucleic acids:US,6376194[P/OL]. 2002-04-23 [2010-12-02]. http://patft.uspto.gov/netacgi/nph-Parser?Sectl=PTO2&Sect2=HITOFF&p=1&u =%2Fnetahtml%2FPTO%2Fsearch-bool.html&r=1&f=G&1=50&co1=AND&d= PTXT&sl=%22Mixed-bed+solid+phase+isolation+nucleic+acids%22.TI.&OS= TTL/%22Mixed-bed+solid+phase+and+its+use+in+the+isolation+of+nucleic+aci ds%22&RS=TTL/%22Mixed-bed+solid+phase+and+its+use+in+the+isolation+o f+nucleic+acids%22.
7. Esser KH, Marx WH, Lisowsky T. MaxXbond:first regeneration system for DNA binding silica matrices [J]. Nat Methods,2006,3(1):1-2.
8. Dederich DA, Okwuonu G, Garner T, et al. Glass bead purification of plasmid template DNA for high throughtput sequencing of mammalian genomes [J]. Nucleic Acids Res,2002,30(7):e32.
9. Padhye W, York C, Burkiewiez A. Nucleic acid purification on silica gel and glass mixtures:US,5658548 [P/OL].1997-08-19[2010-12-02]. http://patft.uspto.gov/netacgi/nph-Parser?Sect1=PTO1&Sect2=HITOFF&d=PAL L&p=1&u=%2Fnetahtml%2FPTO%2Fsrchnum.htm&r=1&f=G&1=50&s1=5658 548.PN.&OS=PN/5658548&RS=PN/5658548
10. Little MC. Process for the purification of DNA on diatomaceous earth:US, 5075430 [P/OL].1991-12-24[2010-12-02]. http://patft.uspto.gov/netacgi/nph-Parser?Sect1=PTO2&Sect2=HITOFF&p=1&u =%2Fnetahtml%2FPTO%2Fsearch-bool.html&r=1&f=G&1=50&co1=AND&d= PTXT&s1=%22Process+purification+DNA+diatomaceous+earth%22.TI.&OS=T TL/
11. Berensmeier S. Magnetic particles for the separation and purification of nucleic acids [J]. Appl Microbiol Biotechnol,2006,73(3):495-504.
12. Nargessi RD. Magnetic isolation and purification of nucleic acids:US,6855499 [P/OL].2005-02-15[2010-12-02]. http://patft.uspto.gov/netacgi/nph-Parser?Sect1=PTO2&Sect2=HITOFF&p=1&u =%2Fnetahtml%2FPTO%2Fsearch-bool.html&r=1&f=G&1=50&co1=AND&d= PTXT&s1=%22Magnetic+isolation+purification+nucleic+acids%22.TI.&OS=TT L/.
13. Bio-Nobile. QuickPickTM kits for magnetic particle purifications [EB/OL]. [2010-10-22]. http://www.bionobile.com/Quick_kits.html.
14. QIAGEN Inc. QAsymphony DNA Handbook [EB/OL]. [2010-10-15]. http://www.qiagen.com/products/qiasymphonydnakit.aspx#Tabs=t1.
15. Applied Biosystems, MagMAXTM Total Nucleic Acid Isolation Kit [EB/OL]. [2010-11-18].https://products.appliedbiosystems.com/ab/en/US/adirect/ab?cmd=c atNavigate2&catID=604554&tab=DetailInfo.
16. Beckman Coulter Inc. Agencourt AMPure XP [EB/OL]. [2010-11-05]. http://www.beckmangenomics.com/products/dna_purification_and_cleanup/agen court_ampure_xp.html
17. QIAGEN Inc. QIAGEN Genomic DNA Handbook [EB/OL]. [2010-11-20]. http://www.qiagen.com/products/genomicdnastabilizationpurification/qiagengeno mictipsystem/qiagengenomic-tip 100g.aspx#Tabs=t2.
18. QIAGEN Inc. AllPrep DNA/RNA/Protein Mini Handbook [EB/OL]. [2010-12-10]. http://www.qiagen.com/products/rnastabilizationpurification/allprepdnarnaprotein minikit. aspx#Tabs=t2.
19. Terra-Ju Group. DeRiPRO DNA, RNA and Proteins Extraction Technology [EB/OL]. [2010-11-16]. http://terra-ju.com/TLS.htm.
20. Boyd J. Robotic laboratory automation [J]. Science,2002,295(5554):517-518.
21. Promega Corporation. Maxwell(?)16:Personal AutomationTM for the Forensic Lab [EB/OL]. [2010-11-25]. http://www.promega.com/maxwell16/forensic/default.htm.
22. Analytik Jena AG. InnuPure C12 Extraction System [EB/OL]. [2010-11-25]. http://www.analytik-jena.com/en/bio-solutions/Products-/Instruments/Autornatic-Purfication 4046/.
© 2004-2018 中国地质图书馆版权所有 京ICP备05064691号 京公网安备11010802017129号 地址：北京市海淀区学院路29号 邮编：100083 电话：办公室：(+86 10)66554848；文献借阅、咨询服务、科技查新：66554700 |