Displaying publications 1 - 20 of 26 in total

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  1. Hossain MI, Ahmad K, Vadamalai G, Siddiqui Y, Saad N, Ahmed OH, et al.
    Biology (Basel), 2021 Sep 03;10(9).
    PMID: 34571739 DOI: 10.3390/biology10090862
    Colletotrichum falcatum Went causes red rot disease in sugarcane farming in the tropical and sub-tropical regions. This disease causes significant economic loss to the sugarcane production industry. Successful disease management strategies depend on understanding the evolutionary relationship between pathogens, genetic diversity, and population structure, particularly at the intra-specific level. Forty-one isolates of C. falcatum were collected from different sugarcane farms across Bangladesh for molecular identification, phylogeny and genetic diversity study. The four genes namely, ITS-rDNA, β-tubulin, Actin and GAPDH sequences were conducted. All the 41 C. falcatum isolates showed a 99-100% similarity index to the conserved gene sequences in the GenBank database. The phylogram of the four genes revealed that C. falcatum isolates of Bangladesh clustered in the same clade and no distinct geographical structuring were evident within the clade. The four gene sequences revealed that C. falcatum isolates from Bangladesh differed from other countries´ isolates because of nucleotides substitution at different loci. The genetic structure of C. falcatum isolates were determined using ISSR marker generated 404 polymorphic loci from 10 selected markers. The percentage of polymorphic loci was 99.01. The genetic variability at species level was slightly higher than at population level. Total mean gene diversity at the species level was 0.1732 whereas at population level it was 0.1521. The cluster analysis divided 41 isolates into four main genetic groups and the principal component analysis was consistent with cluster analysis. To the best of our knowledge, this is the first finding on characterizing C. falcatum isolates infesting sugarcane in Bangladesh. The results of this present study provide important baseline information vis a vis C. falcatum phylogeny analysis and genetic diversity study.
  2. Sivaprakasam S, Mohd Azim Khan NA, Yee Fan T, Kumarasan Y, Sicheritz-Pontén T, Petersen B, et al.
    Microbiol Resour Announc, 2024 Apr 11;13(4):e0113723.
    PMID: 38506531 DOI: 10.1128/mra.01137-23
    Here, we present the complete genome of a plant growth-promoting strain, Bacillus stratosphericus AIMST-CREST02 isolated from the bulk soil of a high-yielding paddy plot. The genome is 3,840,451 bp in size with a GC content of 41.25%. Annotation predicted the presence of 3,907 coding sequences, including genes involved in auxin biosynthesis regulation and gamma-aminobutyric acid (GABA) metabolism.
  3. Naderali N, Nejat N, Vadamalai G, Davis RE, Wei W, Harrison NA, et al.
    Int J Syst Evol Microbiol, 2017 Oct;67(10):3765-3772.
    PMID: 28905707 DOI: 10.1099/ijsem.0.002187
    Landscape-grown foxtail palm (Wodyetia bifurcata A. K. Irvine) trees displaying symptoms of severe foliar chlorosis, stunting, general decline and mortality reminiscent of coconut yellow decline disease were observed in Bangi, Malaysia, during 2012. DNA samples from foliage tissues of 15 symptomatic palms were analysed by employing a nested PCR assay primed by phytoplasma universal ribosomal RNA operon primer pairs, P1/P7 followed by R16F2n/R2. The assay yielded amplicons of a single band of 1.25 kb from DNA samples of 11 symptomatic palms. Results from cloning and sequence analysis of the PCR-amplified 16S rRNA gene segments revealed that, in three palms, three mutually distinct phytoplasmas comprising strains related to 'Candidatus Phytoplasma asteris' and 'Candidatus Phytoplasma cynodontis', as well as a novel phytoplasma, were present as triple infections. The 16S rRNA gene sequence derived from the novel phytoplasma shared less than 96 % nucleotide sequence identity with that of each previously describedspecies of the provisional genus 'Ca. Phytoplasma', justifying its recognition as the reference strain of a new taxon, 'Candidatus Phytoplasma wodyetiae'. Virtual RFLP profiles of the R16F2n/R2 portion of the 16S rRNA gene and the pattern similarity coefficient value (0.74) supported the delineation of 'Ca. Phytoplasma wodyetiae' as the sole representative subgroup A member of a new phytoplasma ribosomal group, 16SrXXXVI.
  4. Lim FH, Rasid OA, Idris AS, As'wad AWM, Vadamalai G, Parveez GKA, et al.
    Mol Biol Rep, 2023 Mar;50(3):2367-2379.
    PMID: 36580194 DOI: 10.1007/s11033-022-08131-4
    BACKGROUND: The basidiomycete fungus, Ganoderma boninense is the main contributor to oil palm Basal Stem Rot (BSR) in Malaysia and Indonesia. Lanosterol 14α-Demethylase (ERG11) is a key enzyme involved in biosynthesis of ergosterol, which is an important component in the fungal cell membrane. The Azole group fungicides are effective against pathogenic fungi including G. boninense by inhibiting the ERG11 activity. However, the work on molecular characterization of G. boninense ERG11 is still unavailable today.

    METHODS AND RESULTS: This study aimed to isolate and characterize the full-length cDNA encoding ERG11 from G. boninense. The G. boninense ERG11 gene expression during interaction with oil palm was also studied. A full-length 1860 bp cDNA encoding ERG11 was successfully isolated from G. boninense. The G. boninense ERG11 shared 91% similarity to ERG11 from other basidiomycete fungi. The protein structure homology modeling of GbERG11 was analyzed using the SWISS-MODEL workspace. Southern blot and genome data analyses showed that there is only a single copy of ERG11 gene in the G. boninense genome. Based on the in-vitro inoculation study, the ERG11 gene expression in G. boninense has shown almost 2-fold upregulation with the presence of oil palm.

    CONCLUSION: This study provided molecular information and characterization study on the G. boninense ERG11 and this knowledge could be used to design effective control measures to tackle the BSR disease of oil palm.

  5. Lim FH, Rasid OA, Idris AS, As'wad AWM, Vadamalai G, Parveez GKA, et al.
    Folia Microbiol (Praha), 2021 Aug;66(4):677-688.
    PMID: 34041694 DOI: 10.1007/s12223-021-00852-6
    The basidiomycete fungus, Ganoderma boninense, has been identified as the main causal agent of oil palm basal stem rot (BSR) disease which has caused significant economic losses to the industry especially in Malaysia and Indonesia. Various efforts have been initiated to understand the disease and this plant pathogen especially at the molecular level. This is the first study of its kind on the development of a polyethylene glycol (PEG)-mediated protoplast transformation system for G. boninense. Based on the minimal inhibitory concentration study, 60 µg/mL and above of hygromycin were effective to completely inhibit G. boninense growth. Approximately 5.145 × 107 cells/mL of protoplasts with the viability of 97.24% was successfully obtained from G. boninense mycelium tissue. The PEG-mediated G. boninense protoplast transformation using 1 µg of transformation vector, 25% of PEG solution, 10 min of pre-transformation incubation, and 30 min of post-transformation incubation has improved the transformation rate as compared with the previous reported protocols for other basidiomycete fungi. Optimization of four transformation parameters has improved the transformation efficiency of G. boninense from an average of 2 to 67 putative transformants. The presence of hygromycin phosphotransferase (hpt) and enhanced green fluorescent protein (eGFP) genes in the putative transformants was detected by PCR and verified by gene sequence analysis. Southern hybridization result further confirmed the integration of hpt gene in G. boninense transformants, and the green fluorescent signal was detected in the G. boninense transformants under the microscopic analysis. The establishment of this transformation system will accelerate the gene function studies of G. boninense especially those genes that may contribute to the pathogenesis of this fungus in oil palm.
  6. Abidin N, Ismail SI, Vadamalai G, Yusof MT, Hakiman M, Karam DS, et al.
    PLoS One, 2020;15(6):e0234350.
    PMID: 32530926 DOI: 10.1371/journal.pone.0234350
    Jackfruit-bronzing is caused by bacteria Pantoea stewartii subspecies stewartii (P. stewartii subsp. stewartii), showing symptoms of yellowish-orange to reddish discolouration and rusty specks on pulps and rags of jackfruit. Twenty-eight pure bacterial strains were collected from four different jackfruit outbreak collection areas in Peninsular Malaysia (Jenderam, Maran, Muadzam Shah and Ipoh). Positive P. stewartii subsp. stewartii verification obtained in the study was based on the phenotypic, hypersensitivity, pathogenicity and molecular tests. Multilocus sequence analysis (MLSA) was performed using four housekeeping genes (gyrB, rpoB, atpD and infB) on all 28 bacterial strains. Single gyrB, rpoB, atpD and infB phylogenetic trees analyses revealed the bootstrap value of 99-100% between our bacterial strains with P. stewartii subsp. stewartii reference strains and P. stewartii subsp. indologenes reference strains. On the other hand, phylogenetic tree of the concatenated sequences of the four housekeeping genes revealed that our 28 bacterial strains were more closely related to P. stewartii subsp. stewartii (99% similarities) compared to its close relative P. stewartii subsp. indologenes, although sequence similarity between these two subspecies were up to 100%. All the strains collected from the four collection areas clustered together, pointing to no variation among the bacterial strains. This study improves our understanding and provided new insight on the genetic diversity of P. stewartii subsp. stewartii associated with jackfruit-bronzing in Malaysia.
  7. Nejat N, Vadamalai G, Davis RE, Harrison NA, Sijam K, Dickinson M, et al.
    Int J Syst Evol Microbiol, 2013 Feb;63(Pt 2):540-548.
    PMID: 22523165 DOI: 10.1099/ijs.0.041467-0
    This study addressed the taxonomic position and group classification of a phytoplasma responsible for virescence and phyllody symptoms in naturally diseased Madagascar periwinkle plants in western Malaysia. Unique regions in the 16S rRNA gene from the Malaysian periwinkle virescence (MaPV) phytoplasma distinguished the phytoplasma from all previously described 'Candidatus Phytoplasma' species. Pairwise sequence similarity scores, calculated through alignment of full-length 16S rRNA gene sequences, revealed that the MaPV phytoplasma 16S rRNA gene shared 96.5 % or less sequence similarity with that of previously described 'Ca. Phytoplasma' species, justifying the recognition of the MaPV phytoplasma as a reference strain of a novel taxon, 'Candidatus Phytoplasma malaysianum'. The 16S rRNA gene F2nR2 fragment from the MaPV phytoplasma exhibited a distinct restriction fragment length polymorphism (RFLP) profile and the pattern similarity coefficient values were lower than 0.85 with representative phytoplasmas classified in any of the 31 previously delineated 16Sr groups; therefore, the MaPV phytoplasma was designated a member of a new 16Sr group, 16SrXXXII. Phytoplasmas affiliated with this novel taxon and the new group included diverse strains infecting periwinkle, coconut palm and oil palm in Malaysia. Three phytoplasmas were characterized as representatives of three distinct subgroups, 16SrXXXII-A, 16SrXXXII-B and 16SrXXXII-C, respectively.
  8. Wu YH, Cheong LC, Meon S, Lau WH, Kong LL, Joseph H, et al.
    Arch Virol, 2013 Jun;158(6):1407-10.
    PMID: 23397332 DOI: 10.1007/s00705-013-1624-8
    A 246-nt variant of Coconut cadang-cadang viroid (CCCVd) has been identified and described from oil palms with orange spotting symptoms in Malaysia. Compared with the 246-nt form of CCCVd from coconut, the oil palm variant substituted C(31)→U in the pathogenicity domain and G(70)→C in the central conserved domain. This is the first sequence reported for a 246-nt variant of CCCVd in oil palms expressing orange spotting symptoms.
  9. Nejat N, Cahill DM, Vadamalai G, Ziemann M, Rookes J, Naderali N
    Mol Genet Genomics, 2015 Oct;290(5):1899-910.
    PMID: 25893418 DOI: 10.1007/s00438-015-1046-2
    Invasive phytoplasmas wreak havoc on coconut palms worldwide, leading to high loss of income, food insecurity and extreme poverty of farmers in producing countries. Phytoplasmas as strictly biotrophic insect-transmitted bacterial pathogens instigate distinct changes in developmental processes and defence responses of the infected plants and manipulate plants to their own advantage; however, little is known about the cellular and molecular mechanisms underlying host-phytoplasma interactions. Further, phytoplasma-mediated transcriptional alterations in coconut palm genes have not yet been identified. This study evaluated the whole transcriptome profiles of naturally infected leaves of Cocos nucifera ecotype Malayan Red Dwarf in response to yellow decline phytoplasma from group 16SrXIV, using RNA-Seq technique. Transcriptomics-based analysis reported here identified genes involved in coconut innate immunity. The number of down-regulated genes in response to phytoplasma infection exceeded the number of genes up-regulated. Of the 39,873 differentially expressed unigenes, 21,860 unigenes were suppressed and 18,013 were induced following infection. Comparative analysis revealed that genes associated with defence signalling against biotic stimuli were significantly overexpressed in phytoplasma-infected leaves versus healthy coconut leaves. Genes involving cell rescue and defence, cellular transport, oxidative stress, hormone stimulus and metabolism, photosynthesis reduction, transcription and biosynthesis of secondary metabolites were differentially represented. Our transcriptome analysis unveiled a core set of genes associated with defence of coconut in response to phytoplasma attack, although several novel defence response candidate genes with unknown function have also been identified. This study constitutes valuable sequence resource for uncovering the resistance genes and/or susceptibility genes which can be used as genetic tools in disease resistance breeding.
  10. Iftikhar Y, Khoo YW, Murugan T, Roslin NA, Adawiyah R, Kong LL, et al.
    Mol Biol Rep, 2022 Feb;49(2):1581-1586.
    PMID: 34773552 DOI: 10.1007/s11033-021-06930-9
    BACKGROUND: A 328-nucleotide variant of citrus bent leaf viroid (CBLVd) was characterized by citrus varieties in Malaysia. After the first report in Malaysia, the emerging CBLVd was detected in five citrus species, namely Citrofortunella microcarpa, Citrus aurantifolia, Citrus hystrix, Citrus maxima, and Citrus sinensis.

    METHODS AND RESULTS: CBLVd was detected in 23 out of 133 symptomatic samples through RT-PCR. Sequence analysis of the RT-PCR amplicons from this study showed 99-100% sequence identity to the reference CBLVd Jp isolate and CBLVd isolates reported in Malaysia. Inoculation of sap, obtained from a CBLVd positive sample, into 6-month old healthy C. microcarpa seedlings showed symptoms of slight leaf bending, reduced leaf size of matured leaves, and mild mosaic between 4 to 6 months after inoculation. Moreover, the observed symptoms of chlorosis, midvein necrosis, leaf rolling, and smalling of leaves in calamondin, C. microcarpa (Bunge) Wijnands, were not reported in earlier studies and opened a new avenue for the study of symptomology. The mechanical transmissibility of CBLVd in the inoculated seedlings was reconfirmed by RT-PCR assay and sequencing.

    CONCLUSIONS: Based on the results, the sequence similarity of CBLVd isolates from different areas of Malaysia showed no significant difference among each other and the reference isolate. The CBLVd is mechanically transmissible and could produce variable symptoms in different hosts.

  11. Mohd Azim Khan NA, Jaya Jothi S, Mohd Hata E, Parimannan S, Vadamalai G, Rajandas H
    Microbiol Resour Announc, 2022 Feb 17;11(2):e0115621.
    PMID: 35084217 DOI: 10.1128/mra.01156-21
    We characterized the draft genome of the potentially beneficial Bacillus tropicus strain UPM-CREST01, which was isolated from the bulk soil at a paddy cultivation area in Kampung Gajah, Perak, Malaysia. The final draft assembly of 5,252,705 bp, with a G+C content of 35.23%, was found to harbor 5,368 coding sequences, including several plant-growth-promoting genes.
  12. Thanarajoo SS, Kong LL, Kadir J, Lau WH, Vadamalai G
    J Virol Methods, 2014 Jun;202:19-23.
    PMID: 24631346 DOI: 10.1016/j.jviromet.2014.02.024
    A reverse transcription loop-mediated isothermal amplification (RT-LAMP) detected Coconut cadang-cadang viroid (CCCVd) within 60 min at 60 °C in total nucleic acid extracted from oil palm leaves infected with CCCVd. Positive reactions showed colour change from orange to green in the reaction mix after the addition of fluorescent reagent, and a laddering pattern band on 2% agarose gel electrophoresis. Conventional RT-PCR with LAMP primers produced amplicons with a sequence identical to the 297-nt CCCVd oil palm variant with the primers being specific for CCCVd and not for other viroids such as PSTVd and CEVd. RT-LAMP was found to be rapid and specific for detecting oil palm CCCVd.
  13. Roslan ND, Sundram S, Hong LW, Ling KL, Vadamalai G
    Mol Biol Rep, 2023 Nov;50(11):9699-9705.
    PMID: 37676433 DOI: 10.1007/s11033-023-08771-0
    BACKGROUND: Sequence variation has been attributed to symptom variations but has not been investigated in Orange Spotting-Coconut cadang-cadang viroid (OS-CCCVd) infected palms. Likewise, the relationship between Coconut cadang-cadang viroid (CCCVd) variants, Orange Spotting (OS) severity and the accumulation of the viroid in the palms have not been elucidated. This paper describes the characterization of CCCVd variants by cloning and sequencing, followed by correlation with symptom expression.

    METHODS AND RESULTS: Total nucleic acids were extracted from leaf samples harvested from frond 20 of seven Dura × Pisifera (D × P) African oil palm (Elaeis guineensis Jacq.) aged between 13 and 21 years old collected from local plantations. The nucleic acids were fractionated using 5% non-denaturing polyacrylamide gel electrophoresis (PAGE) before being subjected to detection by reverse transcribed polymerase chain reaction (RT-PCR). The PCR products were cloned into a plasmid vector and the sequence of the clones was analyzed. CCCVd variants were quantified using real-time qPCR assay with CCCVd specific primers. Sixteen randomly selected clones of (OP246) had an arbitrary 100% identity with CCCVdOP246 (GeneBank Accession No: HQ608513). Meanwhile, four clones had >93% similarity with several minor sequence variations forming variants of OP234, OP235, OP251 and OP279.

    CONCLUSION: The OS symptoms observed in the field were characterized into three categories based on the size and morphology of the orange spots on the affected fronds. In addition, there was no direct correlation between disease severity and the accumulation of CCCVd variants in oil palm. This finding is the first report describing the sequence variation of the CCCVd RNA and symptom variation in OS oil palm field samples.

  14. Wong CKF, Saidi NB, Vadamalai G, Teh CY, Zulperi D
    J Appl Microbiol, 2019 Aug;127(2):544-555.
    PMID: 31077517 DOI: 10.1111/jam.14310
    AIMS: This study sought to investigate the effect of bioformulation on the biocontrol efficacy, microbial viability and storage stability of a consortium of Pseudomonas aeruginosa DRB1 and Trichoderma harzianum CBF2 against Foc Tropical Race 4 (Foc-TR4).

    MATERIALS AND RESULTS: Four bioformulations consisting of dry (pesta granules, talc powder and alginate beads) and liquid formulations were evaluated for their ability to control Foc-TR4, sustain microbial populations after application and maintain microbial stability during storage. All tested bioformulations reduced disease severity (DS) by more than 43·00% with pesta granules producing the highest reduction in DS by 66·67% and the lowest area under the disease progress curve value (468·75) in a glasshouse trial. Microbial populations of DRB1 and CBF2 were abundant in the rhizosphere, rhizoplane and within the roots of bananas after pesta granules application as compared to talc powder, alginate beads and liquid formulations 84 days after inoculation (DAI). The stability of both microbial populations after 180 days of storage at 4°C was the greatest in the pesta granule formulation.

    CONCLUSION: The pesta granule formulation was a suitable carrier of biological control agents (BCA) without compromising biocontrol efficacy, microbial population and storage stability as compared to other bioformulations used in this study.

    SIGNIFICANCE AND IMPACT OF THE STUDY: Pesta granules could be utilized to formulate BCA consortia into biofertilizers. This formulation could be further investigated for possible applications under agricultural field settings.

  15. Anggraini E, Vadamalai G, Kong LL, Mat M, Lau WH
    Microbiol Resour Announc, 2023 Mar 16;12(3):e0001923.
    PMID: 36853043 DOI: 10.1128/mra.00019-23
    Two members of the species Oryctes rhinoceros nudivirus (OrNV) were detected in O. rhinoceros haplotype G beetles collected from an oil palm plantation in Kluang and a wild coconut tree in Batu Pahat (Johor, Malaysia). OrNV strain Kluang comprised 125,794 bp, encoding 125 open reading frames (ORFs), while OrNV strain Batu Pahat comprised 124,925 bp, encoding 126 ORFs.
  16. Anggraini E, Vadamalai G, Kong LL, Mat M, Lau WH
    Sci Rep, 2023 Oct 06;13(1):16850.
    PMID: 37803044 DOI: 10.1038/s41598-023-43691-w
    The CRB (coconut rhinoceros beetle) haplotype was classified into CRB-S and CRB-G, based on the presence of single nucleotide polymorphisms (SNPs) in the mitochondrial cox1 gene. Mitochondrial genomes (mitogenomes) are the most widely used genetic resources for molecular evolution, phylogenetics, and population genetics in relation to insects. This study presents the mitogenome CRB-G and CRB-S which were collected in Johor, Malaysia. The mitogenome of CRB-G collected from oil palm plantations in 2020 and 2021, and wild coconut palms in 2021 was 15,315 bp, 15,475 bp, and 17,275 bp, respectively. The CRB-S was discovered in coconut and oil palms in 2021, and its mitogenome was 15,484 bp and 17,142 bp, respectively. All the mitogenomes have 37 genes with more than 99% nucleotide sequence homology, except the CRB-G haplotype collected from oil palm in 2021 with 89.24% nucleotide sequence homology. The mitogenome of Johor CRBs was variable in the natural population due to its elevated mutation rate. Substitutions and indels in cox1, cox2, nad2 and atp6 genes were able to distinguish the Johor CRBs into two haplotypes. The mitogenome data generated in the present study may provide baseline information to study the infection and relationship between the two haplotypes of Johor CRB and OrNV in the field. This study is the first report on the mitogenomes of mixed haplotypes of CRB in the field.
  17. Wong CKF, Zulperi D, Saidi NB, Vadamalai G
    Trop Life Sci Res, 2021 Mar;32(1):23-45.
    PMID: 33936549 DOI: 10.21315/tlsr2021.32.1.2
    Fusarium wilt of banana cannot be effectively controlled by current control strategies. The most virulent form that caused major losses in the banana production is Fusarium oxysporum f. sp. cubense Tropical Race 4 (Foc-TR4). Biocontrol of Foc-TR4 using microbial antagonists offers a sustainable and eco-friendly alternative. A consortium of biocontrol agents (BCAs), Pseudomonas aeruginosa DRB1 and Trichoderma harzianum CBF2 was formulated into pesta granules, talc powder, alginate beads and liquid bioformulations. Previous study indicated bioformulations containing both BCAs successfully reduced the disease severity of Foc-TR4. To date, the biocontrol mechanism and plant growth promoting (PGP) traits of a consortium of BCAs on infected bananas have not been explored. Therefore, the study was undertaken to investigate the effect of a consortium of DRB1 and CBF2 in the growth and biochemical changes of Foc-TR4 infected bananas. Results indicated pesta granules formulation produced bananas with higher biomass (fresh weight: 388.67 g), taller plants (80.95 cm) and larger leaves (length: 39.40 cm, width: 17.70 cm) than other bioformulations. Applying bioformulations generally produced plants with higher chlorophyll (392.59 μg/g FW-699.88 μg/g FW) and carotenoid contents (81.30 μg/g FW-120.01 μg/g FW) compared to pathogen treatment (chlorophyll: 325.96 μg/g FW, carotenoid: 71.98 μg/g FW) which indicated improved vegetative growth. Bioformulation-treated plants showed higher phenolic (49.58-93.85 μg/g FW) and proline contents (54.63 μg/g FW-89.61 μg/g FW) than Foc-TR4 treatment (phenolic: 46.45 μg/g FW, proline: 28.65 μg/g FW). The malondialdehylde (MDA) content was lower in bioformulation treatments (0.49 Nm/g FW-1.19 Nm/g FW) than Foc-TR4 treatment (3.66 Nm/g FW). The biochemical changes revealed that applying bioformulations has induced host defense response by increasing phenolic and proline contents which reduced root damage caused by Foc-TR4 resulting in lower MDA content. In conclusion, applying bioformulations containing microbial consortium is a promising method to improve growth and induce significant biochemical changes in bananas leading to the suppression of Foc-TR4.
  18. Vadamalai G, Hanold D, Rezaian MA, Randles JW
    Arch Virol, 2006 Jul;151(7):1447-56.
    PMID: 16470341
    Variants of Coconut cadang-cadang viroid have been identified in a plantation oil palm growing in Malaysia. Three size classes are described, comprising 297, 293, and 270 nt. Compared with the 296-nt form of coconut cadang-cadang viroid (CCCVd), all variants substituted C31 --> U in the pathogenicity domain and A175 --> C in the right-hand terminus. Other mutations and deletions accounted for the different sizes. These are the first sequences reported for variants of Coconut cadang-cadang viroid in a species other than coconut palm, and this is the first evidence that variants closely related to CCCVd occur outside the Philippines.
  19. Nazerian E, Sijam K, Zainal Abidin MA, Vadamalai G
    Plant Dis, 2011 Nov;95(11):1474.
    PMID: 30731752 DOI: 10.1094/PDIS-10-10-0754
    Cucumber (Cucumis sativus L.) is one of the most important vegetable fruits in Malaysia. Cucumber is principally grown in the states of Johor, Kelantan, and Perak. The broad host range Enterobacteriaceae pathogen, Pectobacterium carotovorum, can cause soft rot on stems or cucumber fruit. In Malaysia, cucumber is produced in a warm, humid climate, thus the plant is susceptible to attack by P. carotovorum at any time during production. In 2010, cucumber samples with wilted and chlorotic leaves, water-soaked lesions, and collapsed fruits were found in multiple fields. Small pieces of infected stems and fruit were immersed in 5 ml of saline solution (0.85% NaCl) for 20 min and then 50 μl of this suspension was spread onto nutrient agar (NA) and incubated at 27°C for 24 h. White-to-pale gray colonies with irregular margins were selected for analysis. For pathogenicity tests, cucumber fruits were surface sterilized by ethyl alcohol 70%, washed with sterilized distilled water, cut into small pieces, and inoculated with 20 μl of 108 CFU/ml suspensions of five representative strains. Cucumber plants were grown for 3 weeks in sterilized soil and their stems were inoculated with 20 μl of 108 CFU/ml of bacterial suspension. Inoculated samples and control (noninoculated) plants were placed in a growth chamber with 80 to 90% relative humidity at 27°C. Symptoms occurred on fruit slices and stems after 1 to 3 days and appeared the same as naturally infected samples, but the control samples remained healthy. Koch's postulates were fulfilled with the reisolation of cultures with the same characteristics as described earlier. Hypersensitivity reaction (HR) assays were done by infiltrating 108 CFU/ml of bacterial suspension into tobacco leaf epidermis and HR developed. All strains were subjected to biochemical and morphological assays, as well as molecular assessment. The strains were gram negative, facultative anaerobes, rod shaped, able to macerate potato slices and growth at 37°C; catalase positive; oxidase and phosphatase negative; able to degrade pectate; sensitive to erythromycin; negative for utilization of α-methyl glycoside, indole production, and reduction of sugars from sucrose; acid production from arabitol, sorbitol, and utilization of citrate were negative, but positive for raffinose and melibiose utilization. PCR amplification of the pel gene by Y1 and Y2 primers produced a 434-bp fragment on agarose gel 1% (1). Amplification of intergenic transcribed spacer region by G1 and L1 primers gave two main bands at approximately 535 and 580 bp on agarose gel 1.5%. The ITS-PCR products were digested with RsaI restriction enzyme (3). On the basis of biochemical and morphological characteristics, PCR-based pel gene and characterization of the ITS region, and digestion of the ITS-PCR products with RsaI restriction enzyme, all isolates were identified as P. carotovorum subsp. carotovorum. To our knowledge, this is the first report of soft rot caused by P. carotovorum subsp. carotovorum on cucumber from Malaysia. References: (1) A. Darraas et al. Appl. Environ. Microbiol. 60:1437, 1994. (2) N. W Schaad et al. Laboratory Guide for the Identification of Plant Pathogenic Bacteria. 3rd ed. The American Phytopathological Society Press, St. Paul, 2001. (3) I. K. Toth et al. Appl. Environ. Microbiol. 67:4070, 2001.
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