scholarly journals A Phytoplasma Belonging to a 16SrIII-A Subgroup and dsRNA Virus Associated with Cassava Frogskin Disease in Brazil

Plant Disease ◽  
2014 ◽  
Vol 98 (6) ◽  
pp. 771-779 ◽  
Author(s):  
Adriana N. de Souza ◽  
Fábio N. da Silva ◽  
Ivan P. Bedendo ◽  
Claudine M. Carvalho
Keyword(s):  
Dsrna Virus ◽  
Nested Polymerase Chain Reaction ◽  
First Report ◽  
Reverse Transcription Pcr ◽  
Minas Gerais State ◽  
Bacillus Spp ◽  
Total Dna ◽  
Polymerase Chain ◽  
Rna Polymerase Gene

Cassava frogskin disease (CFSD) is a particular threat in cassava because symptoms remain hidden until harvest and losses can be total. The information related to the etiological agent of this disease is contradictory, because some authors believe it is caused by phytoplasmas while others believe that it is caused by a virus. In order to refine detection protocols and to characterize organisms associated with CFSD in Brazil, 32 symptomatic and 20 asymptomatic cassava plants were collected in Minas Gerais state. Total DNA was extracted and used for nested polymerase chain reaction (PCR) to detect phytoplasmas. Because endophytic Bacillus spp. led to false positives, primers were designed to facilitate the detection of phytoplasma in the presence of bacteria. In addition, double-stranded (ds)RNA was extracted from tubers and used in reverse-transcription PCR for the detection of the RNA-dependent RNA polymerase gene from Cassava frogskin virus segment 4. The detected phytoplasma was identified as belonging to the group 16SrIII-A by restriction fragment length polymorphism (RFLP), sequencing, and RFLP in silico. This is the first report of a phytoplasma belonging to the 16SrIII-A group associated with cassava plants, the first molecular characterization of a phytoplasma associated with CFSD in Brazil, and a first report of phytoplasma and a dsRNA virus (possible reovirus) co-infecting cassava plants with CFSD symptoms.

scholarly journals Characterization of Xanthomonas axonopodis pv. phaseoli isolates

2008 ◽  
Vol 34 (3) ◽  
pp. 228-231 ◽  
Author(s):  
Willian Mário de Carvalho Nunes ◽  
Maria Júlia Corazza ◽  
Silvana Aparecida Crestes Dias de Souza ◽  
Siu Mui Tsai ◽  
Eiko Eurya Kuramae
Keyword(s):  
Xanthomonas Campestris ◽  
Random Amplified Polymorphic Dna ◽  
Hypersensitive Reaction ◽  
Chain Reaction ◽  
Xanthomonas Axonopodis ◽  
Paulo State ◽  
Total Dna ◽  
Polymerase Chain ◽  
Bacterial Plant Pathogen

A simple, quick and easy protocol was standardized for extraction of total DNA of the bacteria Xanthomonas axonopodis pv. phaseoli. The DNA obtained by this method had high quality and the quantity was enough for the Random Amplified Polymorphic DNA (RAPD) reactions with random primers, and Polymerase Chain Reaction (PCR) with primers of the hypersensitivity and pathogenicity gene (hrp). The DNA obtained was free of contamination by proteins or carbohydrates. The ratio 260nm/380nm of the DNA extracted ranged from 1.7 to 1.8. The hrp gene cluster is required by bacterial plant pathogen to produce symptoms on susceptible hosts and hypersensitive reaction on resistant hosts. This gene has been found in different bacteria as well as in Xanthomonas campestris pv. vesicatoria (9). The primers RST21 and RST22 (9) were used to amplify the hrp gene of nine different isolates of Xanthomonas axonopodis pv. phaseoli from Botucatu, São Paulo State, Brazil, and one isolate, "Davis". PCR amplified products were obtained in all isolates pathogenic to beans.

scholarly journals Identification of Sugarcane Striate Mosaic-Associated Virus by Partial Characterization of Its Double-Stranded RNA

1999 ◽  
Vol 89 (10) ◽  
pp. 877-883 ◽  
Author(s):  
Yoon Gi Choi ◽  
Barry J. Croft ◽  
John W. Randles
Keyword(s):  
Double Stranded Rna ◽  
Virus Particles ◽  
Reverse Transcription Pcr ◽  
Protein Size ◽  
Putative Coat Protein ◽  
Base Pair Fragment ◽  
Potato Virus M ◽  
Polymerase Chain ◽  
Pair Fragment

Sugarcane striate mosaic (ScSM)-affected sugarcane leaves contain a disease-associated 9-kilobase (kb) double-stranded RNA (dsRNA), usually together with 6- and 2.6-kb dsRNAs. The purified 9-kb dsRNA was amplified by the randomly primed polymerase chain reaction (PCR) and cloned. The nucleotide sequences of three separate regions, representing about 2.55 kb (28%) of the dsRNA sequence, were found to have significant similarities to viruses in the genera Capillo-, Carla-, Fovea-, Potex-, Poty-, Tricho-, and Tymovirus. Greatest overall similarity was found to apple stem pitting virus, with less similarity to blueberry scorch virus and potato virus M. A standard virus purification procedure was used to identify slightly flexuous filamentous particles that copurified with the disease-associated RNA. Particle modal lengths were approximately 950 and 1,900 nm with a diameter of 15 nm. Preparations contained a 51-kDa putative capsid protein and a 9-kb single-stranded RNA with a probable 3′ polyadenylate tract. These ScSM-associated virus particles differ physically from viruses in existing genera because of their relative rigidity, length, and putative coat protein size. Reverse-transcription PCR with a primer pair designed from the sequenced segments amplified a 820-base pair fragment from ScSM-affected but not healthy sugarcane plants.

scholarly journals Isolation and molecular characterization of Acanthamoeba genotypes in recreational and domestic water sources from Jamaica, West Indies

2015 ◽  
Vol 13 (3) ◽  
pp. 909-919 ◽  
Author(s):  
Cheridah D. Todd ◽  
María Reyes-Batlle ◽  
José E. Piñero ◽  
Enrique Martínez-Carretero ◽  
Basilio Valladares ◽  
...  
Keyword(s):  
Water Sources ◽  
Skin Infections ◽  
Domestic Water ◽  
Free Living ◽  
Water Contact ◽  
First Report ◽  
The Caribbean ◽  
Polymerase Chain ◽  
Neurological Infections

Free living amoebae (FLA) are amphizoic protozoa that are ubiquitous in nature. Infection with FLA may result in neurological, ocular and skin infections. Exposure to Acanthamoeba occurs frequently through water contact and knowledge of the presence of the organisms in water sources is important in understanding transmission dynamics. The distribution of Acanthamoeba was studied in recreational and domestic water samples collected from across Jamaica. Morphological assessment and polymerase chain reaction revealed Acanthamoeba spp. isolates in 50.6% (42/83) and 17.3% (14/81) of recreational and domestic water, respectively. Sequencing of the DF3 region of the 18S rDNA resulted in the identification of genotypes T3, T4, T5, T10 and T11 corresponding to Acanthamoeba spp: A. griffini, A. triangularis, A. lenticulata, A. culbertsoni and A. hatchetti. Moreover, T4 was the most frequently isolated genotype in both recreational and domestic water. Thermotolerance and osmotolerance assays indicated that most isolates were potentially pathogenic. This is the first report of T3 and T10 genotypes in the Caribbean and the first report of these Acanthamoeba spp. in Jamaican waters. The study shows that there is potential risk of infection to contact wearers who practise poor lens care. Further, Acanthamoeba should be considered as a cause of neurological infections in Jamaica.

scholarly journals First Report of Pear Decline Phytoplasmas on Pear in Serbia

Plant Disease ◽  
2005 ◽  
Vol 89 (7) ◽  
pp. 774-774 ◽  
Author(s):  
B. Duduk ◽  
M. Ivanović ◽  
A. Obradović ◽  
S. Paltrinieri ◽  
A. Bertaccini
Keyword(s):  
Nested Pcr ◽  
Restriction Enzymes ◽  
Diagnostic Procedures ◽  
Nested Polymerase Chain Reaction ◽  
Universal Primer ◽  
First Report ◽  
Taxonomy Group ◽  
Pyrus Communis L ◽  
Polymerase Chain ◽  
Symptomatic Plant

During August of 2004, pear (Pyrus communis L.) plants with typical symptoms of pear decline (PD) were observed in orchards in central Serbia. The affected plants showed premature reddening and upward rolling of leaves that often showed down-turned petioles. In some cases, premature defoliation was observed. Although a similar decline of pear was observed earlier, until now, the causal agent had not been identified. DNA was extracted with a chloroform/phenol procedure from fresh leaf midribs and branch phloem scrapes of four symptomatic and one asymptomatic pear plants separately. A nested polymerase chain reaction assay (PCR) was used for phytoplasma detection (first PCR round with P1/P7 (4) phytoplasma universal primer pair, followed by nested PCR with group 16SrX specific primers f01/r01) (3). With these primers, the expected products from phloem scrapes and midrib extracts of symptomatic plant samples were obtained. Restriction fragment length polymorphism (RFLP) analyses of the f01/r01 amplicon, with RsaI and SspI restriction enzymes, discriminating among 16SrX subgroup phytoplasmas, showed profiles corresponding to those of the apple proliferation phytoplasma group, 16SrX-C subgroup, “Candidatus Phytoplasma pyri” (2). A 1,155-bp sequence of 16S rDNA gene for one of the PA2f/r (1) amplicons obtained in nested PCR on P1/P7 products from one of the leaf midrib samples was deposited in GenBank (Accession No. AY949984); both strands of the fragment were sequenced with the Big Dye Terminator reaction kit (Applied Biosystems, Foster City, CA). The sequences were analyzed with the Chromas 1.55 DNA sequencing software (Technelysium, Queensland, Australia) and aligned with BLAST software ( http://www.ncbi.nlm.nih.gov ). The blast search showed 100% homology of this sequence with that of PD strain Y16392, confirming the identity with PD of the phytoplasma detected. To our knowledge, this is the first report of pear decline phytoplasmas in Serbia. References: (1) M. Heinrich et al. Plant Mol. Biol. Rep. 19:169, 2001. (2) IRPCM Phytoplasma/Spiroplasma Working Team-Phytoplasma Taxonomy Group. Int. J. Syst. Evol. Microbiol. 54:1243, 2004. (3) K.-H. Lorenz et al. Phytopathology 85:771, 1995. (4) Schneider et al. Pages 369–380 in: Molecular and Diagnostic Procedures in Mycoplasmology. Vol I. S. Razin and J. G. Tully, eds. The American Phytopathological Society, 1995.

scholarly journals First report of Entamoeba moshkovskii in human stool samples from symptomatic and asymptomatic participants in Kenya

2019 ◽  
Vol 5 (1) ◽  
Author(s):  
Cecilia Kyany’a ◽  
Fredrick Eyase ◽  
Elizabeth Odundo ◽  
Erick Kipkirui ◽  
Nancy Kipkemoi ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Chain Reaction ◽  
Human Gut ◽  
Nested Polymerase Chain Reaction ◽  
First Report ◽  
Stool Samples ◽  
Polymerase Chain ◽  
Entamoeba Moshkovskii ◽  
Human Stool

AbstractEntamoeba moshkovskii is a member of the Entamoeba complex and a colonizer of the human gut. We used nested polymerase chain reaction (PCR) to differentiate Entamoeba species in stool samples that had previously been screened by microscopy. Forty-six samples were tested, 23 of which had previously been identified as Entamoeba complex positive by microscopy. Of the 46 specimens tested, we identified nine (19.5%) as E. moshkovskii-positive. In seven of these nine E. moshkovskii-positive samples, either E. dispar or E. histolytica (or both) were also identified, suggesting that co-infections may be common. E. moshkovskii was also detected in both symptomatic and asymptomatic participants. To the best of our knowledge, this is the first report of E. moshkovskii in Kenya.

scholarly journals Identification of a 16SrII-E Phytoplasma in Calendula arvensis, Solanum nigrum, and Chenopodium spp.

Plant Disease ◽  
2006 ◽  
Vol 90 (3) ◽  
pp. 325-330 ◽  
Author(s):  
G. Tolu ◽  
S. Botti ◽  
R. Garau ◽  
V. A. Prota ◽  
A. Sechi ◽  
...  
Keyword(s):  
Solanum Nigrum ◽  
Nested Polymerase Chain Reaction ◽  
Bois Noir ◽  
Gene Coding ◽  
Epidemiological Surveys ◽  
Polymerase Chain ◽  
Pcr Assays ◽  
Taxonomic Groups ◽  
Stolbur Phytoplasma

Epidemiological surveys were performed in Northern Sardinia (Italy) in a 10-year-old vineyard affected by “Bois noir” disease. Samples collected between May and October 2003 from chlorotic and stunted weeds belonging to 14 different taxonomic groups were indexed molecularly for detection of phytoplasmas. Nested polymerase chain reaction (PCR) assays using primers specific for the phytoplasma 16SrDNA gene showed three of six Calendula arvensis, one of two Solanum nigrum, and one of seven Chenopodium spp. assayed positive. Restriction fragment length polymorphism analyses and sequencing of amplified 16SrDNA fragments identified a putative phytoplasma in the ribosomal subgroup 16SrII-E. Further characterization of the rps3 gene, coding a ribosomal protein, confirmed the identification. However, the weeds and leafhop-per species collected in the vineyard tested negative by PCR assays for the Stolbur phytoplasma, the causal agent of “Bois noir”. This is the first report of a phytoplasma of the 16SrII-E subgroup infecting C. arvensis, S. nigrum, and Chenopodium spp.

scholarly journals Phytoplasma associated with shoot proliferation in begonia

2006 ◽  
Vol 63 (5) ◽  
pp. 475-477 ◽  
Author(s):  
Luiz Fernando Caldeira Ribeiro ◽  
Ana Paula de Oliveira Amaral Mello ◽  
Ivan Paulo Bedendo ◽  
Ricardo Gioria
Keyword(s):  
Ornamental Plants ◽  
Shoot Proliferation ◽  
Plant Size ◽  
Universal Primers ◽  
Chain Reaction ◽  
Specific Primers ◽  
Nested Polymerase Chain Reaction ◽  
Phytoplasma Infection ◽  
Total Dna ◽  
Polymerase Chain

Begonia is a very appreciated genus of ornamental plants, of economic relevancy, having species of flowers and foliage. In commercial croppings, plants exhibiting characteristic symptoms of phytoplasma infection have been observed, such as shoot proliferation, reduced plant, size small leaves and flowers, and phyllody. Leaves were sampled and total DNA was extracted to be used in nested Polymerase Chain Reaction (PCR), in order to detect and identify an expected phytoplasma. The results confirmed consistently the presence of a phytoplasma associated with symptomatic plants through the amplification of a typical genomic fragment of 1.2 kb by using the universal primers R16mF2/mR1 and R16F2n/R2. The use of specific primers R16(III)F2/R1 allowed to identify the phytoplasma detected as a representative of the group 16SrIII. This information is very expressive, because different diseases caused by fungus, bacteria, virus and nematodes have been reported for begonia, however, reports have not been found for begonia diseases associated with phytoplasmas.

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