Molecular Characterization of Indigenous Bacillus thuringiensis Strains Possessing Nematode-Active cry Genes
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Date
2024
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MPUAT, Udaipur
Abstract
Plant parasitic nematodes (PPNs) are significant phyto-parasites responsible
for severe crop damage and yield losses. The toxicological and environmental risks
associated with the use of chemicals motivate the adoption of more eco-friendly
alternatives such as the use of biological agents as biocontrol agents to control PPNs
is gaining momentum in recent times. Bacillus thuringiensis (Bt) possessing
nematicidal crystal proteins, is being used widely to mitigate nematode infestation in
agricultural crops. In the present study the potential of the native Bacillus
thuringiensis strains was investigated against the root knot nematode. Morphological
characterization of Bacillus thuringiensis (Bt) strains revealed that all Bt strains were
Gram positive and endospore forming. The 20 Bt strains was screened for the
presence of their nematicidal cry genes by PCR and 12 Bt strains viz., Bt1, Bt5, Bt6,
Bt7, Bt9, Bt10, Bt16, Bt17, Bt19, Bt23, Bt24 and Bt36 showed the presence of
nematicidal cry genes viz., cry5, cry6, cry12, cry13, cry14, cry21, cry55 and cry31Aa
by gene specific primers. The SDS-PAGE studies of spore crystal mixture revealed
different sizes of protein bands viz., 135, 90, 79, 75, 65, 59, 54, 45, and 30 kDa which
represent the presence of different Cry proteins including the nematicidal Cry
proteins. The in-vitro efficacy of the Bt strains were performed against Meloidogyne
incognita using cavity block assay and the Bt strains viz., Bt7, Bt17, and Bt19
completely inhibited the hatching of Meloidogyne incognita eggs & egg masses and
lethal to nematode larvae (J2 stage). In pot studies, Bt7 and Bt19 strains exhibited
highest gall suppuration ability and depicted significantly higher PGP- attributes and
are most effective in biological control of nematodes. The molecular characterization
of potent Bt strains viz., Bt-7, and Bt-19 based on 16S rDNA sequencing revealed
their molecular identity as Bacillus thuringiensis.
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Paramjeet and Jain D.