Combined effect of organic manure and phosphate on growth of Azolla microphylla

Introduction

There are seven well-known species of the fast-growing water fern Azolla (family Salviniaceae) include A. pinnata, A. caroliniana, A. filiculoides, A. mexicana, A. microphylla, A. nilotica, and A. rubra. The name is typically derived from the Greek words azo, which means "to dry," and ollyo, which means "to kill," emphasizing the plant's vulnerability to drying out. Azolla and cyanobacteria (Anabaena azollae or Nostoc azollae), which are housed in cavities within the dorsal leaf lobes and where heterocysts efficiently fix atmospheric nitrogen, create an obligatory symbiotic relationship. The device can fix roughly 1.1 kg N per hectare each day, supplying up to 20-40 kg N/ha per cropping cycle in rice paddies or more depending on conditions. In Asian rice fields, Azolla produces roughly 8-10 t fresh weight/ha/week and doubles its biomass very quickly often in 2-10 days, depending on species and conditions. It has been used as green manure for rice for generations in China and Vietnam, but its use in India and other countries picked up speed in the late 20th century. Each plant is tiny, delicate, and freefloating, with horizontal rhizomes (0.5-7 cm broad) and densely overlapping branching fronds. Each leaf features a bigger dorsal lobe that houses the symbiotic cyanobacteria and a thinner ventral lobe with few stomata and hairs for buoyancy. Azolla reduces reliance on artificial fertilizers by naturally supplying nitrogen to rice and other crops. Cheap, high-yield organic materials that can be used as mulch, compost, or animal feed. It can help with phytoremediation (removing heavy metals) suppress weeds, and lower evaporation. Experiments demonstrate its possible future uses in reducing atmospheric CO2 as well as its historical significance in carbon sequestration (the so-called Azolla Event). Because of its great nutritional value and rapid development, it is often referred to as a "green gold mine" and a "super plant." The purpose of this assessment was to ascertain the impact of zinc, phosphorus, and other micronutrient combinations on Azolla growth.

Material and method

Freshly harvested blotted fern Azolla fronds from mother culture kept in tap water made up the inoculum. Fresh biomass and the dry weight of the fronds were used to record growth observations. Azolla growth was investigated using micronutrients such as potassium hydrogen phosphate (K2HPO4), and zinc phosphate (Zn3(PO4)2) @ 0.6 mg/cm3 and organic manure such as Vermicompost, cowdung @ 2kg in a 200-liter FRP tank measuring 4 × 2 × 1 ft³. Threequarters of the tank's capacity, or 150 liters of water each tank, was filled with tap water. Every micronutrient and organic manure was added according to the specifics of the treatment. After growing the Azolla inoculum in water for five days, 50 grams of Azolla were added to each tank. On the 15th, 30th, 45th, and 60th DAI, observations were made.

Treatments details

The colorimetric method recommended by Subudhi and Watanabe (1981) [13] was used to measure the total chlorophyll content (mg g-1) of the dried leaves. Jackson (1967) provided the vanadomolybdate yellow color method, which was used to assess the phosphorus content of leaves. The Micro Kjeldhal technique described by Ali and Watanabe (1986) [6] was used to analyze the nitrogen content of Azolla. The following formula was used to compute the heterocyst percentage in accordance with.

Result and Discussion

Statistical analysis revealed significant differences among the treatments in terms of both fresh and dry biomass of Azolla microphylla when compared with the uninoculated control. Among the treatments, T5 (Cowdung + K₂HPO₄) recorded the maximum fresh biomass throughout the experimental period. The fresh weight under T5 was 830.66 g/pot at 15 DAI, which increased to 1330.66 g/pot at 30 DAI, 1930.89 g/pot at 45 DAI, and 2034.22 g/pot at 60 DAI. Thus, T5 maintained its superiority over the other treatments at all observation intervals. This result are in accordance with earlier findings of Hamid et al. (2007) [6] , Ali and Watanabe (1986) [6] , Dehane (2022) [3] . A comparable pattern was observed for dry biomass. [5] , Andreeilee et al. (2015) [2] , and Ismail et al. (2010) [7] . Heterocyst percentage, an important indicator of nitrogenfixing activity in Azolla, was evaluated at each observation interval. Treatment T5 (Cowdung + K₂HPO₄) recorded the highest heterocyst percentage, with values of 12.05% at 15 DAI, 29.27% at 30 DAI, 33.01% at 45 DAI and 35.01% at 60 DAI. This was followed by T9 (K₂HPO₄ + Zn₃(PO₄)₂ + Cowdung + Vermicompost) and T7 (Vermicompost + K₂HPO₄). The treatments differed significantly from the uninoculated control. The heterocyst percentage generally showed an increasing tendency up to 60 DAI, while a decline was observed from 45 to 60 DAI according to the overall trend described for the treatments. Despite these variations, several treatments were statistically at par with one another. The reported by Potdukhe et al (2020) [10] documented the same. Phosphorus and nitrogen concentrations of Azolla microphylla. Treatment T5 consistently recorded the highest nitrogen concentration, whereas T9 exhibited the highest phosphorus concentration throughout the observation period. Phosphorus concentration among the treatments ranged from 0.36 to 0.80% at 15 DAI, 0.42 to 0.83% at 30 DAI, 0.48 to 0.88% at 45 DAI and 0.45 to 0.95% at 60 DAI. Similarly, nitrogen concentration varied from 3.58 to 4.58% at 15 DAI, 3.78 to 4.75% at 30 DAI, 3.89 to 4.83% at 45 DAI and 3.72 to 4.93% at 60 DAI. The findings indicated that the important role of phosphorus in the growth and productivity of Azolla microphylla. Phosphate deficiency was associated with reduced biomass production, lower chlorophyll concentration, decreased heterocyst percentage and reduced nitrogen and phosphorus accumulation. Earlier studies conducted by Subudhi et al. (1981) [13] , Hamid et al (2007) [6] , Potdukhe et al (2020) [10] , Gatole (2019) [4] . Frond size of Azolla microphylla was recorded at 15, 30, 45 and 60 DAI. Treatment T5 (Cowdung + K₂HPO₄) produced the largest fronds throughout the experimental period. The corresponding values were 1.67, 1.73, 1.81 and 1.89 cm² at 15, 30, 45 and 60 DAI, respectively, indicating its superior performance over the remaining treatments. In contrast, T2 (Zn₃(PO₄)₂ resulted in the smallest frond size during the observation period. The values recorded under this treatment were 1.12, 1.20, 1.25 and 1.66 cm² at 15, 30, 45 and 60 DAI, respectively. The results of the present investigation are similar to those of Dehane (2022) [3] . The present investigation revealed that varying levels of organic matter had a significant effect on the growth, yield, and physiological characteristics of Azolla. Among the treatments evaluated, T5 (Cowdung+ (K 2 HPO 4 )) exhibited the best overall performance throughout the observation period. This treatment produced the maximum fresh and dry biomass, chlorophyll content, heterocyst percentage, nitrogen and phosphorus content, along with greater frond size at all observation intervals.The improved growth and physiological attributes may be associated with enhanced nutrient availability, which promoted efficient nutrient assimilation and symbiotic activity. These results are consistent with earlier studies that highlighted the beneficial role of organic manure and micronutrient amendments in improving Azolla production. Hence, the application of Cowdung+ (K 2 HPO 4 ) at an appropriate level may be recommended as a suitable practice for sustainable Azolla cultivation and effective nutrient management. The longterm effectiveness of cowdung + K₂HPO₄ on Azolla growth, nutrient accumulation, and its potential as a biofertilizer may also be assessed.

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