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Practical Handbook onAgricultural Microbiology

Edited by

Natarajan Amaresan, Prittesh Patel,and Dhruti Amin

Uka Tarsadia University, Surat, Gujarat, India

EditorsNatarajan AmaresanUka Tarsadia UniversitySurat, Gujarat, India

Prittesh PatelUka Tarsadia UniversitySurat, Gujarat, India

Dhruti AminUka Tarsadia UniversitySurat, Gujarat, India

ISSN 1949-2448 ISSN 1949-2456 (electronic)Springer Protocols HandbooksISBN 978-1-0716-1723-6 ISBN 978-1-0716-1724-3 (eBook)https://doi.org/10.1007/978-1-0716-1724-3

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Preface

Agricultural Microbiology is a part of the microbiology branch dealing with beneficial orharmful microbes associated with either plants or soil. This manual focuses on beneficialmicrobes dealing with soil fertility, microbial degradation of organic matter, soil nutrienttransformations, and biocontrol agents. Nowadays, techniques involved in the study ofbeneficial microbes in agricultural microbiology toward enhancing global agricultural pro-ductivity are in trend. This manual covers a wide range of basic and advanced techniquesassociated with research on the isolation of agriculturally important microbes, identification,biological nitrogen fixation, microbe-mediated plant nutrient use efficiency, and biologicalcontrol of plant diseases and pests. Introduction to each protocol explains the role/impor-tance of chemicals involved, uniqueness, and protocol application. A proper understandingof the protocol helps the researchers to manipulate them as per their need.

This book is composed of seven parts with 52 protocol chapters. Parts I and II representthe importance, isolation, and purification methods of agriculturally important microbesand include mineral-solubilizing microbes. Part III deals with phytohormones quantitativeprotocols directly or indirectly associated with microbes. Parts IV and V provide deepinsights into protocols for screening agriculturally important enzymes and compoundsrelated to biocontrol activity. Part VI represents assessment methods of soil microbialactivity by soil respiration. The final Part VII deals with protocols for selecting microbialstrains for inoculant production and quality control ultimately representing commercialbiofertilizers production criteria. This book will help postgraduate students, research scho-lars, postdoctoral fellows, and teachers belonging to different disciplines of Plant Microbi-ology and Pathology. Moreover, this manual may also serve as a textbook for undergraduatecourses like Techniques on Plant-Microbe Interaction/Biological Control of Plant Dis-eases/Nutrient Use Efficiency.

Surat, Gujarat, India Natarajan AmaresanSurat, Gujarat, India Prittesh PatelSurat, Gujarat, India Dhruti Amin

v

Contents

Preface . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . vContributors. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . xi

PART I ISOLATION AND IDENTIFICATION OF AGRICULTURALLY

IMPORTANT MICROBES

1 Methods for Isolation and Identification of Rhizobia . . . . . . . . . . . . . . . . . . . . . . . . 3Vrutuja Naik and Praveen Rahi

2 Isolation of Frankia from Casuarina Root Nodule . . . . . . . . . . . . . . . . . . . . . . . . . 15Narayanasamy Marappa, Dhanasekaran Dharumadurai,and Thajuddin Nooruddin

3 Isolation and Identification of Nonsymbiotic Azotobacterand Symbiotic Azotobacter Paspali–Paspalum notatum . . . . . . . . . . . . . . . . . . . . . . 25Bhavana V. Mohite and Satish V. Patil

4 Isolation and Identification of Azospirillum . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 35Harshida A. Gamit and Natarajan Amaresan

5 Isolation and Identification of Gluconacetobacter diazotrophicus . . . . . . . . . . . . . . . 41K. Sowmiya and Mahadevaswamy

6 Isolation and Identification of Nitrogen Fixing Bacteria: AzoarcusSpecies. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 47Rashmi Thakor, Harsh Mistry, and Himanshu Bariya

7 Isolation and Identification of Derxia Species from the Soil Sample . . . . . . . . . . . 57Harshida A. Gamit and Natarajan Amaresan

8 Isolation and Characterization of Enterobacter, Klebsiella,and Clostridium . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 63V. Mageshwaran and K. Pandiyan

9 Isolation and Characterization of Genus Desulfotomaculum . . . . . . . . . . . . . . . . . . 71Mohini Pimpalse, Harshida A. Gamit, and Natarajan Amaresan

10 Isolation and Identification of Associative Symbiotic N2 Fixing Microbes:Desulfovibrio . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 77Harsh Mistry, Rashmi Thakor, and Himanshu Bariya

11 Cyanobacteria . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 85Dhruti Amin, Abhishek Sharma, and Sanket Ray

12 Pseudomonas . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 93Sanket Ray and Harsh Patel

13 Isolation and Identification of Entomopathogenic Bacillus Species . . . . . . . . . . . . 99Preeti Parmar, B. K. Rajkumar, and Naresh Butani

14 Methylobacterium . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 111Harshida A. Gamit and Natarajan Amaresan

vii

15 Isolation and Identification of Beijerinckia . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 119Harshida A. Gamit and Natarajan Amaresan

16 Isolation of Streptomyces from Soil Sample . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 127Vishnu Raja Vijayakumar and Dharumadurai Dhanasekaran

17 Isolation and Identification of Trichoderma Spp. from DifferentAgricultural Samples . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 131Harsh Mistry and Himanshu Bariya

18 Extraction, Isolation and Culturing of Mycorrhizal Sporesfrom Rhizospheric Soil . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 145Satish V. Patil, Bhavana V. Mohite, and Chandrashekhar D. Patil

19 Isolation and Identification of Metarhizium . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 151Tarun Kumar Patel

20 Isolation and Identification of Bacteriophage for Biocontrol . . . . . . . . . . . . . . . . . 161Mitesh Dwivedi

21 Isolation of Bacterivorous Protozoan, Acanthamoeba Spp.,as New-Age Agro Bio-Input . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 173Chandrashekhar D. Patil, Bhavana V. Mohite, and Satish V. Patil

PART II ISOLATION OF MINERAL SOLUBILIZING MICROBES

22 Isolation and Screening of Zinc Solubilizing Microbes: As EssentialMicronutrient Bio-Inputs for Crops. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 181Satish V. Patil, Hemant P. Borase, Jitendra D. Salunkhe,and Rahul K. Suryawanshi

23 Isolation and Screening of Mineral Phosphate SolubilizingMicroorganisms . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 187Swati Patel, Vimalkumar Prajapati, and Prittesh Patel

24 Isolation and Identification of Potassium-Solubilizing Microbes . . . . . . . . . . . . . . 193Prittesh Patel and Swati Patel

25 Isolation and Identification of Sulfur-Oxidizing Bacteria . . . . . . . . . . . . . . . . . . . . . 197Vimalkumar Prajapati, Swati Patel, Radhika Patel,and Vaibhavkumar Mehta

26 Isolation of Ammonia Oxidizing Bacteria . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 203Naresh Butani, Shruti Satashia, Hemanshi Kanpariya,and Preeti Parmar

27 Isolation and Identification of Nitrite-Oxidizing Microbes . . . . . . . . . . . . . . . . . . . 219Prittesh Patel, Ami Naik, and Abhishek Sharma

28 Isolation and Identification of Iron-Oxidizing Microbes . . . . . . . . . . . . . . . . . . . . . 223Ami Naik and Pooja Patel

29 Isolation and Identification of Phytin Mineralizing Microbes . . . . . . . . . . . . . . . . . 231Swati Patel and Prittesh Patel

30 Isolation and Screening of Silicate Solubilizing Microbes: ModernBioinputs for Crops . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 237Chandrashekhar D. Patil, Bhavana V. Mohite,Rahul K. Suryawanshi, and Satish V. Patil

viii Contents

31 Isolation of Selenium Biotransforming Microbes as New Age Bioinputs . . . . . . . 243Pradnya B. Nikam, Narendra Salunkhe, Vishal Marathe,Bhavana V. Mohite, Satish V. Patil, and Vikas S. Patil

PART III ESTIMATION OF PHYTOHORMONES BY BENEFICIAL

MICROBES

32 Auxin . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 251Dixita Panchal, Jemisha Mistry, and Dhruti Amin

33 Abscisic Acid . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 257Natarajan Amaresan, A. Sankaranarayanan, and Dhruti Amin

34 Cytokinins . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 263Jawahar Ganapathy, Jemisha Mistry, and Dhruti Amin

35 Ethylene . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 269Ketankumar J. Panchal and Dhruti Amin

36 Gibberellin . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 273Dhruti Amin, Sanket Ray, and Abhishek Sharma

37 Brassinosteroids . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 277Ketankumar J. Panchal, Dhruti Amin, and Tejas Gohil

38 Strigolactones: Extraction and Characterization . . . . . . . . . . . . . . . . . . . . . . . . . . . . 283Hemant P. Borase, Satish V. Patil, and Dhruti Amin

39 Estimation of Jasmonic Acid Using Non-pathogenic MicrobesJasmonic Acid . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 289Krutika S. Abhyankar and Monisha Kottayi

PART IV SCREENING OF AGRICULTURALLY IMPORTANT ENZYMES

40 Chitinase . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 301Purvesh B. Bharvad and Harsha J. Algotar

41 Glucanase . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 309Purvesh B. Bharvad and Harsha J. Algotar

42 Identification of Cellulase Enzyme Involved in Biocontrol Activity. . . . . . . . . . . . 317Ketankumar J. Panchal

43 Identification of Protease Enzymes Involved in Biocontrol Activity . . . . . . . . . . . 323Vimalkumar Prajapati, Swati Patel, Sanket Ray,and Kamlesh C. Patel

44 Isolation and Screening of Naringinase Producing Microbes:As Industrial Inputs for Agro Waste Base Enzyme Industry . . . . . . . . . . . . . . . . . . 331Satish V. Patil, Jitendra D. Salunkhe, and Vishal Marathe

45 Isolation and Screening of Phytase Producing Microorganisms:An Essential Bioinput for Soil Fertility. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 337Bhavana V. Mohite, Kiran Marathe, Narendra Salunkhe,and Satish V. Patil

Contents ix

PART V IDENTIFICATION OF COMPOUNDS INVOLVED IN BIOCONTROL

ACTIVITY

46 Hydrogen Cyanide. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 345Khyati Bhatt and Dhruti Amin

47 Siderophores . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 351Nafisa Patel

48 Isolation and Screening of ACC Deaminase-Producing Microbesfor Drought Stress Management in Crops . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 361Satish V. Patil, Chandrashekhar D. Patil, and Bhavana V. Mohite

49 Exopolysaccharides. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 369Sapna Chandwani and Natarajan Amaresan

PART VI ASSESSMENT OF SOIL MICROBIAL ACTIVITY BY SOIL RESPIRATION

50 Enzymatic Analyses in Soils . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 377Serdar Bilen and Veysel Turan

PART VII SELECTION OF MICROBIAL STRAINS FOR INOCULANT

PRODUCTION AND QUALITY CONTROL

51 Selection of Rhizobium Strain for Inoculum Production . . . . . . . . . . . . . . . . . . . . . 389Shreya Desai and Natarajan Amaresan

52 Mother Culture, Broth, and Peat Test . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 395Dhruti Amin, Sanket Ray, and Vrushali Wagh

Index . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 405

x Contents

Contributors

KRUTIKA S. ABHYANKAR • School of Science, Navrachana University, Vadodara, Gujarat,India

HARSHA J. ALGOTAR • D. L. Patel Science College, Himatnagar, Gujarat, IndiaNATARAJAN AMARESAN • C.G. Bhakta Institute of Biotechnology, Uka Tarsadia University,

Surat, Gujarat, IndiaDHRUTI AMIN • C. G. Bhakta Institute of Biotechnology, Uka Tarsadia University, Surat,

Gujarat, IndiaHIMANSHU BARIYA • Department of Life sciences, Hemchandracharya North Gujarat

University, Patan, Gujarat, IndiaPURVESH B. BHARVAD • D. L. Patel Science College, Himatnagar, Gujarat, IndiaKHYATI BHATT • Post Graduate Department of Biosciences, Sardar Patel University, Gujarat,

IndiaSERDAR BILEN • Department of Soil Science and Plant Nutrition, Faculty of Agriculture,

Ataturk University, Erzurum, TurkeyHEMANT P. BORASE • C.G. Bhakta Institute of Biotechnology, Uka Tarsadia University,

Bardoli, Gujarat, IndiaNARESH BUTANI • Department of Microbiology, Faculty of Science, Sarvajanik University,

Surat, IndiaSAPNA CHANDWANI • C.G. Bhakta Institute of Biotechnology, Uka Tarsadia University,

Surat, IndiaSHREYA DESAI • C.G. Bhakta Institute of Biotechnology, Uka Tarsadia University, Surat,

IndiaDHANASEKARAN DHARUMADURAI • Department of Microbiology, School of Life Sciences,

Bharathidasan University, Tiruchirappalli, Tamil Nadu, IndiaMITESH DWIVEDI • C. G. Bhakta Institute of Biotechnology, Faculty of Science, Uka Tarsadia

University, Surat, Gujarat, IndiaHARSHIDA A. GAMIT • C.G. Bhakta Institute of Biotechnology, Uka Tarsadia University,

Surat, Gujarat, IndiaJAWAHAR GANAPATHY • C. G. Bhakta Institute of Biotechnology, Uka Tarsadia University,

Surat, Gujarat, IndiaTEJAS GOHIL • Sabarmati Ashram Gaushala, Kheda, Gujarat, IndiaHEMANSHI KANPARIYA • Department of Microbiology, Atmanand Saraswati Science College,

Surat, Gujarat, IndiaMONISHA KOTTAYI • School of Science, Navrachana University, Vadodara, Gujarat, IndiaV. MAGESHWARAN • ICAR-National Bureau of Agriculturally Important Microorganisms,

Mau, Uttar Pradesh, IndiaMAHADEVASWAMY • University of Agricultural Sciences, Raichur, Karnataka, IndiaNARAYANASAMY MARAPPA • Department of Microbiology, School of Life Sciences,

Bharathidasan University, Tiruchirappalli, Tamil Nadu, IndiaKIRAN MARATHE • School of Life Sciences, KBC North Maharashtra University, Jalgaon,

Maharashtra, IndiaVISHAL MARATHE • N.E.S. Science College, Nanded, Maharashtra, India

xi

VAIBHAVKUMAR MEHTA • Aspee Shakilam Biotechnology Institute, Navsari AgriculturalUniversity, Surat, Gujarat, India

HARSH MISTRY • Department of Life sciences, Hemchandracharya North GujaratUniversity, Patan, Gujarat, India

JEMISHA MISTRY • C. G. Bhakta Institute of Biotechnology, Uka Tarsadia University, Surat,Gujarat, India

BHAVANA V. MOHITE • Department of Microbiology, Bajaj College of Science, Wardha,Maharashtra, India

AMI NAIK • C. G. Bhakta Institute of Biotechnology, Uka Tarsadia University, Bardoli,Gujarat, India

VRUTUJA NAIK • National Centre for Microbial Resource, National Centre for Cell Science,Pune, India

PRADNYA B. NIKAM • School of Life Sciences, KBC North Maharashtra University, Jalgaon,Maharashtra, India

THAJUDDIN NOORUDDIN • Department of Microbiology, School of Life Sciences,Bharathidasan University, Tiruchirappalli, Tamil Nadu, India

DIXITA PANCHAL • C. G. Bhakta Institute of Biotechnology, Uka Tarsadia University, Surat,Gujarat, India

KETANKUMAR J. PANCHAL • Department of Animal Biotechnology, College of VeterinaryScience and Animal Husbandry, Anand Agricultural University, Anand, India

K. PANDIYAN • ICAR-Central Institute for Research on Cotton Technology, Mumbai, IndiaPREETI PARMAR • Main Cotton Research Station (MCRS), Navsari Agricultural University

(NAU), Surat, Gujarat, IndiaHARSH PATEL • P. G. Department of Biosciences, Sardar Patel University, Vallabh

Vidyanagar, Gujarat, IndiaKAMLESH C. PATEL • PG Department of Biosciences, Sardar Patel University, Anand,

Gujarat, IndiaNAFISA PATEL • Department of Microbiology, Naran Lala College of Professional and Applied

Sciences, Navsari, Gujarat, IndiaPOOJA PATEL • Government Medical College, Surat, Gujarat, IndiaPRITTESH PATEL • C. G. Bhakta Institute of Biotechnology, Uka Tarsadia University, Bardoli,

Gujarat, IndiaRADHIKA PATEL • PG Department of Biosciences, Sardar Patel University, Gujarat, IndiaSWATI PATEL • Aspee Shakilam Biotechnology Institute, Navsari Agricultural University,

Surat, Gujarat, IndiaTARUN KUMAR PATEL • Department of Biotechnology, Sant Guru Ghasidas Government

P.G. College, Dhamtari , Chattisgarh, India; Department of Biotechnology, Guru GhasidasVishwavidyalaya (a Central University), Bilaspur, Chattisgarh, India

CHANDRASHEKHAR D. PATIL • Department of Ophthalmology and Visual Sciences, Universityof Illinois at Chicago, Chicago, IL, USA

SATISH V. PATIL • School of Life Sciences, Kavayitri Bahinabai Chaudhari NorthMaharashtra University, Jalgaon, Maharashtra, India

VIKAS S. PATIL • University Institute of Chemical Technology, KBC North MaharashtraUniversity, Jalgaon, Maharashtra, India

MOHINI PIMPALSE • C.G. Bhakta Institute of Biotechnology, Uka Tarsadia University,Bardoli, Surat, India

VIMALKUMAR PRAJAPATI • Aspee Shakilam Biotechnology Institute, Navsari AgriculturalUniversity, Surat, Gujarat, India

xii Contributors

PRAVEEN RAHI • National Centre for Microbial Resource, National Centre for Cell Science,Pune, India

VIJAYAKUMAR VISHNU RAJA • Department of Microbiology, School of Life Sciences,Bharathidasan University, Tiruchirappalli, Tamil Nadu, India

B. K. RAJKUMAR • Main Cotton Research Station (MCRS), Navsari Agricultural University(NAU), Surat, Gujarat, India

SANKET RAY • Department of Microbiology, Naran Lala College of Professional and AppliedSciences, Navsari, Gujarat, India

NARENDRA SALUNKHE • School of Life Sciences, KBCNorth Maharashtra University, Jalgaon,Maharashtra, India

JITENDRA D. SALUNKHE • School of Life Sciences, KBC North Maharashtra University,Jalgaon, Maharashtra, India

A. SANKARANARAYANAN • C. G. Bhakta Institute of Biotechnology, Uka Tarsadia University,Surat, Gujarat, India

SHRUTI SATASHIA • Department of Microbiology, Atmanand Saraswati Science College,Surat, Gujarat, India

ABHISHEK SHARMA • C. G. Bhakta Institute of Biotechnology, Uka Tarsadia University,Surat, Gujarat, India

K. SOWMIYA • University of Agricultural Sciences, Raichur, Karnataka, IndiaRAHUL K. SURYAWANSHI • Department of Ophthalmology and Visual Sciences, University of

Illinois at Chicago, Chicago, IL, USARASHMI THAKOR • Department of Life Sciences, Hemchandracharya North Gujarat

University, Patan, Gujarat, IndiaVEYSEL TURAN • Department of Soil Science and Plant Nutrition, Faculty of Agriculture,

Bingol University, Bingol, TurkeyVRUSHALI WAGH • Department of Microbiology, Naran Lala College of Professional and

Applied Sciences, Navsari, Gujarat, India

Contributors xiii