Genetic diversity and pathogenicity of Xanthomonas axonopodis strains inducing citrus canker disease in Iran and South Korea

For the first time in 1989 citrus bacterial canker disease has seen on Citrus aurantiifolia in southern Iran. A total of 43 strains from affected citrus trees, ten strains from South Korea and representative from all known five pathotypes of Xanthomonas axonopodis pathogenic on citrus trees were used in this study. Isolated strains from Iran were indistinguishable by phenotypic, FAMEs, and SDS-PAGE analyses but showed different host range. First group were pathogenic on all tested citrus seedlings including C. aurantiifolia, C. limettioides, C. limon, C. jambhiri, Poncirus trifoliata X C. paradisi, C. aurantium, C. paradise, C. medica, P. trifoliate, C. grandis, C. sinensis, C. reticulate and C. sinensis X P. trifoliate. Pathogenicity of the second group were limited to C. aurantiifolia, C. limettioides, C. limon, C. jambhiri, P. trifoliata X C. paradis, and C. aurantium. Among the strains studied by AFLP fingerprinting six clusters were found. These clusters were: (1) strains of pathotype C; (2) strains of pathotypes B and D; (3) strains of pathotype A together with the main group of the Iranian strains; (4) strains isolated from Korea; (5) strains of pathotype E; and (6) seven strains from Iran which made a completely separate cluster. Strains from pathotypes B and D could not be differentiated by AFLP. The tested Iranian strains belongs to the two different groups and strains from Korea grouped as a subcluster from main cluster of Iranian strains belong to the pathotype A

Biological Control of Sclerotinia sclerotiorum the Causal Agent of Potato White Mold by Different Trichoderma spp. and Coniothyrium minitans

The identity of outbreak of stem drying and early death on potato in vast area of potato fields of Hamedan province was studied. As a result, the causal agent of the disease was isolated and identified as Sclerotinia sclerotiorum. Considering the serious damage of this pathogen that can not be easily controlled by cultural and chemical control methods, alternative approaches such as biocontrol method using five species of Trichoderma including T. ceramicum, T. koningii, T. koningiopsis, T. virens, T. viridescens and one isolate of Coniothyrium minitans were evaluated against this pathogen. All mentioned biocontrol agents caused some control on the pathogen in vitro. In assessment of the effect of volatile metabolites on pathogen growth rate, the most inhibition percent was realized about T. koningiopsis with 56% and in case of extracellular compounds, the most inhibition percent in concentrations of 15 and 30% were showed T. ceramicum as 69% and 53%, respectively. In dual culture, T. koningiopsis, T. viridescens and T. ceramicum prevented sclerotia formation as such. They also grew and sporulated on the pathogen mycelia. In assessment of the affect of spore suspension of the biological agents on myceliogenic germination of sclerotia, the most inhibition was observed in T. ceramicum and T. koningiopsis and the less inhibition was in T. virens and Coniothyrium minitans. T. ceramicum, T. koningiopsis and T. viridescens are recently described Trichoderma spp. and this study is the first assessment of their biocontrol effect on plant diseases in Iran

Assessment of Rice Associated Bacterial Ability to Enhance Rice Seed Germination and Rice Growth Promotion

ABSTRACT The application of beneficial bacteria has recently been used for sustainable agriculture. In current research, 71 bacterial isolates were obtained from rice plant and the rhizosphere soil of different paddy fields in Guilan province, Iran. After primitive investigation, 40 bacteria with typical predominant characteristics were selected. By PCR-RFLP of their 16S r-DNA gene, 8 Operational Taxonomic Units (OTUs) totally consisted of 33 isolates were obtained. From all of them, 8 isolates were selected for rice seed germination experiment, then, effective isolates were used for pot experiment to evaluate their ability for promoting rice growth. All of them were able to increase rice growth and yield, but in different potential. These tested isolates were identified as Alcaligenes faecalis (DEp8, O1R4), Pantoea ananatis (AEn1), Bacillus vietnamensis (MR5), Bacillus idriensis (MR2) and Stenotrophomonas maltophilia by partial sequencing of their 16S r-DNA gene. Among them, AEn1 and MR5 produced indole-3- acetic acid (IAA) in larger amounts than the other isolates and the isolates AEn1 and O1R4 were able to solubilize phosphate in higher amounts. According to the results obtained, it can be concluded that AEn1, O1R4 and MR5 can be considered as bacterial inoculants to use as alternatives for chemical fertilizers

Characterization of 16SrII group phytoplasmas associated with alfalfa (Medicago sativa) witches\u2019 broom disease in diverse areas of Iran.

Alfalfa witches\u2019 broom (AWB) is one of the most important alfalfa diseases in Iran. To characterize 16SrII group phytoplasmas associated with this disease, symptomatic and asymptomatic plants were collected during 2013-2015 and subjected to direct and nested polymerase chain reaction (PCR) using P1/P7, R16mF2/R16mR2 and R16F2n/R16R2. PCR amplicons of ~1.8, ~1.4 and ~1.25 kb respectively, were obtained only from all symptomatic plants. Restriction fragment length polymorphism (RFLP) analysis of R16F2n/R2 amplicons showed that the phytoplasma associated with AWB disease were members of 16SrII group subgroups 16SrII-D and -C. Blast analysis of these amplicon sequences and sequence homology of collected strains and strain sequences retrived from GenBank (AWB strains Chahgeer, Juyom and Bushehr) confirmed that AWB phytoplasmas collected from Bafg, Ardakan, Bahabad and Herat (Yazd province), Nikshahr (Sistan-Baluchestan), Bam, Zarand, Jiroft (Kerman province), Bushehr (Bushehr province), Tabas (South Khorasan province), Jowkar (Hamedan province) and Zardenjan (Esfahan province) cluster with phytoplasma strains enclosed in the 16SrII-D subgroup, while AWB strains from Chahgeer (Yazd province) and Juyom (Fars province) cluster with phytoplasma strains in the 16SrII-C subgroup. Based on these results the predominant strains of 16SrII phytoplasmas associated with AWB disease in Iran were classified in the 16SrII-D subgroup. In Ashkezar and Abarkouh in Yazd province entire alfalfa farm was infected with witches\u2019 broom disease. In 3 year alfalfa stands in Ashkezar alfalfa farms were plowed due to high incidence of the diseas

First report of 16SrVI-A and 16SrXII-A phytoplasmas associated with alfalfa witches\u2019 broom disease in Iran

In 2014-15 surveys, alfalfa witches\u2019 broom (AWB) disease was observed in a number of Iranian provinces. The main symptoms were crown proliferation, witches' broom, little leaf, flower virescence, phyllody, sterility and stunting. To investigate the phytoplasma presence, total DNAs were extracted from 132 symptomatic and 19 asymptomatic alfalfa plants and were tested by direct PCR using P1/P7 (Deng and Hiruki, 1991; Schneider et al., 1995) and nested PCR using R16F2n/R16R2 (Gundersen and Lee, 1996) primers. Amplicons were obtained only from all the symptomatic alfalfa plants and RFLP analyses with KpnI, AluI, HaeIII, HhaI, HpaII, MseI, RsaI, TaqI and HpaI enzymes showed the presence of two profiles. Mixed infections were never detected. From AWB samples collected in East Azarbayijan and Zanjan provinces, six amplicons were directly sequenced and showed 100% identity to each other (GenBank accession No. KU240021). The second profile was present in samples from Qom, Markazi, East Azarbayijan, Kordestan, Fars, Kermanshah, Kohgiluyeh and Boyer-Ahmad, Lorestan and Chaharamahal and Bakhtiari provinces; the 15 amplicons from Tabriz, Dehgolan, Arak, Qhorveh, and Shahrekian showed 100% identity in each area and formed 5 groups having 99% identity (GenBank accession Nos. KT763372, KT763371, KT781662, KT763373 and KT750060, respectively). The KU240021 strain sequence and its virtual RFLP indicated that it could be enclosed in the 16SrVI-A subgroup. The five-sequence group showed the highest homology with 16SrXII-A phytoplasma subgroup that was confirmed by virtual RFLP and phylogenetic analysis (MEGA 6.0). This is the first report of 16SrVI-A and 16SrXII-A phytoplasmas associated with AWB in Iran

Molecular identification and phylogenetic analysis of phytoplasmas associated with alfalfa witches’ broom diseases in the western areas of Iran.

During 2014-2015 phytoplasmas associated with alfalfa witches’ broom (AWB) in the western areas of Iran were characterized by molecular analyses studies. Polymerase chain reaction was carried out using direct and nested as says to amplify the 16S ribosomal gene for phytoplasma identification. RFLP analysis of R16F2n/R16R2 amplicons with selected restriction enzymes and phylogenetic studies showed that AWB associated phytoplasma strains from Salafchegan (Qom province), Arak (Markazi province), Tabriz (East Azerbaijan province), Dehgolan and Qhorveh (Kordestan province), Eghlid (Fars province), Mahidasht (Kermanshah province), Dehdasht (Kohgiluyeh and Boyer-Ahmad province), Borujerd (Lorestan province), Chenar-e Mahmudi and Shahrekian (Chaharmahal and Bakhtiari province) belong to 16SrXII-A subgroup (“stolbur”), while AWB associated phytoplasmas from Jowkar (Hamedan province) belong to 16SrII-D subgroup. Disease incidence was variable in the different areas and the highest infection rate (14.7%) was observed in Chenar-e Mahmudi (Chaharmahal and Bakhtiari province). The highly AWB affected areas were generally located in the warmer parts of each region in sparse fields. This is the first report and characterization of 16SrXII-A phytoplasmas associated with alfalfa witches’ broom diseases in Iran and occurrence of 16SrIID in western areas of Iran

Biologic, serologic and molecular characteristics of two 16SrII-C-related phytoplasma strains associated with alfalfa witches\u2019 broom disease in Yazd and Fars provinces, Iran

Alfalfa witches\u2019 broom (AWB) is a limiting factor for alfalfa growth and production in Iran, especially in the tropical regions of the country. AWB phytoplasma strains from two severely affected areas, Chahgeer (Abarkooh, Yazd province) and Juyom (Larestan, Fars province), were compared for main biologic, serologic and molecular characteristics. Based on disease symptoms in alfalfa farms, Chahgeer and Juyom AWB (CAWB and JAWB, respectively) strains were not differentiable. In dodder inoculated periwinkle and tomato plants JAWB phytoplasma induced stronger little leaf compared to the one induced by CAWB phytoplasma. In these experiments the two JAWB and CAWB phytoplasma strains are confirmed as vectored by different leafhopper species, Circulifer haematoceps and Orosius albicinctus respectively. Based on 16S rRNA gene and 16S-23S intergenic spacer region sequences, CAWB and JAWB were not differentiable, however no serologic relationship was observed between the two phytoplasmas in ELISA and DIBA tests using polyclonal antibodies prepared against each of them. Due to the lack of serological relationship, different insect vectors and induction of different symptoms in common host plants, CAWB and JAWB phytoplasmas should be considered as two AWB strains both belonging to 16SrII-C subgroup