Rural do Semi-Árido

Nodules in roots of Anadenanthera colubrina.

The study of native species from Caatinga and the Biological Nitrogen Fixation (BNF) promoted by bacteria generally known as rhizobia have major relevance when “reforestation” is in question, mainly due to the role of nitrogen on vegetal growth, which is appears in energy molecules (ATP, NADH, and NADPH), chlorophyll, proteins, and enzymes. These bacteria, when associated to plant roots, especially leguminous plants, form nodules and have the capacity to fix atmospheric nitrogen (N₂), which plants are unable to directly absorb, and provides it as nitrate and ammonium (NO₃^– and NH₄⁺). From all native species of Caatinga, only plants from the Fabaceae botany family, such as Angico (Anadenanthera colubrina), Catanduva (Pytirocarpa moniliformis), Jurema Branca (Piptadenia stipulacea), Sabiá (Mimosa caesalpiniifolia) and Timbaúba (Enterolobium contortisiliquum), are capable of forming such symbiosis.

The Table bellow shows some of the Native species studied within the Projeto Caatinga, classified as a nodulating or non-nodulating species:

RHIZOBIA ISOLATION

When studying nitrogen-fixing bacteria, or rhizobia, it is necessary to isolate them for posterior inoculation in plants from Fabaceae botany family and verify its efficiency, since these microorganisms form a symbiotic relationship, fixing atmospheric nitrogen for the plants and, consequently, improving their growth.

The process of isolation begins with roots being washed in running water. After cleaning roots, nodules are detached, dried out in a filter paper, and disinfested in alcohol (70%) and sodium hypochlorite (1%). Sterile distilled water is used to wash nodules after disinfestation. One single nodule is placed in a test tube filled with sterile distilled water (no more than 3 mL) and crushed with a sterile glass rod. The water suspension containing bacteria (extracted from the nodule) is then transferred to Petri dishes filled with Yeast Mannitol Agar (YMA) (a specific culture medium for the cultivation of rhizobia) using a bacteriological loop. Plates are stored in BOD (Biological Oxygen Demand) type incubators (MACHADO et al., 2006) for bacterial growth.

Before the isolation, it is important to confirm if nodules are active. This confirmation may be done visually by the cutting the nodule in two pieces and observation of a reddish color inside. This tone is seen when a protein known as Leghemoglobin, responsible for oxygen transportation required for respiration in the nodule, is present. The protein is important for the Nitrogenase, the enzyme that catalyzes the biological fixation of atmospheric nitrogen (N₂) into Ammonium (NH₄⁺), inactivated in the presence of O₂. This way, oxygen transportation – performed by Leghemoglobin – does not inhibit the activity of nitrogenase complex.

PUBLICATIONS

Efeito da inoculação de rizóbios sobre a germinação e vigor de plântulas de Mimosa tenuiflora Willd.

• Authors: Paulo César da Silva Santos, Tatianne Raianne Costa Alves, Afonso Luiz Almeida Freires, Luan Vítor Nascimento, Márcia Michelle de Queiroz Ambrósio
• Event: III Simpósio de Ciências e Tecnologia Agroalimentar (III SIMTAGRO, 2017)
• Language: Portuguese
• Access: Download here

Efeito da inoculação de rizóbios e adubação nitrogenada no desenvolvimento de mudas de Mimosa tenuiflora Willd.

• Authors: Afonso Luiz Almeida Freires, Márcia Michelle de Queiroz Ambrósio, Luan Vítor Nascimento, Tatianne Raianne Costa Alves, Paulo César da Silva Santos
• Event: XXIV Seminário de Iniciação Científica da UFERSA (XXIV SEMIC, 2018)
• Language: Portuguese
• Access: Download here

Caracterização morfofisiológica de rizóbio de Piptadenia stipulacea e a resposta da espécie à adubação nitrogenada e à inoculação com rizóbio

• Authors: Maria Ianca Dias Oliveira, Márcia Michelle de Queiroz Ambrósio, Luan Vítor Nascimento, Afonso Luiz Almeida Freires, Tatianne Raianne Costa Alves
• Event: XXIV Seminário de Iniciação Científica da UFERSA (XXIC SEMIC, 2018)
• Language: Portuguese
• Access: Download here

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REFERENCES