Bio mechanisms changes of Downy mildew fungi Pseudoperonospora cubensis on Cucurbits

Journal of Research and Development Peer Reviewed International, Open Access Journal. ISSN : 2230-9578 | Website: https://jrdrvb.org Volume-17, Issue-9| September 2025 22 Bio mechanisms changes of Downy mildew fungi Pseudoperonospora cubensis on Cucurbits Dr. Vasantrao P. Pawar Dept. of Botany Arts & Science College Bhalod, Tal. Yawal, Dist. Jalgaon. (M.S.) [email protected] Manuscript ID: JRD -2025-170904 ISSN: 2230-9578 Volume 17 Issue 9| Pp. 22-25 September 2025 Submitted:15 Aug. 2025 Revised: 25 Aug . 2025 Accepted: 10 Sept. 2025 Published: 30 Sept. 2025 Abstract Impact of downy mildew on chlorophyll, nitrogen, protein and reducing sugar content of cucurbits was studied. Chlorophyll a, chlorophyll b pigment of the infected leaf was found to be decreased due to downy mildew. Loss in nitrogen, protein and reducing sugar contain due to infection of downy mildew were also observed. Downy mildew caused by Pseudoperonospora cubensis significantly affects the physiological and biochemical composition of cucurbits. The present study evaluated its impact on chlorophyll, nitrogen, protein, reducing sugars, phenols, and starch in infected leaves. Results revealed a marked reduction in chlorophyll a and b, nitrogen, protein, and reducing sugar contents across most cucurbit species, with the greatest protein loss recorded in Cucumis melon. Conversely, phenolic content increased consistently in infected plants, indicating a possible defense response. Interestingly, starch levels remained largely unaffected or showed a slight increase under infection. These findings highlight the biochemical alterations induced by P. cubensis, providing insight into host–pathogen interactions and contributing to strategies for disease management in cucurbits. KeywordsDowny mildew fungi, biochemical, common name, crop leaves Pseudoperonospora cubensis, cucurbits, biochemical changes, chlorophyll, phenols. Introduction Downy mildew of cucurbits is caused by Pseudoperonospora cubensis is obligate bio trophic member of Oomycete Babadoost ( 2016). This disease occurrence in wild and cultivated plants as a fruits and vegetables in all over the word Berkeley and Curtis .(1868) Cohen.(1980) It has worldwide distribution and probably occurs wherever cucurbits are grown except unirrigated, very dry climates and especially prevalent in area with a warm, humid climates Most wild and cultivated plants as a fruits and vegetables in all over the word Sashadri (1986) Reddy (2002).In literature cited very little information was availed regarding the host range of the pathogen biochemical changes occur due to the pathogen and ecofriendly management of the disease. Materials and Methods Chlorophyll contentChlorophyll content from healthy and infected plant leaves were estimated by the method suggested by Aronon [1949],for this 2 gram of plant leaves were crushed in Mortar and pestle in 80% chilled action containing 4 ml of liquor ammonia per liter in dark code room. A pinch of Magnesium carbonate [MgCo3] was added during crushed. The extracts were filtrated trough Buchnners funnel using Whitman filter paper No.1 and volume of the filtrated was made up to 100 ml 80% with acetone 5 ml.of the extract was diluted up to 50 ml. with acetone in volumetric flask. In ordered to avoid destruction of chlorophyll by light, the flasks containing chlorophyll extracts were covered with black paper and stored at low temperature. Quick Response Code: Website: https://jrdrvb.org/ DOI: Creative Commons (CC BY-NC-SA 4.0) This is an open access journal, and articles are distributed under the terms of the Creative Commons Attribution-NonCommercial-ShareAlike 4.0 International Public License, which allows others to remix, tweak, and build upon the work noncommercially, as long as appropriate credit is given and the new creations ae licensed under the idential terms. Address for correspondence: Dr. Vasantrao P. Pawar, Dept. of Botany Arts & Science College Bhalod, Tal. Yawal, Dist. Jalgaon. (M.S.) How to cite this article: Vasantrao P. Pawar.,(2025) Bio mechanisms changes of Downy mildew fungi Pseudoperonospora cubensis on Cucurbits. Journal of Research & Development, 17(9), 22-25. Original Article Journal of Research and Development Peer Reviewed International, Open Access Journal. ISSN : 2230-9578 | Website: https://jrdrvb.org Volume-17, Issue-9| September 2025 23 The absorbance was measured at 645 nm and 663 nm double beam spectrophotometer. Chlorophyll [mg/g fresh weights]were calculated using formula given below. Chlorophyll a=12.7 O.D663-2.69 O.D 645 D.F Where 12.7 and 2.69 are constant D.F= Dilution factor Chloropyll b= 22.9 O.D 645-4.68 O.D 663 D.F Where 22.9 and 4.68 are constant D.F.= Dilution factor Total Clorophhyll =20.2 O.D 645 -8.02 O.D 663 D.F. Where 20.2 and 8.02 constant. D.F=Dilution Factor. Nitrogen contentEstimations of nitrogen contents were made by Microkjeldahl method ( A.O.A.C.,1970).For this 300mg dry sample of healthy and infected leaves were taken in Microkjeldhl flasks. A pinch of catalyst was added to it with the help of spatula 7.5 ml. of concentrated sulphuric acid(H2 So4) was added to flask. The flask were heated on a digestion stand unit ( 6 to 10 hr.). The mixture was clearing i.e. apple green in color or colorless. During digestion care was taken to avoid principles of indigested carbon sticking on the sides of the tube. 5 ml of the diluted material was introduced in Markham’s distillation apparatus, through the side tube funnel to which glass stopper was fitted 50 ml conical flask containing 10 ml of 2% boric acid solution mixed with indicate or was at the delivery end of the condenser to collect the ammonium tetra borate( (NH4)3 Bo3) .It was then titrated with 0.035 N HCl/toll the pink color obtained and titration values were recorded .Nitrogen content present in mg/g day sample was calculated by calculating the strength of NH3 in the distillate using equation. 1ml of 0.35 NHCL=0.5 of Nitrogen. From the above equation the amount of nitrogen for 5ml of sample was calculated, which will be equivalent to that of present in 300mg. of dry sample .It was recorded as percentage nitrogen of dry sample. Estimation of crude protein This was done by estimating N content in the sample with the help of Microkjeldahl technique(A.O.A.C.,1970) .The amount of N content was multiplied by 6.25 factor which gave crude protein content of the sample.300 mg seed powder were taken in Microkjeldahl flask along with 250 mg K2 So4 and 40 mg CuSo4 and kept overnight. This was digested till the mixture become white .After complete digestion the flasks were allowed to cool. The digest was processed for distillation with the help of Markham’s distillation test .Digested was diluted to 50 ml. volumetric flask, 5 ml aliquots were taken and introduced in distillation unit through the side tube funnel .The glass stopper was immediately fitted .To this 10 ml 40/NaOH into the digest.NH3 is liberated into 10 ml 2/boric acid (with indicator) containing 50 ml conical flask. After distillation green colored ammonium borate was obtained (This gave 1 ml 0.035 NHCL =0.5 mg N/ crude protein=N/ (6.25) Crude protein of seed was calculated as percent nitrogen liberated 6.5. Estimation of Reducing SugarThe sugar content in the plant material was estimated by the procedure recommended by Oster( 1979) as follow 500 gm. of seed powder was taken in 50 ml. distilled water and boiled it, then filtered it and the filtrate is diluted up to 100ml.Three Following tube taken and added following manner(1) Blank-D.W. 2ml (2) 2ml.glucose C solution (3) 2 ml filtrate .In each tube 3 ml alkaline solution of copper was added then tube was boiled in boiling water bath for 8 minutes .Cooled the tube under tap water and add 2 ml of phosphomolybdic acid solution which gave blue colour .then this solution was diluted up to 25 ml distilled water and optical density determined at 420 nm and calculated the amount of reducing sugar present in seed powder. Estimation of starchStarch contents of healthy and downy mildew infected plant sample were estimated by Anthron Method .For this 0.5 g of the sample was homogenized with hot 80% ethanol and sugars were removed. It was then centrifuged, the residue was dried well over a water bath.0.5 ml of distilled water and 6.5 ml of 52% perchloric acid were added to the residue. It was then centrifuged [after 20 minutes of addition] and the supernatant was collected. The extraction was repeated by using fresh perchloric acid, it was again centrifuged and supernatant was adjusted up to 100 ml with distilled water,0.2 ml of the supernatant was pipetted out and to it 0.8 ml distilled water was added. The standard was prepared by taking 0.2 ml,0.4 ml, 0.6 ml 0.8 ml and 1 ml of the working standard in test tubes. The volume of each tube was made up to 1 ml with distilled water. To each tube 4 ml of Anthron regent was added .It was then heated for 8 minutes in a boiling water bath. The tubes were cooled rapidly in tap water and intensity of green to dark green colour was measured at 630 nm on spectrophotometer. Journal of Research and Development Peer Reviewed International, Open Access Journal. ISSN : 2230-9578 | Website: https://jrdrvb.org Volume-17, Issue-9| September 2025 24 Result and Disccussion It is clear from the (Table-1) that chlorophyll a and chlorophyll b pigment of the infected leaf were found to be decreased due to infection of downy mildew. Nitrogen and protein content of leaves were also depleted to downy mildew .However maximum loss in protein content was reported in the leaves Cucumis melon (Table-2) Reducing sugars were founded decreased in the infected host except Citrullus lanatus and Cucurbita pepo (Table-3) TableNo-1 Effect of Downy Mildew on Chlorophyll content of cucurbits leaves Name of crop Common Name Name of pathogen Chlorophyll a Chlorophyll b Chlorophyll a/b Healthy Infected Healthy Infecte d Healthy Infect ed Citrullus lanatus Water melon Pseudoperonos pora cubensis 0.56 0.23 1.03 0.40 0.54 0.57 Cucumis sativus Cucumber 0.55 0.29 1.13 0.50 0.48 0.58 Cucumis melon Musk melon 0.33 0.18 0.64 0.40 0.51 0.45 Cucurbita pepo Red pumpkin 0.55 0.27 1.07 0.48 0.51 0.56 Lagenaria siceraria Bottle gourd 0.82 0.41 0.52 0.24 1.67 1.70 Luffa actangula Ridge gourd 0.93 0.36 0.87 0.29 1.06 1.24 Luffa cylindria Sponge gourd 0.76 0.32 0.56 0.19 1.35 1.68 TableNo-2 Effect of Downy Mildew on Nitrogen and Protein content of Cucurbits Name of crop Common Name Name of pathogen % Nitrogen % Protein Healthy Infected Healthy Infected Citrullus lanatus Water melon Pseudoperonospo ra cubensis 3.24 2.67 20.00 16.68 Cucumis sativus Cucumber 2.89 2.43 18.06 15.18 Cucumis melon Musk melon 3.14 1.83 19.60 11.43 Cucurbita pepo Red pumpkin 3.07 2.41 19.18 15.06 Lagenaria siceraria Bottle gourd 2.81 1.82 17.56 11.37 Luffa actangula Ridge gourd 3.17 2.18 19.81 13.62 Luffa cylindria Sponge gourd 3.03 2.02 18.93 12.62 Table No-3 Effect of Downy Mildew on Reducing Sugar content of Cucurbits leaves Name of crop Common Name Name of pathogen Reducing sugar ( mg/g) Non-reducing sugar ( mg/g) Total sugar ( mg/g) Healthy Infected Healthy Infected Healt hy Infec ted Citrullus lanatus Water melon Pseudoperono spora cubensis 1.01 0.14 0.34 1.42 1.35 1.56 Cucumis sativus Cucumber 0.96 0.12 0.03 0.98 0.99 1.11 Cucumis melon Musk melon 0.98 0.34 0.02 0.67 1.00 1.01 Cucurbita pepo Red pumpkin 0.64 0.28 0.25 0.71 0.90 0.99 Lagenaria siceraria Bottle gourd 0.84 0.33 0.13 0.69 0.97 1.02 Luffa actangula Ridge gourd 0.71 0.16 0.26 0.87 0.98 1.04 Luffa cylindria Sponge gourd 0.82 0.28 0.16 0.77 0.98 1.05 Journal of Research and Development Peer Reviewed International, Open Access Journal. ISSN : 2230-9578 | Website: https://jrdrvb.org Volume-17, Issue-9| September 2025 25 TableNo-4Effect of Downy Mildew on composition of Phenol and Starch in Cucurbits Name of crop Common Name Name of pathogen Phenol Starch Healthy Infected Healthy Infected Citrullus lanatus Water melon Pseudoperonospora cubensis 0.51 0.74 1.02 0.12 Cucumis sativus Cucumber 0.47 0.69 1.09 0.12 Cucumis melon Musk melon 0.48 0.62 1.08 0.12 Cucurbita pepo Red pumpkin 0.46 0.70 1.09 0.12 Lagenaria siceraria Bottle gourd 0.49 0.69 1.14 0.13 Luffa actangula Ridge gourd 0.54 0.75 0.94 0.12 Luffa cylindria Sponge gourd 0.50 0.71 0.99 0.10 It is clear from the result summarized in table .4 that phenols were found to be increased significantly due to infection in all the cucurbits plant .the increase was maximum in case of with Citrullus lanatus infected Cucumis melon and Luffa actangula infected with Lagenaria siceraria.It was surprising to note that there was no decrease in starch content due to infection of any species of Pseudoperonospora cubensis however there was slight increase in starch contents due to infection in all the cucurbits References 1. 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Salmon( 1900) A morphology of Erysipheae .mem. Torr.Bot.9: 1/292. 10. Sashadri (1986) Chemical Composition of vegetable and Their Products-Scientific .Figure on Research gate. Avabilable Form;[https;//www.researchgate.net/Nuttitional –value-of –cucumber –per 100-g 35-oz-Adapted – after _tabl5_302493121[accessed 12 Nov,2018]