Family Cucurbitaceae is primarily found in the warmer regions of the world. It is the major family for economically important species, particularly edible fruits. In Pakistan cucurbits occupies an area of 28,600 ha with a very low production in Khyber Pukhtunkhwa due to many biotic and abiotic stresses. The reason is also the lack of growers’ awareness about the diseases and the cultural practices adopted to provide favorable environment for development of epidemics. Viral diseases such as Cucumber mosaic virus (CMV) cause losses as high as 100%. Various control strategies are being used to control CMV. The aim of the current study was to screen out different verities and to find the most resistant one against CMV. CMV isolate was collected from farmer’s field at the site of TaruJaba during a survey of cucurbit crops. The identity of the virus was confirmed through DAS-ELISA using diagnostic kit (ADGEN, UK). Seventeen cucumber germplasm seeds were sown in earthen pots in which fourteen were germinated and exhibited characteristics symptoms of the virus while none of them showed resistance against CMV. Symptoms’ expression was delayed in summer green and local green till 12 days post inoculation. While in khyber, Diamond, VEGAF1 and Yousaf, symptoms started to appear soon after inoculation categorizing them as highly susceptible. No resistance is found in available commercial germplasm, so more germplasm from different area of Pakistan should be tested for resistance against CMV. If no resistance is found locally imported, germplasm can be evaluated for a source of resistance against the prevalent isolates of CMV.
Cucurbitaceae or cucurbit family (also commonly referred to as the cucumber, gourd, melon, or pumpkin family) is primarily found in the warmer regions of the world. It is the major family for economically important species, particularly those edible fruits. It includes about 118 genera and 825 species [
Globally, cucurbits are grown on an area of 8.5 million hectares with production of 17.9 million tones [
The reason for such low yield of these crops is the lack of growers’ awareness about the diseases and the cultural practices adopted to provide favourable environment for development of epidemics. The cultural practices adapted by the growers, lack of awareness of growers about viral diseases and their control, high population of weeds, and type of cropping pattern that includes the host crops of the prevalent viruses are some of the major contributing factors that result in a high incidence of viral diseases.
Cucurbits viral diseases are a problem worldwide that causes losses as high as 100% [
Cucumber mosaic virus (CMV) is one of the most important pathogen, which belongs to genus Cucumovirus in the family Bromoviridae. It is an economically important virus of agricultural crops and has the broadest host range known for any plant virus with approximately 1000 susceptible plant species [
Various control strategies are used for controlling these viruses and there is a dire need to evaluate different control strategies under each set of environmental condition and with different virus host combination to formulate an effective integrated control approach. Incidence of cucurbit viruses has been reported from Pakistan showing quite prevalence of these viruses in different cucurbit growing areas [
The study was conducted at the Department of Plant Pathology, The University of Agricultural Peshawar Pakistan, with the objective to screen different cucumber cultivars against (CMV).
CMV isolate was collected from farmer’s field at TaruJaba during a survey of cucurbit crops. The identity of the virus was confirmed though DAS-ELISA using diagnostic kit (ADGEN, UK).
Seventeen germplasm of cucumber collected from local market and Agricultural Research Institute (ARI) Tarnab and were used for screening purpose. Twenty seeds of each cultivar were sown in earthen pots with two seeds in each pot. Percent germination of each cultivar was observed after one week.
After germination, number of plants in each pot was reduced to one. One plant, out of ten, for each cultivar was used as control where inoculation was done using phosphate buffer only without crushing plant sample, while in the remaining 9 plants inoculation was done using virus infected sap.
Mechanical inoculation was done on carborandum dusted cotyledonary leaves at cotyledonary leaf stage with the help of a cotton swab. The inoculum was prepared by crushing Ig of infected leaf in 10 ml phosphate buffer (2.4 gm of KH2PO4 and 5.4 gm of K2HPO4 in one liter water). Plants were washed with tap water after inoculation and kept in dark for overnight for symptoms development.
After inoculation the plants were kept in screen house and were regularly monitored (twice a week) till four weeks post inoculation. The symptoms on the host plants were assessed using the following disease rating scale (1 - 5) as by [
1. = Highly resistant (no symptoms; 0% - 10% infection);
2. = Resistant (vein clearing after some time; 11% - 20%);
3. = Moderately resistant (vein clearing and mild mottle; 21% - 30%);
4. = Moderately susceptible (mild mosaic on few leaves; 31% - 40%);
5. = Susceptible (mosaic, wrinkling, mottling; >60%).
According to the disease rating scale the cucumber plants were divided into different categories.
Finally all the samples were assayed by double antibody sandwich ELISA (DAS-ELISA) using commercial kits from ADGEN, UK to confirm the identity of the virus, as well as to check the possibility of any latent infection.
The following feature of the procedure, as recommended by the manufacturer kit, was used for the assay:
100 µl of the antibodies were coated on the ELISA plates with the help of coating buffer and the plates were incubated at 37˚C for 4 hours. Following incubation, the plates were washed three times with the help of washing buffer;
Samples were homogenized in the extraction buffer and 100/l of the sap was applied to the wells. The plates were incubated at 4˚C overnight followed by washing, thrice with the help of washing buffer;
100 µl of conjugated antibodies were applied to the wells and plates were incubated at 37˚C for 1 hr;
The plates were incubated at about 25˚C (or room, temperature) in dark;
Data were recorded after 20, 30 and 60 minutes.
Seventeen cucumber germplasm seeds were sown in earthen pots in which fourteen germinated and exhibited characteristics symptoms of the virus as described in
At seven days of post inoculation, symptoms of CMV were observed in five germplasm that is in Khyber, Yousaf, VEGAF1, Waleed, and Diamond. Among the other germplasm symptoms started appearing after 12 days of post inoculation that was in (Betialpha, Punjab, Point pett-76, Baran, Surnmer green, Local green, Kurnar, Diamond 2 Fl, Sahill). No symptoms were observed on plants used as a control (inoculated with buffer only).
Symptoms expression was delayed in summer green and Local green till twelve days post inoculation while in Khyber, Diamond, VEGAFl, Yousaf, symptoms start appearing soon after inoculation, categorized them highly susceptible. Germplasm Beitalpha, Punjab, and Barandidnt show any symptom during the first two observations
S. No. | Germplasm name | 7 days | 12 days | 18 days | 24 days | Disease rating | Symptoms appearance |
---|---|---|---|---|---|---|---|
1. | Khyber | 3/9 | 6/9 | 8/9 | 9/9 | 5 | Sm & M |
2. | Beitalpha | 0/9 | 0/9 | 8/9 | 9/9 | 4 | Mm |
3. | Punjab | 0/9 | 0/9 | 8/9 | 9/9 | 4 | Mm |
4. | Point pett-76 | 0/9 | 3/9 | 6/9 | 9/9 | 4 | Mm |
5. | Baran | 0/9 | 0/9 | 8/9 | 9/9 | 4 | Mm |
6. | Summer green | 0/9 | 3/9 | 6/9 | 9/9 | 4 | Mm |
7. | Localgreen | 0/9 | 3/9 | 6/9 | 9/9 | 4 | Mm |
8. | Kumar | 0/9 | 3/9 | 6/9 | 9/9 | 4 | Mm |
9. | Yousaf | 3/9 | 6/9 | 8/9 | 9/9 | 5 | Sm & M |
10. | VEGAF1 | 3/9 | 6/9 | 8/9 | 9/9 | 5 | Sm & M |
11. | Diomond 2F 1 | 0/9 | 3/9 | 6/9 | 9/9 | 4 | Mm |
12. | Sahill | 0/9 | 3/9 | 6/9 | 9/9 | 4 | Mm |
13. | Iqbal | NG | NG | ||||
14. | Waleed | 3/9 | 6/9 | 8/9 | 9/9 | 5 | Sm & M |
15. | Market more | NG | NG | ||||
16. | Holandgrcen | NG | NG | ||||
17. | Diamond | 6/9 | 8/9 | 9/9 | 9/9 | 5 | Sm & M |
18 | Control | 0/9 | 0/9 | 0/9 | 0/9 | 1 | NS |
NG = No germination; NS = No symptoms; Sm & M = Severe mosaic & mottling; Mm = Mild mosaic.
and during the third observation 88%, (8/9), and almost all plants were infected during the 4th observation reaching to 100%. Among all of these germplasm, Diamond showed maximum susceptibilityas the symptoms appeared soon after inoculation and all its 9 plants became infected. Thus showing high level of susceptibility compared to other susceptible cultivar.
Cucumber mosaic virus (CMV) is undoubtedly an important, serious and destructive disease in all growing countries of the world including Pakistan. In view of ubiquitous nature of CMV disease, 17 cucumber germplasm seeds were screened for resistance to CMV. These seeds were selected by the farmer’s on locally based criteria over many years, which are strongly adapted for their local source conditions. The genotypes were classified into five reaction groups based upon % infected plants and ELISA test. These were: highly resistant, resistant, moderately resistant, moderately susceptible and susceptible. It is clear from the data that most of cucumber germplasm are moderately susceptible and if we observe
In Pakistan no systematic work has been done to determine the yield losses due to CMV diseases on crops. CMV is one of the major viral diseases recorded in world elsewhere including Pakistan [
S. No. | Scale/reaction | Cucumber |
---|---|---|
1. | Highly susceptible | Khyber, Yousaf, VEGAF1, Diamond Waleed |
2. | Moderately susceptible | Punjab, Beitalpha, pointpett-76, Baran, Local green, Summer green, Diomond 2F l, Sahill |
3. | Moderately resistant | ―― |
4. | Resistant | ―― |
5. | Highly resistant | ―― |
for a long time and transportation of infested materials by human activities from one place to another, whereas for long area dispersal of the inocula is due to viruliferous aphids to a new cultivating area. Virus infected weeds within crop are serious threat because of the primary source of inoculum to the healthy ones [
Recent reports showed that aphid population has increased tremendously in the recent past due to favorable environmental conditions [
It is concluded from the present findings that no resistance is found in available commercial germplasm, so more germplasm from different area of Pakistan should be tested for resistance against CMV. If no resistance is found locally imported, germplasm could be evaluated for source of resistance against the prevalent isolates of CMV. One main problem with germplasm evaluation is that some genotypes found resistance at one location turn out to be susceptible at another place [