Data from: Development and proof of concept evaluation for a low-resource compatible Chikungunya virus diagnostic

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. # 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.