Fig. 1.

(a) 5×5 sample pool: A total of 25 samples were included in 5 × 5 matrix pool. Forty µL from each sample was added in two pools i.e row and column. e.g. 40 µL VTM from sample no. 1 was added to pool no. C1 and R1; Sample no.6 was added in pool no. C1 and R2, and so on. One to three positives were introduced at random positions in an operator- blinded manner; (b) 10 × 10 sample pool: A total of 100 samples were included in 10 × 10 pool. Twenty or Forty µL from each VTM was added in two pools. e.g. 40 µL VTM from Sample no. 1 was added in pool no. C1 and R1; Sample no.11 was added in pool no. C1 and R2, and so on. 1–3 positives were introduced at random positions in an operator-blinded manner [C- column, R-Row, numerical- sample nos.].