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Improvement of widespectrum detection of different potyviruses infecting passion fruit by the mixture of two kinds of polyclonal antibodies against east Asian passiflora virus
(A) Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and (B, C) western blotting assays of the bacterial-expressed coat protein (CP) of East Asian Passifl ora virus (EAPV)-AO and EAPV-IB. (A) The bacterial-expressed fusion proteins of EAPV-AO (37 kDa) and EAPV-IB (33 kDa) (lane eBEP) were observed by SDS-PAGE. (B) The monoclonal antibody to <i>Potyvirus</i> purchased from Agdia, and (C) the polyclonal antibodies to EAPV-AO CP and EAPV-IB CP prepared in this study were used to react with the bacterial-expressed viral CPs. Lane M: protein markers, lane pET: IPTG-induced bacterial cell lysate containing the empty vector pET28a(+), lane BEP:IPTG-induced bacterial cell lysate containing pET28a(+) with viral CP insert, and lane eBEP: the purified bacterial-expressed CPs of EAPV-AO and EAPV-IB, respectively.
(A) Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and (B, C) western blotting assays of the bacterial-expressed coat protein (CP) of East Asian Passifl ora virus (EAPV)-AO and EAPV-IB. (A) The bacterial-expressed fusion proteins of EAPV-AO (37 kDa) and EAPV-IB (33 kDa) (lane eBEP) were observed by SDS-PAGE. (B) The monoclonal antibody to Potyvirus purchased from Agdia, and (C) the polyclonal antibodies to EAPV-AO CP and EAPV-IB CP prepared in this study were used to react with the bacterial-expressed viral CPs. Lane M: protein markers, lane pET: IPTG-induced bacterial cell lysate containing the empty vector pET28a(+), lane BEP:IPTG-induced bacterial cell lysate containing pET28a(+) with viral CP insert, and lane eBEP: the purified bacterial-expressed CPs of EAPV-AO and EAPV-IB, respectively.

Author:Chin-Chih Chen*, Fen-Lang Chiang, Ying-Huey Cheng, and Jia-Yi Liao

Abstract:

When the virus-infected passion fruits showed woodiness symptoms on fruits, the fruits were deformed and smaller resulting in a low juice rate, poor flavor and quality, which could cause a great impact on yield. Two East Asian Passiflora viruses (EAPV-AO and EAPV-IB) and Telosma mosaic virus (TeMV) are the three main potyviruses infecting passion fruits in Taiwan. In this study, the polyclonal antibodies against the coat proteins (CP) of EAPV-AO and EAPV-IB were respectively prepared from immunizing rabbits with the bacterial expressed viral CPs, revealing that both antibodies have a wide-spectrum response with 19 tested potyviruses. In indirect enzyme-linked immunosorbent assay (indirect ELISA), the equivalent mixture of polyclonal antibodies against the CPs of EAPV-AO and EAPV-IB, denoted Anti-EAPVmix, could also be used to detect passionfruit isolates of TeMV from Taiwan and Thailand, and to improve the detection rate of EAPV-AO, EAPV-IB, and TeMV. The distribution of viruses on plant branches was detected by Anti-EAPVmix. The virus was detected at a high rate from the upper part near new leaves, which is recommended for sampling. Our result showed that the mixture of EAPV polyclonal antibodies can be used to detect not only the three major passiflora viruses in Taiwan but also other potyviruses to save the detection time and costs. The detection method can strengthen the field survey of passiflora viruses and the internal control of healthy seedlings. 

Key words:Passiflora potyviruses, Polyclonal antibody, Antibody mixture, Wide-spectrum detection

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Journal of
Taiwan Agricultural Research
Vol. 72, No.3 (2023) ISSN: 2790-086X Vol. 72, No.3 (2023)
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