MOLECULAR IDENTIFICATION AND  MANAGEMENT OF FUSARIUM WILT  PATHOGEN IN CHRYSANTHEMUM  (Dendranthema grandiflorum Tzelev)

N. UMALATHA

MOLECULAR IDENTIFICATION AND MANAGEMENT OF FUSARIUM WILT PATHOGEN IN CHRYSANTHEMUM (Dendranthema grandiflorum Tzelev)

Date

2019-07-08

Authors

N. UMALATHA

Publisher

COLLEGE OF HORTICULTURE ANANTHARAJUPETA - 516 105, Y.S.R KADAPA DISTRICT, ANDHRA PRADESH

Abstract

The present investigation entitled ʽʽMolecular identification and management of Fusarium wilt pathogen in chrysanthemum (Dendranthema grandiflorum Tzelev)ʼʼ was carried out at College of Horticulture, Anantharajupeta during the year 2018-2019. A roving survey was conducted to record Fusarium wilt incidence under field conditions in Kadapa, Chittoor, Ananthapuramu and Kurnool of Andhra Pradesh (A.P.). The maximum mean disease incidence was observed in Kadapa district (24.68%) followed by Chittoor district (19.85%), Ananthapuramu district (15.78%), whereas, minimum mean disease incidence was observed in Kurnool district (6.95%). The wilt pathogen of Fusarium oxysporum f. sp. chrysanthemi was isolated from collar region of infected chrysanthemum plant and their pathogenicity proved. Based on cultural and morphological characters, of isolates were identified as Fusarium oxysporum f. sp. chrysanthemi (NCFT, 9374.18) and confirmed by National Centre for Fungal Taxonomy (NCFT), New Delhi. Molecular characterization was conducted for Fusarium oxysporum f. sp. chrysanthemi. For identification of pathogen at molecular level, the genomic DNA was amplified with universal primes ITS 1 (TCCGTAGGTGAACCTGCGG) and ITS 4 (TCCTCCGCTTATTGATATGC). From the results obtained, the genomic DNA amplified at a 562 bp. Sequences were analysed through NCBI-BLAST programme database search system. BLAST (mega blast) analysis for sequence similarity of ITS rDNA region confirmed the identity of the pathogen. Sequence was deposited in NCBI, the Accession number (MK956193) for that sequence. Phylogenetic tree was constructed by using neighbour-joining method and maximum likelihood for the ITS regions. Results from the BLAST (Mega blast) data base showed, that the studied isolate have 98% similarity with Name of the author : N. UMALATHA Title of the thesis : ʽʽMolecular identification and management of Fusarium wilt pathogen in chrysanthemum (Dendranthema grandiflorum Tzelev)ʼʼ Degree to which it is submitted : M.Sc. (HORTICULTURE) Faculty Department : : HORTICULTURE PLANT PATHOLOGY Major Advisor : Dr. Ch. RUTH University : Dr.Y.S.R. HORTICULTURAL UNIVERSITY Year of submission : 2019 Fusarium oxysporum (Genbank KJ082096.1) and Fusarium sp. (Genbank ID - KU612374.) Rhizosphere antagonists (Bacteria, fungi and actinomycetes) were isolated from healthy rhizosphere soil samples collected from Kadapa, Chittoor, Ananthapuramu and Kurnool of Andhra Pradesh (A.P.). A total of 25 rhizosphere microbes were isolated. Among these, 20 isolates, seven fungi (RFA1 Phoma glomerata, RFA2 Aspergillus niger strain 1 RFA3 Aspergillus fumigates, RFA4 Aspergillus niger strain2 , RFA5 Aspergillus nidulans, RFA6 Aspergillus flavus strain 1, RFA7 Aspergillus flavus strain 2 ) three bacteria (RBA1 Bacillus cereus, RBA2 and RBA3) and three actinomycetes (RA1Streptomyces griseus, RA2 Streptomyces griseolus and RA3 Streptomyces griseoflavus)were found to exhibit antagonism against chrysanthemum wilt pathogen. Molecular identification of effective rhizosphere bacterial antagonists (RBA1) was conducted by the use of primers, 27F(AGAGTTTGATCMTGGCTCAG) and 1492R (TACGGYTACCTTGTTACGACTTS) the bacteria were identified as Bacillus cereus which have amplified at 874 base pairs. They were sequenced and the sequenced nucleotides were compared against Gen Bank database using the NCBI BLAST algorithm, the BLAST results shown the 88% similarity of the isolate (RBA 1) with Bacillus cereus. Among the rhizosphere antagonistic fungi, recorded highest inhibition over control, RFA 5- Aspergillus nidulans (84.68%), bacterial antagonist RBA 1- Bacillus cereus (65.88%) and rhizospheric actinomycetes RA1- Streptomyces griseus (70.74%) were more antagonistic against F.oxysporum f. sp. chrysanthemi respectively. An in vitro experiment was conducted on Fusarium oxysporum f. sp. chrysanthemi with five fungicides, viz., Copper oxy chloride 50% WDP, Carbendazim 12% + Mancozeb 64% WP, CuSO₄ + Lime + Water (Bordeaux mixture), Thiophanate methyl, Difenconazole 25% EC for their inhibitory effect. Among the fungicides, tested against pathogen Carbendazim + Mancozeb concentration (0.1% 0.2% 0.3%) and Difenconazole at (0.2%) showed 100 per cent inhibition against Fusarium wilt pathogen. The compatibility of the effective rhizosphere fungal antagonist Aspergillus nidulans (RFA 5) and effective rhizosphere bacterial antagonist Bacillus cereus (RBA1) to five fungicides viz., Copper oxy chloride 50% WDP, Carbendazim 12% + Mancozeb 64% WP, Bordeaux mixture, Thiophanate methyl, Difenconazole 25% EC were assessed. Copper oxychloride (0.3%) and Bordeaux mixture (0.5%) are more compatible with Aspergillus nidulans and Carbendazim 12% + Mancozeb 64% WP, Thiophanate methyl, Difenconazole 25% EC were compatible with Bacillus cereus. The most effective treatments proved effective under in vitro studies were tested against Fusarium oxysporum f. sp. chrysanthemi under pot culture conditions. Among the six treatments, recorded lowest disease incidence in the treatment T1- soil application of Streptomyces griseus (27.00%) and T2 - soil application of Streptomyces griseolus (27.25%) were the most effective antagonists respectively.

Description

➢ The indiscriminate use of harmful residual chemical pesticides and fungicides in disease management should be replaced with use of beneficial rhizosphere antagonists. ➢ The production and productivity of chrysanthemum has to be increased by proper utilization of PGPR. ➢ The antagonistic activity of rhizosphere microbes against the Fusarium wilt pathogen must be tested in large scale under field conditions. ➢ The compatibility of beneficial rhizosphere antagonists should be tested with fungicides, insecticides, herbicides and botanicals at field level. ➢ There is a need for development of wilt resistant chrysanthemum varieties.