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20212
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Subject: Agricultural Microbiology × Author: KAU × End date: 2021 × Start date: 2021 × Type: Thesis × Thesis Type: M.Sc ×
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    ThesisItemOpen Access
    Plant growth promoting actinobacteria from rhizosphere soils of Black pepper in Wayanad
    (Department of Agricultural Microbiology, College of Agriculture, Vellanikkara, 2021) Rineesha Backer, A; KAU; Girija, D
    Black pepper (Piper nigrum L.) known as the ‘King of spices’ is one of the important export-oriented spice crops that provides major source of income and employment for rural households in Kerala. Wayanad is one of the main pepper growing tracts in Kerala. However, the production of black pepper in Wayanad has been declining over the past many years, mainly due to the poor soil health status, improper land management and changes in climatic factors. An increasing demand for low-input agriculture has resulted in a greater interest in soil microorganisms which are able to enhance plant health and soil quality. However, actinobacteria have not yet been exploited for this purpose. Hence the present study focused on isolation, screening and characterization of actinobacteria from rhizosphere soils of black pepper in Wayanad and evaluation of their plant growth promoting activity. Rhizosphere soil samples were collected from five different locations of flood affected and non-flood affected black pepper growing areas of Wayanad district. Enumeration of actinobacteria on three different media revealed that the population of actinobacteria ranged from 1.00 x104 cfu to 12.00 x104 cfu g -1 soil in non-flooded soil. Actinobacteria could not be detected in flooded soils, even at a dilution of 10-1 , except in Meppadi. Among the different media, starch casein agar recorded higher population of actinobacteria. A total of 35 isolates were purified and maintained for characterization and screening for plant growth promoting activities. Cultural, morphological and biochemical characters of all 35 isolates of actinobacteria were studied. All the isolates were Gram positive, and they varied in the colony morphology, colour of aerial mycelium, spore chain morphology and pigmentation. Two isolates Vlt-K and Plp-B produced yellow water-soluble pigment and Vlt-I produce purple water insoluble pigment. Screening of all the 35 isolates under in vitro conditions for plant growth promoting (PGP) activities revealed that the isolates were highly variable. Two isolates (Ptd-A and Amb-C) were superior to other isolates in IAA production. Significantly higher N-fixation was noticed in four isolates (Ptd-A, Ptd-E, Ptd-B and Ptr-A). Six isolates solubilized insoluble phosphate to available P, and also reduced the pH of the medium. Reduction in pH by phosphate solubilizers has been reported earlier and this is attributed to the production of organic and inorganic acids. None of the isolates solubilized K or Zn. Three isolates with multiple PGP activities were selected, based on ranking for PGP activities in vitro and identified by 16S rRNA gene sequencing (Ptd-A and Ptr-A as Streptomyces sp. and Ptd-E as Actinobacteria bacterium). These actinobacterial isolates evaluated in-planta for PGP activities with black pepper cuttings (variety Panniyur-1). Treatments also included PGPR mix-1 (KAU commercial formulation) and Organic Package of Practices Recommendations (2017). All the three native isolates exhibited significant increase in shoot length, number of leaves and internode length throughout the growth period from planting to five months. Significantly higher root growth was observed in treatment T2: Ptd-A (Streptomyces sp.), with significantly higher root volume, fresh and dry root weight. Significantly higher plant fresh weight was observed in T2: Ptd-A (Streptomyces sp.) and significantly higher plant dry weight was observed in T2: Ptd-A (Streptomyces sp.) and T3: Ptr-A (Streptomyces sp). The present investigation revealed that native actinobacteria have the potential to improve the growth of black pepper. These isolates may further be evaluated under field conditions before commercialization. Screening for other beneficial traits like disease suppression and drought tolerance may also be carried out. The compatibility of the actinobacteria with other PGP microorganisms may also be evaluated. Actinobacterial spores survive in soil for longer periods and this may be an added advantage. Actinobacterial diversity and the biochemical molecules produced by actinobacteria could be exploited to build up sustainable and eco-friendly agriculture.
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    ThesisItemOpen Access
    Isolation and characterization of beneficial rhizosphere microorganisms from ragi grown in Attappady hill tract of Kerala
    (Department of Agricultural Microbiology, College of Agriculture, Vellayani, 2021) Gayathri, M; KAU; Chitra, N
    The study entitled “Isolation and characterization of beneficial rhizosphere microorganisms from ragi grown in Attappady hill tract of Kerala” was conducted during the year 2019-2021 in the Department of Agricultural Microbiology, College of Agriculture, Vellayani, Thiruvananthapuram with the objective to isolate nitrogen fixing and phosphate solubilizing bacteria from soil and root samples collected from Attapady and select efficient isolates through in vitro screening process and assess the plant growth promotion activity by roll towel assay. Thirty six nitrogen fixing bacteria and eight phosphate solubilizing bacteria were isolated from root and soil samples from ragi grown in Attappady region. The isolates were designated as ATY1-ATY36 for NFB isolates and PSB1-PSB8 for PSB isolates. Twenty isolates were selected after eliminating isolates with similarities in colony morphological characters, in order to avoid repetition among the isolates obtained from the same sample. Both NFB and PSB isolates were characterized by morphology and biochemical tests. All selected isolates were subjected to plant growth promotion activity studies. The Indole Acetic Acid (IAA) production showed a wide range of variation from 101.22 µg mL-1 to 3.26 µg mL-1 . Gibberellic Acid (GA) production of all the twenty isolates were done and the results ranged between 10.07 µg mL-1 and 2.18 µg mL-1 . Maximum IAA and GA production was recorded in isolate ATY10. Extracellular ammonia production of the bacterial isolates ranged between 176.53±5.07 μmol mL−1 and 75.59 μmol mL−1. The nitrogen content of NFB isolates ranged between 30.75 µg mL-1 and 9 µg mL-1 . The isolate ATY33 had maximum nitrogen content. The selected PSB isolates were subjected to plate assay and broth assay using Pikovskaya’s medium. The isolate PSB1 recorded maximum zone of solubilization (18.23 mm), phosphate solubilizing index (2.82) and quantification of phosphate solubilization (53.41 mg L-1 ). The bacterial isolates were screened based on weighted average ranking. In NFB, ATY10, ATY34 and ATY35 were selected for further studies and in PSB, the isolates PSB1, PSB3 and PSB4 were selected. The selected isolates were 164 identified as Pantoea agglomerans ATY10, Rhizobium sp. ATY34, Ensifer adhaerens ATY35, Burkholderia territorii PSB1, Burkholderia cepacia PSB3 and Aeromonas hydrophila PSB4 by 16s rRNA sequencing. Effect of these isolates were assessed for plant growth promotion in vitro in ragi seeds using roll towel assay. Maximum germination percentage was recorded by Aeromonas hydrophila PSB4 (81.24 per cent) and the minimum number of days taken for germinationwas recorded by Pantoea agglomerans ATY10 treated seeds. Also, the treatment Pantoea agglomerans ATY10 recorded maximum shoot length (3.31 cm), root length (7.76 cm), seedling length (11.08 cm), seedling vigour index (876.62), root dry weight (0.443 mg) and root shoot ratio (0.50). The treatment Burkholderia territorii PSB1 recorded maximum root fresh weight (3.762 mg) and shoot dry weight (0.877 mg). The six isolates were subjected to assess the multiple traits such as phosphorus, potassium and silicate solubilizing capacity, siderophore production and antifungal activity. Burkholderia territorii PSB1 showed maximum zone of inhibition against Rhizoctonia solani (3.16 mm) and Sclerotium rolfsii (0.58 mm). The maximum antagonistic activity against Fusarium sp. and Helminthosporium sp. was recorded by Burkholderia cepacia PSB3 with 4.13 mm and 5.66 mm of zone of inhibition respectively. Burkholderia territorii PSB1 and Burkholderia cepacia PSB3 solubilized potassium alumino silicate in agar also, with clearance zone of 11.00 mm and 12.66 mm (in diameter), respectively. None of the isolates solubilized magnesium trisilicate in Bunt and Rovira medium. The siderophore production was observed in Aeromonas hydrophila PSB4 isolate in Chrome Azurol S (CAS) agar medium plates. Based on the results of the present study, it can be concluded that Pantoea agglomerans ATY10 is the superior isolate among NFB isolates for plant growth promotion. Also, the isolate Burkholderia territorii PSB1 is best among PSB isolates for plant growth promotion and biocontrol activity.