Bottle gourd IC-0262269, a super-susceptible genotype to tomato leaf curl Palampur virus.

While conducting field trial of 82 genotypes of bottle gourd at Delhi during 2020-2021, a particular genotype, IC-0262269 was found to be affected by chlorotic curly stunt disease (CCSD). The affected plants were severely stunted and bearing very small chlorotic and crinkle leaves. The disease incidence in the said genotype was as high as 80% among different replicated trial blocks. The application of PCR using a generic primers specific to begomoviruses, as well as species-specific PCR diagnostics to six tomato-infecting begomoviruses: tomato leaf curl New Delhi virus (ToLCNDV), tomato leaf curl Palampur virus (ToLCPalV), tomato leaf curl Joydebpur virus (ToLCJoV), tomato leaf curl Gujrat virus (ToLCGuV), tomato leaf curl Bangalore virus (ToLCBV), and chilli leaf curl virus (ChiLCV) showed that, only ToLCPalV could be detected in the genotype IC-0262269. Following, rolling circle amplification, cloning and sequencing of full-length DNA-A and DNA-B genome of an isolate BoG1-ND from the genotype IC-0262269 revealed association of ToLCPalV with the disease. The successful agro-infection of the cloned genome of BoG1-ND (DNA-A and DNA-B) in the plants of Nicotiana benthamiana and bottle gourd demonstrated that ToLCPalV is the causal begomovirus of CCSD. The study provides the first evidence of the natural occurrence of ToLCPalV in bottle gourd crop and also showed that the bottle gourd genotype IC-0262269 is super-susceptible to ToLCPalV. Supplementary Information: The online version contains supplementary material available at 10.1007/s13205-023-03838-y.

Geographical distribution of tomato-infecting begomoviruses in major cucurbits in India: a diagnostic analysis using begomovirus species specific PCR.

Cucurbits are an essential summer-season vegetable crops, but they are highly vulnerable from a range of abiotic and biotic factors. One of the significant biotic factors posing a growing menace to the production of major cucurbits in India is the emergence of tomato-infecting begomoviruses. In this study, we utilized PCR-based species-specific primers, developed earlier in our laboratory for the detection of begomoviruses infecting tomato and chilli plants, to identify begomoviruses in cucurbits across various regions of India. Leaf samples from major cucurbits were collected from different regions of Haryana, Delhi, Uttar Pradesh, Chhattisgarh, Maharashtra, Telangana and Karnataka, during the year 2020-2021. Total nucleic acid (TNA) was extracted from the samples and subjected to PCR using a generic primer specific to begomoviruses. The samples that exhibited positive amplification were further tested using six different species-specific primers targeting specific begomovirus species, namely Tomato leaf curl New Delhi virus (ToLCNDV), Tomato leaf curl Palampur virus (ToLCPalV), Tomato leaf curl Bangalore virus (ToLCBV), Tomato leaf curl Joydebpur virus (ToLCJoV), Tomato leaf curl Gujarat virus (ToLCGuV), and Chilli leaf curl virus (ChiLCV). The PCR analysis revealed that among the 551 plant samples tested, a total of 124 samples exhibited positive amplification using the universal begomovirus PCR. Specifically, 47 samples tested positive for ToLCNDV, 73 samples were positive for ToLCPalV and only one sample showed positive amplification for ChiLCV. However, none of the samples tested positive for ToLCJoV, ToLCGuV and ToLCBV. These findings from our study indicate the prevalence of ToLCNDV and ToLCPalV in major cucurbits across India. Furthermore, the study highlights the varied distribution of begomoviruses in major cucurbits between northern and southern regions of India.

Delivery of progeny virus from the infectious clone of cucumber green mottle mosaic virus and quantification of the viral load in different host plants.

Cucumber green mottle mosaic virus (CGMMV, genus Tobamovirus) is a widely occurring tobamovirus in cucurbits. The genome of CGMMV has been used previously for the expression of foreign genes in the plant. High throughput delivery and high viral titer are important requirements of foreign protein expression in plant through virus genome-based vector, in this study, Agrobacterium containing infectious construct of CGMMV was infiltrated through syringe, vacuum and high-speed spray to N. benthamiana, cucumber and bottle gourd leaves. The success rate of systemic infection of CGMMV agro-construct through all three methods was higher (80-100%) in N. benthamiana compared to the cucurbits (40-73.3%). To determine the high-throughput delivery of CGMMV in the plant system, four delivery methods viz. rubbing, syringe infiltration, vacuum infiltration and high-speed spray using the progeny virus derived through CGMMV agro-construct were compared in the three different plant species. Based on the rate of systemic infection and time required to perform delivery by different methods, vacuum infiltration was found most efficient for the high-throughput delivery of CGMMV. The quantification of CGMMV through qPCR revealed that CGMMV load varied considerably in leaf and fruit tissues depending with the time of infection. Immediately after expression of symptoms, a high load of CGMMV (~ 1 µg/100 mg of tissues) was noticed in young leaves of N. benthamiana and cucumber. In bottle gourd leaves, the CGMMV load was far low compared to N. benthamiana and cucumber plants. In the fruit tissues of cucumber and bottle gourd higher virus load was observed in mature fruit but not in immature fruit. The findings of the present study will serve as an important base line information to produce foreign protein through CGMMV genome-vector. Supplementary Information: The online version contains supplementary material available at 10.1007/s13205-023-03630-y.