Middle East Journal of Applied Volume : 09 | Issue :02 |April-June| 2019 Sciences Pages: 319-325 ISSN 2077-4613

Applying biofertilizer and different rates of for the production of squash

Saad Abou-El-Hassan, Hassan G. Elmehrat, Wael M. Ibrahim and Atef A. Ragab

Central Lab of Organic Agriculture, Agricultural Research Center, Giza, Egypt. Received: 10 Mar. 2019 / Accepted 15 April 2019 / Publication date: 30 April 2019

ABSTRACT Field experiment was conducted at Giza Agricultural Research Station, Giza Governorate, Egypt, during the two autumn successive seasons of 2016 and 2017. This study aims to evaluate the production of squash (cv. Eskandarani) using biofertilizers under different rates of compost as an alternative to mineral . Three rates of compost 6, 8 and 10 tons/feddan (feddan = 0.42 hectare) with and without adding biofertilizers were compared to recommended mineral fertilizers of N, P and K. Biofertilizers were used as bacterial mixture of chroococcum, Azospirillium brasilense, Bacillus megaterium and Bacillus circulans. The results indicated that using high compost rate (10 t/fed.) with adding biofertilizers improved significantly vegetative growth (leaves No/plant and chlorophyll reading of leaves), nutritional status of plants, yield and fruit compositions compared to recommended mineral . Treatment of middle compost rate (8 t/fed.) with biofertilizers produced a yield similar to the recommended mineral fertilizers treatment and exceeded in the total soluble solid and dry matter contents of fruits. All compost rates with or without biofertilizers lessened nitrate content in squash fruits. This study demonstrated the possibility of producing a good yield of squash safe and healthy without applying mineral fertilizers via integration of compost and biofertilizers as environmentally friendly practices.

Keywords: Squash, Compost, Biofertilizers, Alternatives of mineral fertilizer

Introduction Squash (Cucurbita pepo L.), is considered one of the main vegetable crops in Egypt and other countries of the Mediterranean region. The total cultivated area in Egypt was 59340 feddans (feddan = 0.42 hectare), produced about 455866 tons on annual basis with an average of 7.682 tons/feddan (Ministry of Agriculture and Land Reclamation, 2015). Chemical fertilizers cause several harmful effects on the environment and human health. These fertilizers are rapidly lost by leaching in drainage water; this leads to dangerous environmental pollution (Hernandez et al., 2010). With the increasing warning of the harmful effects of the chemical fertilizers, substitution of chemical fertilizers with organic and bio fertilizers has become very important for sustainability of agriculture production and maintain of (Abou-El-Hassan et al., 2017). Sustainable farming aim to produce healthy food without causing adverse effects on the natural environment. This goal can be achieved through the use of non-chemical nutrients such as organic and biofertilizers (Lotter, 2003). Application of compost to soil improves their physical, chemical and biological characteristics, such as retention water, aggregation, porosity, increasing of cation exchange capacity, fertility, increasing of microbial and biological activities in the rhizosphere (Miyasaka et al., 1997; Ahmad et al., 2008; Fiorentino and Fagnano, 2011). Compost is commonly added at one dose before the planting this lead to the available nutrients from compost, is inadequate for crop requirements. Therefore, some amendments should be added to help such as bio-fertilizers (Abou-El-Hassan et al., 2014). Biofertilizer is a substance containing living microorganisms, when applied to seed, plant root or soil, colonize the rhizosphere zone and promoting growth by increasing the availability of nutrients to the host plant (Vessey, 2003). Biofertilizers include on microorganisms that fixing, dissolving, potassium releasing and plant growth promoting (Goel et al., 1999). Biofertilizers are influenced the growth, yield and nutrient uptake by an array of mechanisms. They help in promoting free-living nitrogen-fixing bacteria, increase supply of other nutrients, such as , potassium, sulphur and micronutrients, produce plant hormones like cytokinins, Indole

Corresponding Author: Saad Abou-El-Hassan, Central Lab of Organic Agriculture, Agricultural Research Center, Giza, Egypt. E-mail: [email protected] 319 Middle East J. Appl. Sci., 9(2): 319-325, 2019 ISSN 2077-4613

Acetic Acid, Gibberillins (Vessey, 2003 and Saharan & Nehra 2011). Many studies showed that the combination of biofertilizers with organic fertilizers enhanced the growth, yield and quality of plants such as Fawzy et al. (2007), Glala et al., (2010) and Marajan et al., (2017) on tomato, Habibi et al. (2011) on pumpkin, Sarhan et al. (2011), Glala et al. (2012) and Jahan et al. (2012) on squash and Shedeed et al. (2014) and Singh et al. (2015) on onion. The objective of this work was to evaluate the production of squash using biofertilizer under different rates of compost as an alternative to mineral fertilizers.

Materials and Methods The experiment was conducted at Giza Agricultural Research Station, Giza Governorate, Egypt, during the two autumn successive seasons of 2016 and 2017. Seeds of squash (cv. Eskandarani) were sown in the field on 23th and 27th of August in the first and second seasons, respectively. The experimental soil was analyzed according to FAO (2008) and showed in Table 1.

Table 1: Physical and chemical properties of the experimental soil Sand Silt Clay EC Cations meq/l Anions meq/l Texture pH ++ ++ + + = - - = % % % dS/m Ca Mg K Na Co3 HCO3 Cl SO4 23.3 35.2 41.5 Clay loam 7.63 2.05 4.20 2.80 1.80 10.35 - 0.76 16.23 1.80

Experimental soil was tilled and divided into ridges (80 cm width). The seeds were sown at a distance of 40 cm on one side of ridge and irrigated using surface system. The plot area was 12 m2 (3 m length and 4 m width). Each plot included 5 ridges. Three rates of compost 6, 8 and 10 tons/fed. with and without addition of biofertilizers were investigated for production of squash comparing to conventional production by recommended dose of NPK as mineral fertilizers (control). Analyses of the used compost were analyzed according to FAO (2008) and illustrated in Table (2). Biofertilizers were used as mixture of Azotobacter chroococcum, Azospirillium brasilense (nitrogen fixing bacteria), Bacillus megaterium (phosphate dissolving bacteria) and Bacillus circulans (potassium releasing bacteria).

Table 2: Analyses of the used compost EC 1:10 O.C O.M C/N N P2O5 K2O N-NH4 N-NO3 pH 1:10 dS/m (%) (%) Ratio (%) ppm 7.12 3.69 17.15 28.13 18.84 0.91 0.71 1.20 302.00 76.00

The Experimental Treatments were as follow: 1. Recommended dose of NPK as mineral fertilizers (RMF) as a control treatment. 2. Rate 1 of compost as 6 tons/fed. (Comp1) 3. Rate 2 of compost as 8 tons/fed. (Comp2) 4. Rate 3 of compost as 10 tons/fed. (Comp3) 5. Rate 1 of compost as 6 tons/fed. + biofertilizers (Comp1 + biofertilizers) 6. Rate 2 of compost as 8 tons/fed. + biofertilizers (Comp2 + biofertilizers) 7. Rate 3 of compost as 10 tons/fed. + biofertilizers (Comp3 + biofertilizers)

A completely randomized block design with three replicates was used. The recommended dose of N, P and K were applied as follow 60 kg N/fed. as 293 kg ammonium sulphate (20.5% N), 30 kg P2O5/fed. as 194 kg super phosphate (15.5% P2O5) and 48 kg K2O/fed. as 100 kg potassium sulphate (48% K2O). Calcium super phosphate was added as one dose during soil preparation, whereas ammonium sulphate and potassium sulphate were added at three equal portions, after 2, 4 and 6 weeks from set up seeds. Each quantities of compost were added as one dose during soil preparation. Each kind of biofertilizers was added with the irrigation after 2 and 4 weeks from seeds sowing at a rate of 5 L/fed. (1ml contains 106-8cell) according to Mashhoor et al. (2002).

320 Middle East J. Appl. Sci., 9(2): 319-325, 2019 ISSN 2077-4613

After 45 days from sowing, the following parameters were measured; shoot fresh weight, leaf number on plant and chlorophyll reading in the third leaf of inner by using Minolta Chlorophyll Meter Spad 501. Nutrient contents (N, P and K) in squash plants were determined in the inner mature leaf according to Cottenie et al. (1982). Total nitrogen was determined by Kjeldahl method, phosphorus was determined using spectrophotometer and potassium was determined photometrically using flame photometer according to the procedure described by FAO (2008). Harvesting began after 50 days of sowing seeds and lasted for 5 weeks. Fruits were harvested twice a week. Total yield for each plot and fruits number for plant were recorded after each harvesting accumulatively. Percentage of total soluble solids (TSS) in fruits was measured by using Digital Refractometer. As well as, dry matter percent of fruit was determined. Nitrate content of fruits was performed using Cardy Nitrate Meter Model HORIBA, Spectrum Technologies, Inc., as described by Al-Moshileh et al. (2004). Data were statistically analyzed by the analysis of variance according to Snedecor and Cochran (1980) by SAS software, version 2004. Treatment means were compared using Tukey test at 5% level of probability.

Results and Discussion

Data presented in Table (3) showed that all rates of compost with or without biofertilizers decreased shoot fresh weight of squash plants compared to recommended mineral fertilizers in both seasons. On the other hand, using the high rate of compost (10 t/fed.) with adding biofertilizers increased leaves number and chlorophyll reading of squash leaves compared to other treatments. The increment in shoot fresh weight of plants treated with mineral fertilizer over compost treatments might be due to the plants obtained nitrogen from mineral fertilizer more easily than , that increases water content of plant cells, thus increasing the fresh weight of the plants. This interpretation was confirmed by Shedeed et al. (2014), Lasmini et al. (2015) and Singh et al. (2015). Increasing leaves number and chlorophyll reading of plants treated by the high rate of compost with adding biofertilizers might be attributed to increase the amount of compost indo soil, which improves the physical properties of soil, thus improving the growth of plant roots (Miyasaka et al., 1997; Ahmad et al., 2008; Fiorentino and Fagnano, 2011). Besides, adding bio fertilizers in the presence of the high rate of compost led to an increase in the mass of these bio-fertilizers in the soil, which increased their efficiency in improving the growth of plants by production of growth promoting compounds like cytokinins, Indole Acetic Acid, Gibberillins and releasing of many nutrients found in the compost, as well as fixing atmospheric nitrogen in a form available for absorption by plant roots (Vessey, 2003 and Saharan & Nehra 2011). All that have been reflected in improved vegetative growth by increasing leaves number and chlorophyll reading of plants.

Table 3: Effect of treatments on vegetative growth characteristics of squash plants during 2016 and 2017 seasons Shoot fresh weight Leaf Chlorophyll Treatments kg No. SPAD 1st season 2nd season 1st season 2nd season 1st season 2nd season RMF 1.27 a 1.30 a 15.50 b 16.00 b 43.07 bc 43.63 bc Comp1 0.45 e 0.46 e 10.33 d 10.33 d 34.90 e 35.33 f Comp2 0.62 d 0.65 d 12.67 c 12.67 c 38.77 d 38.67 e Comp3 0.75 c 0.77 c 13.67 c 13.83 c 41.33 c 41.77 d Comp1 +Biofertilizers 0.73 c 0.77 c 13.33 c 13.83 c 42.43 bc 42.90 cd Comp2 +Biofertilizers 1.05 b 1.16 b 15.33 b 16.33 ab 44.33 ab 45.07 ab Comp3 +Biofertilizers 1.10 b 1.19 b 17.00 a 17.33 a 45.67 a 46.00 a Means followed in same column by similar letters are not statistically different at 0.05 level according to Tukey test. RMF = recommended dose of mineral fertilizer Comp1 = 6 t/fed of compost Comp2 = 8 t/fed of compost Comp3 = 10 t/fed of compost

Effect of treatments on the nutritional status in squash plants show in Table 4. Using the high rate of compost with adding biofertilizers gave the highest values of N, P and K contents of leaves. Applying the middle rate of compost (8 t/fed) with adding biofertilizers gave N, P and K contents of

321 Middle East J. Appl. Sci., 9(2): 319-325, 2019 ISSN 2077-4613 leaves similar to recommended mineral fertilizers treatment. While, the rest treatments reduced N, P and K contents of leaves compared to recommended mineral fertilizers. High nutrient contents of squash plants treated with high rate of compost and biofertilizers may be explained to the role of biofertilizers, that fixing atmospheric nitrogen and the releasing of essential nutrients found in the compost to be available for plants, where compost is considered a large store of nutrients, especially P, K and micronutrients (Vessey, 2003 Ahmad et al., 2008; Fiorentino and Fagnano, 2011).

Table 4: Effect of treatments on nutritional status of squash plants during 2016 and 2017 seasons % N % P % K Treatments 1st season 2nd season 1st season 2nd season 1st season 2nd season RMF 3.727 b 3.783 b 0.411 b 0.419 b 3.683 c 3.737 bc Comp1 2.057 f 2.067 f 0.243 f 0.245 f 2.400 g 2.373 f Comp2 2.361 e 2.441 e 0.280 e 0.285 e 2.777 f 2.860 e Comp3 2.772 d 2.927 d 0.325 d 0.342 d 3.120 e 3.287 d Comp1 +Biofertilizers 3.167 c 3.211 c 0.365 c 0.375 c 3.443 d 3.523 c

Comp2 +Biofertilizers 3.711 b 3.811 ab 0.428 b 0.435 b 3.860 b 3.940 b Comp3 +Biofertilizers 3.897 a 3.937 a 0.447 a 0.458 a 4.167 a 4.197 a Means followed in same column by similar letters are not statistically different at 0.05 level according to Tukey test. RMF = recommended dose of mineral fertilizer Comp1 = 6 t/fed of compost Comp2 = 8 t/fed of compost Comp3 = 10 t/fed of compost

Effect of different treatments on yield component is presented in Table (5) Data indicated that high rate of compost with adding biofertilizers treatment increased fruits number/plant and total yield of squash compared to recommended mineral fertilizer. There were no significant differences between treatments of middle compost rate with biofertilizers and recommended mineral fertilizers. The rest treatments decreased fruits number/plant and total yield of squash compared to recommended mineral fertilizers. These results were occurred in both seasons. Superior the yield in treatment of high compost rate with biofertilizers might be attributed to add the biofertilizers in the presence of a high compost rate, which led to increased efficiency of biofertilizers in the secretion of growth promoters and the fixation of atmospheric nitrogen and releasing essential nutrients in the form available for absorption by plant roots. Besides, compost improves the physical properties of soil, thus improving the growth of plant roots, thereby increasing its nutrient uptake. All these improve the nutritional status of plants and consequently increasing yield component of squash. These results are concordant with those reported by Habibi et al. (2011) on pumpkin, Sarhan et al. (2011), Glala et al. (2012), and Jahan et al. (2012) on squash, Abou-El-Hassan et al. (2014) on cucumber.

Table 5: Effect of different treatments on fruits/number and yield of squash during 2016 and 2017 seasons Yield Fruits No/plant Treatments t/fed. 1st season 2nd season 1st season 2nd season RMF 14.667 b 15.000 b 9.090 b 9.190 b Comp1 9.500 e 9.667 e 5.773 e 5.810 e Comp2 11.167 d 11.500 d 6.722 d 6.796 d Comp3 12.833 c 13.267 c 7.907 c 8.175 c Comp1 +Biofertilizers 12.667 c 13.000 c 7.869 c 8.125 c

Comp2 +Biofertilizers 14.667 b 15.000 b 9.068 b 9.230 b Comp3 +Biofertilizers 15.833 a 16.167 a 9.566 a 9.643 a Means followed in same column by similar letters are not statistically different at 0.05 level according to Tukey test. RMF = recommended dose of mineral fertilizer Comp1 = 6 t/fed of compost Comp2 = 8 t/fed of compost Comp3 = 10 t/fed of compost

322 Middle East J. Appl. Sci., 9(2): 319-325, 2019 ISSN 2077-4613

Effect of different fertilizers on fruit composition is showed in Table 6. Using high and middle compost rate with biofertilizers increased percent of TSS content in fruits compared to recommended mineral fertilizers. This may be attributed to the role of biofertilizers in the presence of an adequate amount of compost in fixing K element, which increased K content of plant (Table 4), which helps in the transmission of photosynthesis products to the fruits, thus increasing TSS content of fruits. This result was in conformity with the results obtained by Abou-El-Hassan et al. (2014) and Lasmini et al. (2015). All rates of compost with or without biofertilizers increased dry matter percent compared to recommended mineral fertilizers. This result may be due to available nitrogen in organic fertilizer is low, which leads to metabolism more toward to formation of carbon compounds, that are the main component of dry matter in fruits (Brandt and Mølgaard, 2001; Rembialkowska, 2007). On the contrary, all rates of compost with or without biofertilizers reduced nitrate content of squash fruits significantly compared to recommended mineral fertilizer treatment. This can be explained by the fact that applying organic fertilizers decrease nitrate accumulation in vegetables as was reported by Mahmoud et al. (2009), Abou-El-Hassan et al. (2014) on cucumber and Abou-El-Hassan et al. (2017) on green bean.

Table 6: Effect of different treatments on TSS, dry matter and nitrate contents of squash during 2016 and 2017 seasons % TSS % Dry matter % Nitrate Treatments 1st season 2nd season 1st season 2nd season 1st season 2nd season RMF 4.087 c 4.103 cd 2.830 d 2.867 e 0.220 a 0.227 a Comp1 3.007 f 3.060 f 3.073 bc 3.083 d 0.067 e 0.068 d Comp2 3.373 e 3.340 e 3.050 c 3.083 d 0.075 de 0.076 c Comp3 3.740 d 3.840 d 3.090 bc 3.123 cd 0.082 cd 0.083 c Comp1 +Biofertilizers 4.080 c 4.113 c 3.160 ab 3.184 bc 0.080 cd 0.081 c

Comp2 +Biofertilizers 4.320 b 4.460 b 3.217 a 3.243 ab 0.090 bc 0.092 b Comp3 +Biofertilizers 4.773 a 4.793 a 3.243 a 3.267 ab 0.095 b 0.097 b Means followed in same column by similar letters are not statistically different at 0.05 level according to Tukey test. RMF = recommended dose of mineral fertilizer Comp1 = 6 t/fed of compost Comp2 = 8 t/fed of compost Comp3 = 10 t/fed of compost

Conclusion It could be concluded that high compost rate (10 t/fed.) with adding biofertilizers as mixture of Azotobacter chroococcum, Azospirillium brasilense, Bacillus megaterium and Bacillus circulans was superior of yield and quality of squash fruits compared to recommended mineral fertilizers. Integration of compost and biofertilizers can be used as an alternative to mineral fertilizers to produce a good and healthy yield of squash.

Acknowledgment This work has been supported by Central Lab of Organic Agriculture, Agricultural Research Center.

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