Evaluation of Antagonistic of the some Fungal isolates on Golden Potato Cyst Nematode (Globoderarostochiensis) in vitro and Greenhouse Conditions in Hamedan Province

Potato (Solanumtuberosum) is one of the most important crops used as a source of human food. Iran is the third-largest producer of potato in Asia, where the production rate in 2015 was estimated to be about 5 million tonnes. Potato producers inHamedan province produce 21.3% oftotal potato harvestedinIran. Golden potato cyst nematode, Globoderarostochiensis is the most destructive potato pathogen. As the chitin is a dominant composition in middle layer of the eggshell, using the chitinases produced as chitin-degrading enzymes in a wide range of fungiis a good strategy for biological control of the golden potato cyst nematode. We assessedthe ability of various antagonistic fungi to control Globoderarostochiensisunder in vitro and greenhouse conditions.
Thirty four fungal isolates obtained from infected eggs of the potato cyst nematode, Globoderarostochiensisin potato fieldsofHamedan were evaluated in two chitin-agar and water-agar mediums under in vitro and greenhouse conditions.The ability of thechitinase enzyme production was assessedin chitin-agar medium with colloidal chitin as substrate, so the chitin was used as exclusive source of carbon.Colloidal chitin was prepared based onthe procedure of Seyedasliet al. (2004) with 10 g of powder chitin from practical-grade crab shell chitin (Sigma) in 100 ml of 85% H3PO4. Water was added to the above mixtureand was filtered with cheese cloth. To completely remove acid, water addition and filtrationrepeated for several times. The produced unguent materialwas dried and powdered and then used as carbon source in the medium. 0.5 percent of colloidal chitin was added to the medium. Afterwards, a 5 mm disk from the edges of 5days old was placed in the center of Petridish and all of them were kept for 5 days at 25º C. Chitinase detection medium (chitin-agar) was directly supplemented with colloidal chitin (5 g/l) and bromocresol purple (0.15 g/l). The ability of antagonistic activity of the fungi on the cyst nematode was testedin water-agar medium through assessing the interaction between fungi and cysts. The numbers of healthy and parasitized (dead) larvae and eggs were calculated after two weeks. The ability of the antagonistic fungi under greenhouse conditions was also analyzed. To provide fungal inoculum, 20g of soaked wheat seed were cast in nylon with autoclave capability. 2 ml distilled water were added per gram of cast seed and they were autoclaved threetimesduring 24 hours. Four fungi disk with 5 mm diameter from selected isolates were then added to allnylonswiththreerepetitions and werekept in 25ºC and dark conditions. To colonizeall of the seeds and avoid hangingthem, the seeds in nylons were mixed within 48 hours interval. After three weeks all of the seeds were infected with fungal isolates. The ability of the antagonistic fungi under greenhouse conditions was studiedby adding fungal inoculum and 100 cysts to each pot and performance evaluation of potato traits in pot after 90 days.
ANOVA (Analysis of variance) data analysis was conducted using of the SAS software version 9.0 in completely randomized design (CRD) with three replicates under all conditions.
The ANOVA results ofqualitative evaluation for chitinase activity in Petri dish containing chitin-agar showed significant difference among isolates at1% level of significance. Furthermore, there was apositiveassociationbetween diameter and chitanase activity. Isolates 153(P. lilacinus) and 6 (C. parapsilosis) had the maximum and minimumdiameter, respectively.
A significant difference was foundamong isolatesin the 0.1% level of significance. Isolate 152 (L. muscarium) had the greatestantagonistic ability and 62 (F. solani)was the weakest antagonist isolate.
Mean comparisons of measured performance traits of potato in pot showed in all traits thatisolates 154 (T. atroviridae)and 151 (B. bassiana)were the best antagonist isolates under greenhouse conditions. Isolates56 (F. equiseti)and 12 (F. oxysporum)with the lowest measured values in all functional traits were alsothe weakest antagonist fungi.
The results illustrated a strong correlation between antagonistic ability of fungal isolates under in vitro and greenhouse conditions.Therefore, fungal isolates can effectively reduce the nematode damages. Finally, isolates 151(B. bassiana), 152(L. muscarium), 153 (P. lilacinus) and 154(T. atroviridae) were selected as the strongest antagonistic fungi in controlling this nematode.