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Fig. 6. Phylogenetic relationship of the sequence of the Polerovirus isolates from KZN.
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The increasing number of reported viruses infecting cucurbits in the world warranted an updated status of these viruses in KZN. Previous studies of this nature were reported by Cradock et al. (2001) who screened for viruses causing mosaic diseases by means of serological assays. In this study, serological and molecular based techniques were used to identify viruses causing mosaic and yellowing symptoms on cucurbits. Cradock et al. (2001) identified WMV in the previous survey, however, the virus was not identified during this study. Moreover, SqMV was not detected in both surveys. CMV, MWMV and ZYMV remained the mosaic viruses of cucurbits identified from both surveys. Results obtained during this survey showed that MWMV was the most prevalent virus infecting cucurbits in KZN contrary to previous reports by Cradock et al. (2001) in which ZYMV was the most prevalent. MWMV was detected in a variety of hosts including baby marrow, pattypan and pumpkin compared to ZYMV which was only detected in butternut.
Virus, vector, and host are the components that form the tripartite pathosystem of which each component interacts with the environment (Jones, 2014). All viruses identified in this study are vector transmitted. Unlike BPYV that is transmitted only by the greenhouse whitefly T. vaporariorum (Wisler et al., 1998), CMV; MWMV; ZYMV and PABYV are transmitted with varying efficiency by numerous species in various genera in the family Aphididae (Castle et al., 1992; Garzo et al., 2004; Lecoq and Desbiez, 2012). There is currently no information on the aphid species occurring in KZN and their efficiency in transmitting the viruses identified in this study. On the other hand, the increase in the number of cucurbit seed firms over the last decade has provided farmers with a comprehensive selection of hybrids and cultivars.
Commercial growers actively pursue the use of tolerant varieties in order to mitigate virus damages on crops. Subsistence farmer selection of cultivars and varieties are primarily based on their affordability. The variation observed in the frequencies of virus incidence in KZN in regards to the factors involved in virus epidemics can only be speculative and could have been the result of variability in host and vector populations.
Phylogenetic analysis is of crucial importance in the molecular studies of plant viruses.
The coat protein sequences of the CMV isolates detected in this study clustered within the Subgroup IA and are therefore different from the Wemmershoek isolate of CMV detected earlier in South Africa which belongs to the Subgroup II (Bashir et al., 2006). The coat protein sequence of the ZYMV isolates used in this study clustered within Group A of the three major groups that comprise the ZYMV lineage. The group A lineage of ZYMV, also referred to as the worldwide isolates, is further divided into four Subgroups; AI, AII, AIII and AIV (Coutts et al., 2011). Based on the coat protein sequences, the ZYMV isolates clustered within Subgroups AI and AII which consists of isolates from Asia, Europe, Oceania and North
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America. Using Nib C-ter and the N-terminal portion of the coat protein coding region of the genome for analysis, the six MWMV isolates clustered within the Southern African lineage of MWMV which is consistent with previous studies conducted by Yakoubi et al. (2008) which supports the limited long distance dispersal hypothesis of MWMV.
Yellowing symptoms of cucurbits in KZN are the result of infection by the Crinivirus BPYV and a Polerovirus species. In this survey, BPYV was more frequently identified in samples from tunnels than in open fields. Phylogenetic analysis performed using all the available BPYV sequences in the GenBank database confirmed the identity of the sequences as BPYV.
Prior to the development of degenerate primers that target the species of the genus Polerovirus, CABYV was the main Polerovirus reported to infect cucurbits globally. The amplicon of the expected size obtained with the Polerovirus degenerate primers pointed to the presence of a Polerovirus different from CABYV as no amplification was obtained with the CABYV specific primers. The assumption was confirmed following the analysis of the respective sequence data. The sequences of these isolates shared 72 % similarity with CABYV sequences used in this study. The species criterion in the family Luteoviridae according to the International Committee on Taxonomy of Viruses (ICTV) is a difference in amino acid sequences of any gene product greater than 10 % (d’Arcy and Domier; 2012). PABYV was the only species among Polerovirus infecting cucurbits to share amino acid sequence similarity greater than 90 % for the different gene products analysed in this study. Therefore, the Polerovirus detected in this study should be considered as isolates of PABYV. This conclusion was supported by the phylogenetic results in which all KZN Polerovirus isolates clustered within the PABYV group. PABYV was the provisional name given to recent isolates of Polerovirus infecting cucurbit detected in Mali in West Africa (Knierim et al., 2014). Although no complete genome sequence of PABYV is currently available in the GenBank database, the high prevalence and broad host infectivity in cucurbits by PABYV recorded in this study may be an indication of its widespread distribution in the African continent.
The frequent identification of various cucurbit-infecting viruses across KZN is an indication of the lack of resistance in the locally cultivated varieties. This threat to cucurbit production requires the development of better virus control strategies. MWMV and ZYMV which were prevalent viruses of cucurbits detected in this study and are known to cause up to 100 % yield and up to 95 % fruit marketability losses when infections occur early in the season (Lecoq and Desbiez, 2012). These losses are mostly incurred by subsistence farmers who do not possess the knowledge or resources to effectively manage the diseases caused by these
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viruses. The use of virus resistant cultivars can be effectively used in combination with existing methods to reduce the impact of these diseases (Gal-On, 2007). A study by Colvin et al. (2012), emphasizes the enormous socio-economic benefits that arise for resource-poor farmers from growing virus resistant varieties.