Establishment of a reverse genetics system for an epidemic strain of porcine rotavirus JXAY01 type G5P[23]I12

Figures

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  Figure 1. PAGE of viral genomic dsRNA extracted from SA11, native JXAY01 and monoreassortants. The numbers on the side indicate the order of the genomic dsRNA segments of SA11 and JXAY01. The red markers indicate recombinant segments.

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  Figure 2. Plaque size comparison. (A) Plaques were formed on MA104 monolayers and identified at 4 days post-infection using Neutral Red staining. Displayed are representative images of viral plaques. (B) The dimensions of a minimum of 30 plaques, picked randomly from two separate plaque tests, were quantified using the GraphPad Prism software and expressed as area in relative units. The mean values and standard deviation are displayed. The statistical significance was assessed using a Student’s t-test. The asterisks are used to highlight significant differences (*p ≤ 0.05; **p ≤ 0.01; ***p ≤ 0.001).

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  Figure 3. Growth kinetics of reassortant viruses. (A,B) Confluent MA104 cells cultured in 6 wells were exposed to an infection with a multiplicity of infection (MOI) of 0.1. At the specified time intervals, samples were collected and the number of plaque-forming units (PFUs) was calculated using plaque assay. (C) The illustrated data are the means obtained from three separate experiments, and the error bars indicate the standard deviation. The viral titers at 24 h post-infection were used for statistical analysis, and the results showed that the viral titers of the VP2 and VP4 single-segment recombinant strains were significantly reduced (*p ≤ 0.05; **p ≤ 0.01; ***p ≤ 0.001).

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  Figure 4. Establishment of a reverse genetics system for Porcine rotavirus strain JXAY01. (A) Immunofluorescence detection and CPE observation of the whole plasmid rescue strain rJXAY01. (B) Electrophoretic patterns of dsRNA from SA11, native JXAY01 and rJXAY01. (C) Plaque morphologies of native JXAY01 and rJXAY01 on MA104 cells were detected by neutral red staining and sizes. (D) The plaque size were measured using ImageJ software. Statistical analysis shows no significant difference in the plaque sizes between the two strains. (E) Growth kinetics of native JXAY01 and recombinant strain rJXAY01 in MA104 cells. Statistical analysis shows no significant difference in the viral titers between the two strains (*p ≤ 0.05; **p ≤ 0.01; ***p ≤ 0.001).