title: Multi-gene detection and identification of mosquito-borne RNA viruses using an oligonucleotide microarray.
abstract: BACKGROUND
  Arthropod-borne viruses are important emerging pathogens world-wide. Viruses transmitted by mosquitoes, such as dengue, yellow fever, and Japanese encephalitis viruses, infect hundreds of millions of people and animals each year. Global surveillance of these viruses in mosquito vectors using molecular based assays is critical for prevention and control of the associated diseases. Here, we report an oligonucleotide DNA microarray design, termed ArboChip5.1, for multi-gene detection and identification of mosquito-borne RNA viruses from the genera Flavivirus (family Flaviviridae), Alphavirus (Togaviridae), Orthobunyavirus (Bunyaviridae), and Phlebovirus (Bunyaviridae).
  
  
  RESULTS
  The assay utilizes targeted PCR amplification of three genes from each virus genus for electrochemical detection on a portable, field-tested microarray platform. Fifty-two viruses propagated in cell-culture were used to evaluate the specificity of the PCR primer sets and the ArboChip5.1 microarray capture probes. The microarray detected all of the tested viruses and differentiated between many closely related viruses such as members of the dengue, Japanese encephalitis, and Semliki Forest virus clades. Laboratory infected mosquitoes were used to simulate field samples and to determine the limits of detection. Additionally, we identified dengue virus type 3, Japanese encephalitis virus, Tembusu virus, Culex flavivirus, and a Quang Binh-like virus from mosquitoes collected in Thailand in 2011 and 2012.
  
  
  CONCLUSIONS
  We demonstrated that the described assay can be utilized in a comprehensive field surveillance program by the broad-range amplification and specific identification of arboviruses from infected mosquitoes. Furthermore, the microarray platform can be deployed in the field and viral RNA extraction to data analysis can occur in as little as 12 h. The information derived from the ArboChip5.1 microarray can help to establish public health priorities, detect disease outbreaks, and evaluate control programs.
authors: Virology Division, United States Army Medical Research Institute of Infectious Diseases, Fort Detrick, Frederick, Maryland, United States of America. nathan.grubaugh@colostate.edu
authors: Nathan D
authors: ND
authors: Grubaugh
authors: Scott S
authors: SS
authors: McMenamy
authors: Michael J
authors: MJ
authors: Turell
authors: John S
authors: JS
authors: Lee
mesh_terms: D000818:Animals; D009033:Culicidae / Q000821:virology*; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D012328:RNA Viruses / Q000145:classification / Q000235:genetics / Q000302:isolation & purification*; D013785:Thailand; D014773:Virology / Q000379:methods*
references: Microbiol Rev. 1994 Sep;58(3):491-562
references: 7968923
references: Annu Rev Microbiol. 2002;56:371-402
references: 12142476
references: Lancet. 1999 Oct 9;354(9186):1261-2
references: 10520637
references: Arch Virol. 2006 Nov;151(11):2253-60
references: 16820982
references: J Clin Virol. 2013 Mar;56(3):238-43
references: 23219893
references: Biosens Bioelectron. 2007 Apr 15;22(9-10):1853-60
references: 16891109
references: J Gen Virol. 2008 Oct;89(Pt 10):2580-2585
references: 18796727
references: PLoS Biol. 2003 Nov;1(2):E2
references: 14624234
references: J Am Mosq Control Assoc. 1996 Jun;12(2 Pt 1):167-71
references: 8827588
references: Arch Virol. 2007;152(12):2237-47
references: 17891328
references: J Biomol Tech. 2012 Apr;23(1):4-10
references: 22468136
references: J Clin Microbiol. 2006 Apr;44(4):1209-18
references: 16597840
references: PLoS One. 2007 Sep 26;2(9):e924
references: 17895966
references: Anal Chim Acta. 2010 Oct 29;679(1-2):85-90
references: 20951861
references: Nature. 2008 Feb 21;451(7181):990-3
references: 18288193
references: Proc Natl Acad Sci U S A. 2002 Nov 26;99(24):15687-92
references: 12429852
references: J Gen Virol. 2006 Apr;87(Pt 4):735-748
references: 16528021
references: Adv Virus Res. 2003;59:23-61
references: 14696326
references: Virology. 1975 Apr;64(2):367-77
references: 806166
references: Transbound Emerg Dis. 2012 Aug;59(4):336-43
references: 22103739
references: J Clin Microbiol. 2005 Aug;43(8):3779-87
references: 16081910
references: J Med Virol. 2005 Dec;77(4):528-40
references: 16254977
references: J Virol Methods. 1998 May;72(1):27-41
references: 9672130
references: J Clin Microbiol. 2010 Sep;48(9):3386-7
references: 20504990
references: Virology. 2012 Jun 5;427(2):90-7
references: 22425062
references: Arch Virol. 2003 Jun;148(6):1085-93
references: 12756616
references: J Gen Virol. 1990 Oct;71 ( Pt 10):2307-12
references: 2230736
references: Am J Trop Med Hyg. 2011 Jul;85(1):169-77
references: 21734144
references: Antiviral Res. 2010 Feb;85(2):328-45
references: 19857523
references: Am J Trop Med Hyg. 2010 Aug;83(2 Suppl):5-13
references: 20682900
references: J Virol. 2006 Jun;80(11):5627-30
references: 16699043
references: J Clin Microbiol. 2007 Feb;45(2):584-9
references: 17108075
references: Am J Trop Med Hyg. 2013 Feb;88(2):245-53
references: 23249687
references: Arch Virol. 2009;154(5):857-60
references: 19347244
references: Virology. 2007 Mar 15;359(2):405-14
references: 17070886
references: J Gen Virol. 2012 May;93(Pt 5):1023-1034
references: 22278828
references: J Gen Virol. 2008 Nov;89(Pt 11):2754-2760
references: 18931072
references: J Med Virol. 2009 Nov;81(11):1945-50
references: 19774692
references: J Clin Microbiol. 1996 May;34(5):1184-8
references: 8727900
references: Am J Trop Med Hyg. 2004 Feb;70(2):164-71
references: 14993628
references: Emerg Infect Dis. 2007 Jan;13(1):73-81
references: 17370518
references: J Virol Methods. 2010 Sep;168(1-2):147-51
references: 20471427
references: BMC Bioinformatics. 2006 Apr 28;7:232
references: 16643672
references: FEBS J. 2006 Jan;273(2):398-408
references: 16403026
references: Vector Borne Zoonotic Dis. 2009 Feb;9(1):25-32
references: 18759638
references: Am J Trop Med Hyg. 1984 Jan;33(1):176-81
references: 6696176
references: Virol J. 2008 Jan 24;5:16
references: 18218114
references: J Clin Microbiol. 2002 Jul;40(7):2323-30
references: 12089242
references: Virology. 2007 Nov 25;368(2):286-95
references: 17655905
references: Arch Virol. 2003 Jun;148(6):1095-118
references: 12756617
references: J Clin Microbiol. 2009 Aug;47(8):2398-404
references: 19535518
references: J Neuroimmune Pharmacol. 2010 Sep;5(3):428-42
references: 20652430
references: Am J Trop Med Hyg. 1974 Nov;23(6):1153-60
references: 4429185
references: J Biomed Biotechnol. 2012;2012:251364
references: 22829749