To evaluate the cross-reactivity of the LFIA test strip, BBrMV and two other common banana infecting viruses (BBTV, the banana bunchy top virus; CMV, the cucumber mosaic virus) from strongly positive field samples, were tested with the test strip. BBrMV-infected leaveswere used as a positive control, and virus extraction buffer (PBS-T) was used as a negative control. The sensitivity of LFIA strips was evaluated by testing a series of different concentration of the purified BBrMV protein solution 0, 5, 10, 20, 40, 80, 160, 320 and 640 ng per milli litre. Each dilution was then added to the LFIA test strip, and the sensitivity was determined by finding the endpoint dilution. A series of crude extract dilutions from BBrMV-infected leaves were tested using the LFIA, crude extract from BBrMV-uninfected leaves was used as the negative control.
The LFIA was validated using 114 samples of commercial banana cultivars suspected to have BBrMV infection and collected from different orchards located in six states of India and also tissue culture samples received under the NCS-TCP being operative in India. In addition, the same samples were also subjected to ELISA assay to provide a comparison. Based on the results obtained, the samples were classified as true positive (TP), true negative (TN), false positive (FP), or false negative (FN).
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The diagnostic sensitivity and specificity of the LFIA were calculated using the Cohen’s kappa index 6.