Accredited for compliance with ISO/IEC 17043 Accreditation Number: 14863 RCPAQAP Molecular Infectious Diseases Generic Report MAF1:2015 Molecular Alphavirus / Flavivirus Copyright This material is copyright and may not be used in any form for advertising, sales promotion or publicity. The material may not be reproduced in whole or in part for any purpose whatsoever (including presentations at meetings and conferences), without the prior written permission of the RCPA Quality Assurance Programs Pty Limited. Permission must be sought in writing from the Program but will not be unreasonably refused. Confidentiality RCPA Quality Assurance Programs Pty Limited keeps all participant details confidential. No information related to any of the participants will be divulged to a third party, unless required by legislation, without the express written consent of the participant. General information may be discussed at meetings or presented as papers to journals. RCPA Quality Phone +61 2 9045 6000 Assurance Programs Fax +61 2 9356 2003 ABN 32 003 520 072 Suite 201, 8 Herbert Street St Leonards NSW 2065 www.rcpaqap.com.au
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Accredited for compliance with ISO/IEC 17043Accreditation Number: 14863
CopyrightThis material is copyright and may not be used in any form for advertising, sales promotion or publicity. The material maynot be reproduced in whole or in part for any purpose whatsoever (including presentations at meetings and conferences),without the prior written permission of the RCPA Quality Assurance Programs Pty Limited. Permission must be sought inwriting from the Program but will not be unreasonably refused.
ConfidentialityRCPA Quality Assurance Programs Pty Limited keeps all participant details confidential. No information related to any of theparticipants will be divulged to a third party, unless required by legislation, without the express written consent of theparticipant. General information may be discussed at meetings or presented as papers to journals.
Generic Report MAF1:2015Participant ResultsThe Molecular Alphavirus/Flavivirus Program was distributed to 14 participants.12 participants returned results for this survey.
Clerical ErrorThere was a clerical error included in this survey for Specimen 1B.
9/12 (75%) participants detected the clerical error for Specimen 1B.
3/12 (25%) participants (1187, 83000 and 89082) did not identify the clerical error.
Specimen DetailsSamples consist of inactivated virus (gamma inactivation radiation (50 kGy)) from human strains of alphavirus orflavivirus suspended in 250uL of tissue culture fluid or molecular grade water.(Ref. 2015 RCPAQAP Molecular Infectious Diseases Program Plan section MAF1:2015)
1A Positive for Alphavirus, Ross River virus.
1B Negative, to assess contamination.
1C Positive for Flavivirus, Dengue 3 virus.
1D Positive for Alphavirus, Chikungunya virus.
1E Positive for Flavivirus, Dengue 1 virus.
1F Positive for Flavivirus, Japanese Encephalitis.
Source: Molecular Diagnostics & Development, Division of Microbiology and Infectious Diseases, PathWest LaboratoryMedicine, WA and RCPAQAP Serology MDM Serum Bank.
Preliminary & Pre-issue testing: Molecular Diagnostics & Development, Division of Microbiology and Infectious Diseases,PathWest Laboratory Medicine, WA and Public Health Laboratory, Queensland Health Forensic and Scientific Services,QLD.
Survey AimsThe detection and identification of Alphavirus (Ross River and Chikungunya) and Flavivirus (Dengue type 1, Dengue type3 and Japanese Encephalitis). A negative is included to assess contamination.
Clinical Notes (written by Dr Ed Raby)The “Clinical Notes” were created so as to highlight particular issues relating to interpretation of results. They do notnecessarily reflect the clinical history of the patient from whom the sample was obtained.
1A 24 year old female with fever, joint pains, rash. Rubella?
1B 42 year old female with polyarthralgia, tired.
1C 18 year old male, headache, rash, returned from Fiji.
1D 64 year old male, fever, hand and ankle pains, recent travel to Bali.
1E 32 year old male, travel to Thailand, fever, thrombocytopaenia.
1F 20 year old female, holiday work on pig farm in Vietnam, polio‐like syndrome.
Pre-issue ResultsThese results were obtained in pre-issue testing of the survey specimens.
Specimen Alphavirus Result Flavivirus Result Virus Identification[RT PCR]
1A positive negative Ross River
1B negative negative not applicable
1C negative positive Dengue 3
1D positive negative Chikungunya
1E negative positive Dengue 1
1F negative positive Japanese Encephalitis
Method PerformanceIt is up to all users to form their own opinion or conclusions on each kit's performance based on the accumulation ofdata and information that is supplied by this survey, and other information available to the participant.
Please note when primer sequence information is returned by participants it is not published to protect confidentiality.
ALPHAVIRUS Results10 participants returned results for Alphavirus molecular analysis.
Specimen 1A is positive for an Alphavirus; Ross River virus
3/4 participants (10003, 40002 and 83000) correctly detected an Alphavirus using Alphavirus group testing.Participant 10003 further commented on the identification of Ross River virus.
Participant 89112 failed to detect an Alphavirus using specific Alphavirus group testing.
All participants (40002, 60003 and 80071) performing specific Ross River virus testing identified the Ross River virus.
Specimen 1B is negative for an Alphavirus; all participants correctly returned a negative / not detected result.
Specimen 1C is negative for an Alphavirus; all participants correctly returned a negative / not detected result.
Specimen 1D is positive for an Alphavirus; Chikungunya virus.
3/4 participants (10003, 40002 and 83000) correctly detected an Alphavirus using Alphavirus group testing.Participants 10003 and 83000 further commented on the identification of Chikungunya virus.
Participant 89112 failed to detect an Alphavirus using specific Alphavirus group testing.
All participants (10003, 40002, 60003, 80011, 80042, 80069, 80071, 83000, 89082 and 89112) performing specificChikungunya virus testing identified the Chikungunya virus.
Specimen 1E is negative for an Alphavirus; all participants correctly returned a negative / not detected result.
Specimen 1F is negative for an Alphavirus; all participants correctly returned a negative / not detected result.
The results reported by Participant 1187 in the Alphavirus section were moved to the Dengue section as a data entryerror had been made.
Table 1 Alphavirus Group5 participants returned results.
Molecular A/F Alphavirus Group
Part1A 1B 1C 1D 1E 1F
Result Comment Result Comment Result Result Comment Result Result
10003 detected typed as ross rivervirus by sequencing
notdetected
notdetected detected
typed as chikungunyavirus by real-time PCR(Ct=20) & confirmed bysequencing
notdetected
notdetected
40002 detected notdetected
notdetected detected not
detectednot
detected
83000 detected A type of Alphavirusdetected
notdetected - not
detected detected
A type of Alphavirusdetected, typed asChikungunya by real-time RT-PCR
notdetected
notdetected
89112 negThis result does notexclude a recentAlpha virus infection
negThis result does notexclude a recentAlpha virus infection
nottested neg
This result does notexclude a recent Alphavirus infection
nottested
nottested
Participant 89112 appears to have used an expired amplification kit.
Participants 40002 and 89112 have not provided inhibition control information.
Table 2 Barmah Forest Virus2 participants returned results.
Molecular A/F Barmah Forest Virus
Part 1A 1B 1C 1D 1E 1F
40002 not detected not detected not tested not detected not tested not tested
60003 not detected not detected not detected not detected not detected not detected
Participant 40002 has not provided inhibition control information.
Table 3 Chikungunya Virus10 participants returned results.
Molecular A/F Chikungunya Virus
Part 1A 1B 1C 1D 1E 1F
10003 not detected not detected not detected detected not detected not detected
40002 not detected not detected not tested detected not tested not tested
60003 not detected not detected not detected detected not detected not detected
80011 not detected not detected not detected detected not detected not detected
80042 not detected not detected not detected detected not detected not detected
80069 not detected not detected not detected detected not detected not detected
80071 neg neg neg pos neg neg
83000 not detected not detected not detected detected not detected not detected
89082 neg neg neg pos neg neg
89112 neg neg not tested pos not tested not tested
Participant 80069 commented ‘Chikungunya not detected’ for Specimens 1A, 1B, 1C, 1E and 1F.
Participant 89112 commented ‘This result does not exclude a recent Chikungunya virus infection’ for Specimens 1A and1B. Participant 89112 commented ‘This result suggest a recent infection with Chikungunya virus’ for Specimen 1D.
Participants 40002, 80069, 80071, 89082 and 89112 have not provided inhibition control information.
Table 4 Ross River Virus3 participants returned results.
Molecular A/F Ross River Virus
Part 1A 1B 1C 1D 1E 1F
40002 detected not detected not tested not detected not tested not tested
60003 detected not detected not detected not detected not detected not detected
80071 pos neg neg neg neg neg
Participants 40002 and 80071 have not provided inhibition control information.
Table 5 Sindbis Virus1 participant returned results.
Molecular A/F Sindbis Virus
Part 1A 1B 1C 1D 1E 1F
40002 not detected not detected not tested not detected not tested not tested
Participant 40002 has not provided inhibition control information.
FLAVIVIRUS Results12 participants returned results for Flavivirus molecular analysis.
Specimen 1A is negative for a Flavivirus; all participants correctly returned a negative / not detected result.
Specimen 1B is negative for a Flavivirus; most participants correctly returned a negative / not detected result.
Participant 10003 incorrectly detected a Flavivirus using Flavivirus group testing and commented ‘typed as japaneseencephalitis virus by sequencing’. This participant reported a correct result of ‘not detected’ for specific JapaneseEncephalitis virus, however commented ‘positive in Flavivirus Group assay and typed as JEV by sequencing’.
Specimen 1C is positive for a Flavivirus; Dengue 3 virus.
All participants correctly detected a Flavivirus using Flavivirus group testing.Participants 10003 and 83000 also correctly commented on the identification of Dengue virus 3.
All participants performing specific Dengue virus testing identified the Dengue virus.Participants 10003, 40002, 60003, 80042, 80069, 80071 and 83000 also correctly identified the virus as Dengue type 3.
Specimen 1D is negative for a Flavivirus; most participants correctly returned a negative / not detected result.
Participant 10003 incorrectly detected a Flavivirus using Flavivirus group testing and commented ‘typed as dengue virustype 3 by real-time PCR (Ct=36) & confirmed by sequencing’. This participant also incorrectly identified Dengue type 3,using specific Dengue testing.
Specimen 1E is positive for a Flavivirus; Dengue 1 virus.
All participants correctly detected a Flavivirus using Flavivirus group testing.Participants 10003 and 83000 also correctly commented on the identification of Dengue virus 1.
All participants performing specific Dengue virus testing identified the Dengue virus.Participants 10003, 40002, 60003, 80042, 80069, 80071 and 83000 also correctly identified the virus as Dengue type 1.
Specimen 1F is positive for a Flavivirus; Japanese Encephalitis virus.
All participants correctly detected a Flavivirus using Flavivirus group testing.Participants 10003 and 83000 also correctly commented on the identification of Japanese Encephalitis virus.
All participants performing specific Japanese Encephalitis virus testing identified the Japanese Encephalitis virus.
Participant 60003 incorrectly reported ‘detected’ for this specimen using specific West Nile Virus assay, furthercommenting ‘WN and JE POS’.
Table 6 Flavivirus Group5 participants returned results.
Molecular A/F Flavivirus Group
Part1A 1B 1C 1D 1E 1F
Result Result Result Comment Result Result Comment Result Comment
10003 notdetected detected detected
typed as dengue virustype 3 by real-timePCR (Ct=17) &confirmed bysequencing
detected detected
typed as dengue virustype 1 by real-timePCR (Ct=18) &confirmed bysequencing
detected
typed as japaneseencephalitis virus byreal-time PCR (Ct=27)& confirmed bysequencing
40002 notdetected
notdetected detected not
detected detected detected
80071 neg neg pos neg pos pos
83000 notdetected
notdetected detected
A type of Flavivirusdetected, typed asDengue 3 by real-timeRT-PCR
notdetected detected
A type of Flavivirusdetected, typed asDengue 1 by real-timeRT-PCR
detectedA type of Flavivirusdetected, typed as JEby real-time RT-PCR
89112 neg neg posThis result suggest arecent infection withFlavivirus
neg posThis result suggest arecent infection withFlavivirus
posThis result suggest arecent infection withFlavivirus
Participant 10003 incorrectly returned ‘detected’ for Specimens 1B and 1D and further commented ‘typed as japaneseencephalitis virus by sequencing’ and ‘typed as dengue virus type 3 by real-time PCR (Ct=36) & confirmed by sequencing’respectively.
Participant 89112 commented ‘This result does not exclude a recent Flavivirus infection’ for Specimens 1A, 1B and 1D.
Participants 40002, 80071 and 89112 have not provided inhibition control information.
Table 7 Dengue Virus12 participants returned results.
Molecular A/F Dengue Virus
Part1A 1B 1C 1D 1E 1F
Result Result Result Comment Result Result Comment Result
1187 notdetected
notdetected detected not
detected detected notdetected
10003 notdetected
notdetected detected dengue virus type 3 detected detected detected dengue virus type 1
detectednot
detected
40002 nottested
notdetected detected Dengue 3 RNA was detected in
this samplenot
tested detected Dengue 1 RNA was detectedin this sample
notdetected
60003 notdetected
notdetected detected Dengue Type 3 POS not
detected detected Dengue Type 1 POS notdetected
80011 notdetected
notdetected detected not
detected detected notdetected
80042 notdetected
notdetected detected Dengue Type 3 not
detected detected Dengue Type 1 notdetected
80052 neg neg pos neg pos neg
80069 notdetected
notdetected detected DEN 3 Detected not
detected detected DEN 1 Detected notdetected
80071 neg neg pos D3 neg pos D1 neg
83000 notdetected
notdetected detected Dengue type 3 detected not
detected detected Dengue type 1 detected notdetected
89082 neg neg pos neg pos neg
89112 nottested
nottested pos This result suggest a recent
infection with Dengue virusnot
tested pos This result suggest a recentinfection with Dengue virus neg
Clinical Notes [Discussion by Dr Ed Raby, PathWest, WA]The “Clinical Notes” were created so as to highlight particular issues relating to interpretation of results. They do notnecessarily reflect the clinical history of the patient from whom the sample was obtained.
1A 24 year old female with fever, joint pains, rash. Rubella?Typical symptoms of RRV include a rash in about 50% of patients that may be clinically indistinguishable fromthe rash of rubella.
1B 42 year old female with polyarthralgia, tired.
1C 18 year old male, headache, rash, returned from Fiji.Fiji has had more than 25,000 suspected cases of Dengue with 15 deaths and remains an important source ofimported cases to Queensland.
1D 64 year old male, fever, hand and ankle pains, recent travel to Bali.Chikungunya is now endemic in Bali and may be difficult to distinguish clinically from Dengue Fever. The jointpain/arthralgia of CHIK infection typically affects the distal joints, similar to RRV/BFV and in contrast to Denguewhich is typically more central.
1E 32 year old male, travel to Thailand, fever, thrombocytopaenia.Low platelet count (thrombocytopaenia) is a common finding in Dengue Fever although significant haemorrhageremains very rare in adult travellers.
1F 20 year old female, holiday work on pig farm in Vietnam, polio‐like syndrome.Japanese Encephalitis Virus is present throughout Asia and causes epidemics and seasonal disease. It is rare intravellers without prolonged rural exposure. Polio‐like flaccid paralysis as well as more generalised encephalitisoccur in a minority of patients with most infections remaining asymptomatic.
AlphavirusSpecimens 1A and 1D were positive for an Alphavirus.
Four participants submitted group results. Three participants correctly detected the presence of an alphavirus in boththe above specimens. Participant 89112 failed to detected alphavirus in either Specimen 1A or 1D despite successfuldetection of both Ross River virus and Chikungunya virus in previous surveys using a pan-alpha PCR.
Three participants returning alphavirus results identified Ross River virus in Specimen 1A using a specific PCR.
Ten participants returning alphavirus results identified Chikungunya virus in Specimen 1D using a specific PCR.
There were no false positive results.
FlavivirusSpecimens 1C, 1E and 1F were positive for a Flavivirus.
Five participants submitted group results. All participants correctly detected the presence of a flavivirus in the abovespecimens. False positive results reported in Specimens 1B and 1D should be further investigated by Participant 10003.This may represent clerical error, however appears most likely to be due to cross-contamination of specimens duringprocessing.
Twelve participants returned Dengue virus specific assay results and although all were correctly detected for Specimens1C and 1E, only seven were able to go on and provide typing results.
Japanese encephalitis virus (Specimens 1F) was detected by all seven participants using a specific PCR. The mixeddetection reported by Participant 60003 (West Nile virus) has been reported in previous surveys as ‘cross reactivity’ andmost likely relates to primer specificity.
General CommentsIt is noted that the terminology used by several participants is incorrect, inconsistent and/or incomplete. A standardisedapproach to data returned to the RCPAQAP is encouraged. Generally in-house manufactured controls, amplification anddetection systems are poorly defined. Data entry errors are also noted. The general use of a Ct value of 0 to define anegative is not informative and is technically incorrect. Some indication of the upper and lower thresholds for each assaywould be more useful information. i.e. NEG = Ct >45 ; POS = Ct <35.
Participant 60003 reported using 160uL of specimen in the QIAamp Viral RNA Mini Kit. This is a non-standard specimenvolume for this purification method. The same participant reported the use of a 12uL volume of nucleic acid with 8uL ofreaction mix. These results are contradictory to previous submissions and may be clerical errors.
Participant 89112 used an expired Dengue virus kit. Although this did not seem to affect detection of these surveyspecimens, it may not perform as efficiently on specimens nearer to the lower limit of detection of the assay.
All participants are encouraged to incorporate an inhibition control into each assay and consider the development of apan-alpha / pan-flavi approach where possible.
There has been a large increase in the number of RRV cases reported to the Centre for Communicable DiseaseIntelligence from both NSW and QLD in the year to date and Dengue virus detections in returned traveller’s remainshigh. The use of nucleic acid amplification detection techniques in the detection of arboviruses should be limited to thetransient acute phase of infection and typing for epidemiology. All molecular results should be considered in conjunctionwith IgM and IgG serological results
Comments prepared for and on behalf of RCPAQAP by Ms Joanne Montgomery, PathWest Laboratory Medicine,Western Australia.
Alphavirus Amplification Kit Information for Survey MAF1:2015:1A, 1B, 1C, 1D, 1E & 1F
Part
Alphavirus - Amplification
Manufacturer Kit Name Method Automation Lot No. ExpiryDate
MasterVol
SampleVol
Single/Nested
10003 In house - RT PCR Eppendorf GradientS Thermalcycler NA NA 37 3 Nested
40002 In house - RT PCR Eppendorf GradientS Thermalcycler 19032015 Mar 16 15 5 Nested
83000 BioRad iScript One-StepRT-PCR iQ5 170-8893 Dec 15 20 5 Single round
89112 Roche AppliedScience
Expand HighFidelity PLUS
AB 2720 ThermalCycler 13434025 Mar 14 48.8 1 Nested
Part
Barmah Forest Virus - Amplification
Manufacturer Kit Name Method Automation Lot No. ExpiryDate
MasterVol
SampleVol Single/Nested
40002 In house - TaqMan RT-PCR Rotorgene 20052014 May 15 15 5 Single round60003 In house - TaqMan RT-PCR Rotorgene NA NA 8 12 Single round
Part
Chikungunya Virus - Amplification
Manufacturer Kit Name Method Automation Lot No. ExpiryDate
MasterVol
SampleVol
Single/Nested
10003 In house - TaqMan RT-PCR ABI Prism NA NA 17 3 Single round40002 In house - TaqMan RT-PCR Rotorgene 04032015 Mar 16 15 5 Single round60003 In house - TaqMan RT-PCR Rotorgene NA NA 12 8 Single round
80011 Invitrogen SuperScript III qRT-PCR RT PCR Light Cycler 1565698 Apr 16 15 5 Single round
80042 In house - Real-time PCR Light Cycler NA NA 20 5 Single round80069 AITbiotech abTES DEN 5 qPCR RT-PCR CFX96 C151-24 May 15 20 5 Single round80071 In house - Real-time PCR Light Cycler 10035000 Sep 15 9 1 Single round83000 Qiagen OneStep RT-PCR Kit RT-PCR - 148030425 Jul 15 20 5 Single round89082 In house - TaqMan RT-PCR LightCycler 110315 Mar 16 15 5 Single round
89112 RocheDiagnostics
LightCycler RNAAmplification kitHypro
RT-PCR Light Cycler 14748722 Jan 16 15 5 Single round
Part
Ross River Virus - Amplification
Manufacturer Kit Name Method Automation Lot No. ExpiryDate
MasterVol
SampleVol Single/Nested
40002 In house - TaqMan RT-PCR Rotorgene 05022015 Feb 16 15 5 Single round60003 In house - TaqMan RT-PCR Rotorgene NA NA 12 8 Single round80071 In house - Real-time PCR Light Cycler 10035000 Sep 15 9 1 Single round
Part
Sindbis Virus - Amplification
Manufacturer Kit Name Method Automation Lot No. ExpiryDate
MasterVol
SampleVol Single/Nested
40002 In house - TaqMan RT-PCR Rotorgene 21012015 Jan 16 15 5 Single round
Alphavirus Detection Kit Information for Survey MAF1:2015:1A, 1B, 1C, 1D, 1E & 1F
Part
Alphavirus Group - Detection
Manufacturer Kit Name Method Automation Lot No. ExpiryDate Single/Nested
10003 In house - Gelelectrophoresis - NA NA Nested
40002 Qiagen QIAxcel DNA HighResolution Kit QIAxcel C131031033 May 15 Nested
83000 BioRad iScript One-Step RT-PCR iQ5 170-8893 Dec 15 Single round
89112 UV Products UV Transilluminator GeneSnap fromSynGene IN HOUSE Oct 15 Nested
Part
Barmah Forest Virus - Detection
Manufacturer Kit Name Method Automation Lot No. ExpiryDate Single/Nested
40002 In house - TaqMan RT-PCR Rotorgene 20052014 May 15 Single round60003 In house - TaqMan RT-PCR Rotorgene NA NA Single round
Part
Chikungunya Virus - Detection
Manufacturer Kit Name Method Automation Lot No. ExpiryDate Single/Nested
10003 In house - TaqMan RT-PCR ABI Prism NA NA Single round40002 In house - TaqMan RT-PCR Rotorgene 04032015 Mar 16 Single round60003 In house - TaqMan RT-PCR Rotorgene NA NA Nested80011 In house - TaqMan RT-PCR LightCycler 1565698 Apr 16 Single round80042 In house - Real-time PCR Light Cycler NA NA Single round80069 AITbiotech abTES DEN 5 qPCR RT-PCR CFX96 C151-24 May 15 Single round80071 In house - Real-time PCR Light Cycler 10035000 Sep 15 Single round
83000 Qiagen QuantiTect Probe RT-PCR Kit
Fluorescent(FAM channel) - 148030425 Jul 15 Single round
89082 In house - TaqMan RT-PCR LightCycler 110315 Mar 16 Single round89112 In house - TaqMan RT-PCR LightCycler IN HOUSE Jan 16 Single round
Part
Ross River Virus - Detection
Manufacturer Kit Name Method Automation Lot No. ExpiryDate Single/Nested
40002 In house - TaqMan RT-PCR Rotorgene 05022015 Feb 16 Single round60003 In house - TaqMan RT-PCR Rotorgene NA NA Single round80071 In house - Real-time PCR Light Cycler 10035000 Sep 15 Single round
PartSindbis Virus - Detection
Manufacturer Kit Name Method Automation Lot No. Expiry Date Single/Nested
40002 In house - TaqMan RT-PCR Rotorgene 21012015 Jan 16 Single round
40002 N/A in house NA Probe Control, Primer Control 23072014 ,16012015 NA Taqman RT-PCR Probe Fluorescence60003 In-house In-house Reference strain Yes Realtime PCR
West Nile Virus - Controls & In house Methods
PartInhib Control Neg Control Pos Control In-house Methods
Manufacturer Lot No. Expiry Manufacturer Lot No. Expiry Manufacturer Lot No. Expiry In-house Method Detection Method
40002 Probe Control; Primer Control 22072010,12032013 NA TaqMan RT-PCR Probe Flourescence
60003 In-house In-house Reference strain Yes Realtime PCR80069 NA NA NA In house NA NA In house NA NA Conventional RT-PCR Gel electrophoresis89082 In house Mar 15 Mar 16 In house Mar 15 Mar 16
89112 NA Promega (nucleasefree water) 14748722 Jan 2016 In house (WNV RNA as Positive
Control) WN 2651 TC1 NA NA Conventional PCR(Roche Titan one step)
Agarose gelelectrophoresis
Yellow Fever Virus - Controls & In house Methods
PartInhib Control Neg Control Pos Control In-house Methods
Manufacturer Lot No. Expiry Manufacturer Lot No. Expiry Manufacturer Lot No. Expiry In-house Method Detection Method
40002 N/A Inhouse Neg Control NA Primer Control, Probe Control 12122013,12122013 NA Taqman RT-PCR Probe Flourescence
80069 NA NA NA In house NA NA In house NA NA Conventional RT-PCR Gel Electrophoresis
80071 NA NA NA Water NA NA In house (Yellow fever virus celllysate RNA as Positive Control) NA NA Designed in-house
RT-PCR protocolLightCycler® RNAMaster SYBR Green I
89082 In house Mar 15 Mar 16 In house Mar 15 Mar 16
89112 NA Promega (nucleasefree water) 14748722 Jan 2016 In house YFV (RNA) as positive
control YF20/11/121 NA NA Conventional RT-PCR(Roche Titan one step)
Flavivirus Units and Cut-Off Values for Survey MAF1:2015:1A, 1B, 1C, 1D, 1E & 1F
Part UnitsFlavivirus Group - Units & Cut Off Values
Neg Equ Pos
10003 PCR product size no specific electrophoresis sizeband/IC OK
specific electrophoresis sizeband/IC OK
40002 band intensity
80071 Cp No product formation at meltingcurve analysis
Product formation at meltingcurve analysis
83000 Ct No band seen - Band seen89112 No units available
Part UnitsDengue Virus - Units & Cut Off Values
Neg Equ Pos
1187 Ct 40 and below - Less than 4010003 Ct no specific amplification/IC OK specific amplification/IC OK40002 Ct 40 37-39 3660003 Ct 0 39-40 1-3980011 Ct80042 Ct 45 40-45 1-4080052 Ct 39 37-39 3780069 Ct 0.00 & above 40.00 - Below 40.0
80071 Cp No amplification is detected atabs.quantification
Amplification is detected atabs.quantification
83000 Ct Below Ct value - Above Ct value89082 Ct 45 40 2989112 No units available
Part UnitsJapanese Encephalitis Virus - Units & Cut Off Values
Neg Equ Pos
10003 Ct no specific amplification/IC OK specific amplification/IC OK40002 Ct 40 37-39 3660003 Ct 0 39-40 1-3980069 - No specific band seen - Specific band seen
80071 Cp No product formation at meltingcurve analysis
83000 Bioline UK MyTaq One-StepRT-PCR - ABI Veriti MTOS10-
112C May 15 20 5 Nested
89112 Roche AppliedScience
Titan One tubeRT-PCR RT-PCR AB 2720 Thermal
Cycler 10774220 Oct 15 40 10 Single round
Part
Dengue Virus - Amplification
Manufacturer Kit Name Method Automation Lot No. ExpiryDate
MasterVol
SampleVol
Single/Nested
1187 AIT BiotechPte Ltd
abTes TM DEN 4qPCR 1 Kit Real Time PCR Biorad CFX96 C155-26 Aug 15 20 5 Single round
10003 In house - TaqMan RT-PCR ABI Prism NA NA 17 3 Single round40002 In house - TaqMan RT-PCR Rotorgene 18022015 Feb 16 15 5 Single round60003 In house - TaqMan RT-PCR Rotorgene NA NA 12 8 Single round
Manufacturer Kit Name Method Automation Lot No. ExpiryDate
MasterVol
SampleVol
Single/Nested
10003 In house - TaqMan RT-PCR ABI Prism NA NA 17 3 Single round40002 In house - TaqMan RT-PCR Rotorgene 11022015 Feb 16 15 5 Single round60003 In house - TaqMan RT-PCR Rotorgene NA NA Single round
83000 In house - Gel electrophoresis - 0000398806 Nov 15 Single round
89112 UV Products UV Transilluminator GeneSnapfrom SynGene IN HOUSE Oct 15 Single round
Part
Dengue Virus - Detection
Manufacturer Kit Name Method Automation Lot No. ExpiryDate Single/Nested
1187 AIT BiotechPte Ltd
abTes TM DEN 4qPCR 1 Kit Real Time PCR Biorad CFX96 C155-26 Aug 15 Single round
10003 In house - TaqMan RT-PCR ABI Prism NA NA Single round40002 In house - TaqMan RT-PCR Rotorgene 18022015 Feb 16 Single round60003 In house - TaqMan RT-PCR Rotorgene NA NA Nested80011 In house - TaqMan RT-PCR LightCycler 1565698 Apr 16 Single round80042 In house - Real-time PCR ABI 7500 N/A NA Single round80052 In house - TaqMan RT-PCR Rotorgene 6089093 NA Single round80069 AITbiotech abTES DEN 5 qPCR RT-PCR CFX96 C151-24 May 15 Single round80071 In house - Real-time PCR Light Cycler 10772000 Aug 16 Single round
Manufacturer Kit Name Method Automation Lot No. ExpiryDate Single/Nested
10003 In house - TaqMan RT-PCR ABI Prism NA NA Single round40002 In house - TaqMan RT-PCR Rotorgene 11022015 Feb 16 Single round60003 In house - TaqMan RT-PCR Rotorgene NA NA Single round80069 In house - Gel electrophoresis - - NA80071 In house - Real-time Light Cycler 10035000 Sep 15 Single round
83000 Qiagen Quantitect ProbeRT-PCR kit
Fluorescent (FAMchannel) BioRad CFX 96 148030425 Jul 15 Single round
89082 In house - TaqMan RT-PCR LightCycler 030215 Feb 16 Single round
Flavivirus Detection Information for Survey MAF1:2015:1A, 1B, 1C, 1D, 1E & 1F (continued)
Part
Kunjin Virus - Detection
Manufacturer Kit Name Method Automation Lot No. ExpiryDate Single/Nested
40002 In house - TaqMan RT-PCR Rotorgene 29012015 Jan 16 Single round60003 In house - TaqMan RT-PCR Rotorgene NA NA Single round
Part
Murray Valley Encephalitis Virus - Detection
Manufacturer Kit Name Method Automation Lot No. ExpiryDate Single/Nested
40002 In house - TaqMan RT-PCR Rotorgene 07012015 Jan 16 Single round60003 In house - TaqMan RT-PCR Rotorgene NA NA Single round80069 In house - Gel electrophoresis - NA NA
Part
West Nile Virus - Detection
Manufacturer Kit Name Method Automation Lot No. ExpiryDate Single/Nested
40002 In house - TaqMan RT-PCR Rotorgene 05022015 Feb 16 Single round60003 In house - TaqMan RT-PCR Rotorgene NA NA Single round80069 In house - Gel electrophoresis - NA NA89082 In house - TaqMan RT-PCR LightCycler 300414 Apr 15 Single round
89112 UV Products UVTransilluminator
GeneSnapfrom SynGene IN HOUSE Oct 15 Single round
Part
Yellow Fever Virus - Detection
Manufacturer Kit Name Method Automation Lot No. ExpiryDate Single/Nested
40002 In house - TaqMan RT-PCR Rotorgene 10122014 Dec 15 Single round80069 In house - Gel electrophoresis - NA NA80071 In house - Real-time PCR Light Cycler 10035000 Sep 15 Single round89082 In house - TaqMan RT-PCR LightCycler 110315 Mar 16 Single round