DOI: 10.5061/dryad.547d7wmmf
title: Data from: Development and proof of concept evaluation for a low-resource compatible Chikungunya virus diagnostic

description: Chikungunya virus (CHIKV) is a positive sense RNA Alphavirus that
 continues to pose major public health threats throughout the world. CHIKV
 is primarily transmitted via the Aedes genus mosquito; however, it has
 also exhibited transmission routes via blood transfusion and vertical
 transmission (mother to child). With only one approved vaccine thus far
 and no approved medicines, early detection is crucial in mitigating CHIKV
 outbreaks. Here, we designed and evaluated a sensitive and specific CHIKV
 diagnostic using reverse transcription-recombinase aided amplification
 (RT-RAA) coupled lateral flow strip detection (LFD) targeting a highly
 conserved region of the CHIKV E1 gene. Our results demonstrate that using
 our simple sample preparation reagent (TNA-Cifer-E) can inactivate live
 CHIKV in two minutes at room temperature, whilst also sustaining viable
 viral RNA. Our specificity analysis demonstrates that the Iso-CHIKV-Dx
 does not detect any closely related Alphaviruses nor any of the common
 co-circulating Flaviviruses. Proof-of-concept evaluation using urine
 spiked with CHIKV exhibited that in CHIKV-infected urine samples, this
 Iso-CHIKV-Dx can detect as low as 570 copies/μL of CHIKV RNA in 30 minutes
 under isothermal conditions. Contrary to conventional RT-qPCR, our
 Iso-CHIKV-Dx does not require expensive machinery, advanced
 instrumentation or extensively trained personnel. Further performance
 comparisons also show that this Iso-CHIKV-Dx is four times faster than
 conventional RNA isolation and RT-qPCR. As such, pre-clinical evaluation
 demonstrates that this Iso-CHIKV-Dx has the potential to act as a robust,
 point of care CHIKV diagnostic that could prove to be highly beneficial in
 place of, or in the absence of RT-qPCR.
description: # Data from: Development and proof of concept evaluation for a
 low-resource compatible Chikungunya virus diagnostic Dataset DOI:
 [10.5061/dryad.547d7wmmf](10.5061/dryad.547d7wmmf) ## Description of the
 data and file structure These data were generated to develop, optimize,
 and validate a proof-of-concept rapid, isothermal CHIKV test using reverse
 transcription recombinase aided amplification coupled with lateral flow
 detection (RT-RAA/LFD) and a rapid sample preparation for low-resource
 virus detection in Urine. ### Files and variables #### File:
 Raw_Data_for_Figure_1.fasta **Description:** Raw, aligned sequence data of
 667 unique CHIKV E1 gene sequences along with a consensus sequence. ####
 File: Raw_Data_for_Figure_2_.csv **Description:** Limit of detection
 calculation of CHIKV RNA copies per micro litre using RT-RAA-LFD. #####
 Variables: Viral RNA copies/uL, Pixel density (black and white normalised
 values) * Raw image file: Uploaded separately as a PNG file. Depicting raw
 scanned image of lateral flow strips by which statistical analysis was
 based upon.  * Mean test band: Average of the pixel density of the test
 band calculated via image J  * Mean White Space: Average of the pixel
 density of the test band calculated via image * White Space normalized: A
 normalised calculation of the white space of the lateral flow strip *
 Normalized test band: A normalised calculation of the test band of the
 lateral flow strip * Min Thr: The minimum detected value based on the
 normalisation calculations * Max Thr: The upper end detected value based
 on the normalisation calculations * black pixels in test band: Numerical
 value of the intensity of the black bands based on the measurements using
 Image J * logic test (refers to 3*stddev): A calculation based on the
 standard deviation multiplied by three * blank normalised average:
 Calculation of the average black normalised values from three replicates *
 blank normalised stdev: Calculation of the standard deviation of black
 normalised values from three replicates. * Blank normalized: Normalisation
 of blank space (cleanest part of the lateral flow strip) to account for
 any ink run off via lateral flow chromatography  * Cut off Subtracted: The
 cut off value calculation based on the minimum pixel density to result in
 a positive based on the normalised white space values. * Graph: Depicted
 within the figure. #### File: Raw_data_for_Figure_3.csv
 **Description:** Analysis of CHIKV RT-RAA-LFD specificity testing using
 familial and other co-circulating arboviruses. ##### Variables: Viral RNA
 copies/uL, Pixel density (black and white normalised values) * Raw image
 file: Uploaded separately as a PNG file. Depicting raw scanned image of
 lateral flow strips by which statistical analysis was based upon.  * Mean
 test band: Average of the pixel density of the test band calculated via
 image J  * Mean White Space: Average of the pixel density of the test band
 calculated via image * White Space normalized: A normalised calculation of
 the white space of the lateral flow strip * Normalized test band: A
 normalised calculation of the test band of the lateral flow strip * Min
 Thr: The minimum detected value based on the normalisation calculations *
 Max Thr: The upper end detected value based on the normalisation
 calculations * black pixels in test band: Numerical value of the intensity
 of the black bands based on the measurements using Image J * logic test
 (refers to 3*stddev): A calculation based on the standard deviation
 multiplied by three * blank normalised average: Calculation of the average
 black normalised values from three replicates * blank normalised stdev:
 Calculation of the standard deviation of black normalised values from
 three replicates. * Blank normalized: Normalisation of blank space
 (cleanest part of the lateral flow strip) to account for any ink run off
 via lateral flow chromatography  * Cut off Subtracted: The cut off value
 calculation based on the minimum pixel density to result in a positive
 based on the normalised white space values. * Graph: Depicted within the
 figure. #### File: Raw_Data_for_Figure_5.csv **Description:** Limit of
 detection calculation of CHIKV RNA copies per micro litre using RT-RAA-LFD
 in Urine samples. ##### Variables: Viral RNA copies/uL, Pixel density
 (black and white normalised values) * Raw image file: Uploaded separately
 as a PNG file. Depicting raw scanned image of lateral flow strips by which
 statistical analysis was based upon.  * Mean test band: Average of the
 pixel density of the test band calculated via image J  * Mean White
 Space: Average of the pixel density of the test band calculated via image
 * White Space normalized: A normalised calculation of the white space of
 the lateral flow strip * Normalized test band: A normalised calculation of
 the test band of the lateral flow strip * Min Thr: The minimum detected
 value based on the normalisation calculations * Max Thr: The upper end
 detected value based on the normalisation calculations * black pixels in
 test band: Numerical value of the intensity of the black bands based on
 the measurements using Image J * logic test (refers to 3*stddev): A
 calculation based on the standard deviation multiplied by three * blank
 normalised average: Calculation of the average black normalised values
 from three replicates * blank normalised stdev: Calculation of the
 standard deviation of black normalised values from three replicates. *
 Blank normalized: Normalisation of blank space (cleanest part of the
 lateral flow strip) to account for any ink run off via lateral flow
 chromatography  * Cut off Subtracted: The cut off value calculation based
 on the minimum pixel density to result in a positive based on the
 normalised white space values. * Graph: Depicted within the figure. ####
 File: Raw_Data_for_Figure_4_.csv **Description:** Sample processing
 analysis of CHIKV culture exposed to TNA-Cifer-reagent E at 1:1 and 1:2
 ratios for varying time frames. ##### Variables: FFU/mL, TCE concentration
 ratio, Time of exposure to TCE (mins) * 1:1 TCE: CHIKV:TCE as a 1:1 ratio
 for 0.5, 1, 2 ,5, 10 mins * 1:1 PBS: CHIKV:PBS as a 1:1 ratio for 0.5, 1,
 2 ,5, 10 mins (as the control) * 1:2 TCE: CHIKV:TCE as a 1:1 ratio for
 0.5, 1, 2 ,5, 10 mins * 1:2 PBS: CHIKV:PBS as a 1:1 ratio for 0.5, 1, 2
 ,5, 10 mins (as the control) #### File: Figure_2__CHIKV_RT-RAA-LFD_LOD.png
 **Description:** Raw image of scanned lateral flow strips. #### File:
 Figure_3__CHIKV_RT-RAA-LFD_Specificity.png **Description:** Raw image of
 scanned lateral flow strips. #### File:
 Figure_5__CHIKV-RT-RAA-LFD_Urine.png **Description:** Raw image of scanned
 lateral flow strips. #### File: Raw_Data_For_Figure5.csv
 **Description:** RT-qPCR analysis of processed CHIKV urine samples.  #####
 Variables: CT values, RNA copy numbers, FFU/ml equivalent ## Code/software
 Possible software needed: Bioedit, MEGA, Genious, SnapGene or any free
 software that reads .FASTA format.