scholarly journals A New Member of the Clover Proliferation Phytoplasma Group (16SrVI) Associated with Elm Yellows in Illinois

Plant Disease ◽  
2003 ◽  
Vol 87 (3) ◽  
pp. 241-246 ◽  
Author(s):  
K. A. Jacobs ◽  
I.-M. Lee ◽  
H. M. Griffiths ◽  
F. D. Miller ◽  
K. D. Bottner
Keyword(s):  
Disease Severity ◽  
Universal Primers ◽  
Specific Primers ◽  
Detection Rates ◽  
Ulmus Americana ◽  
Disease Symptoms ◽  
Lower Trunk ◽  
Pcr Rflp ◽  
Polymerase Chain ◽  
Elm Yellows

A disease with symptoms similar to elm yellows (EY) was noticed in the early 1990s in suburban Chicago, IL. More than 1,000 mature American elms (Ulmus americana) have since died. Infected trees varied in the incidence and severity of canopy yellowing, leaf epinasty, butterscotch discoloration, and wintergreen odor of the phloem, but all developed a sparse and clumpy crown, uniformly necrotic phloem, and died within 2 years of showing canopy symptoms. Because symptoms were expressed irregularly and phytoplasma detection results by a commercial diagnostic company were inconsistent, a study was initiated to determine if EY phytoplasma was the causal agent. Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) methods using universal or EY phytoplasma specific primers were employed to detect putative phytoplasma(s) associated with 10 trees of varied disease severity within the outbreak region and 10 asymptomatic trees from an uninfected area (controls). Nested PCR using universal primers revealed that 90% of trees from the outbreak region were positive for phytoplasma while asymptomatic elms from another location (controls) tested negative. Phytoplasma-positive trees ranged in disease severity from 1 (asymptomatic) to 5 (near death). Inner bark samples chiseled from the lower trunk had higher phytoplasma detection rates than foliage or drill shavings. RFLP analyses and DNA sequencing of 16S rDNA indicated that the phytoplasma recovered from dying elms in Arlington Heights is not related to the reference EY phytoplasma (group16SrV). It is most closely related to clover proliferation (CP) phy-toplasma (group 16SrVI), and we have designated it Illinois Elm Yellows (ILEY) phytoplasma, and assigned it to a new taxonomic subgroup (16SrVI-C). EY phytoplasma was not detected in any samples, but two ILEY phytoplasma positive trees also were positive for aster yellows (AY) phytoplasma. ILEY phytoplasma was not detected in local leafhopper populations trapped in elm trees between May and September 2000. This is the first report of a phytoplasma related to CP phytoplasma causing elm yellows disease symptoms.

scholarly journals First Report of a Group 16SrII Phytoplasma Infecting Chickpea in Oman

Plant Disease ◽  
2006 ◽  
Vol 90 (7) ◽  
pp. 973-973 ◽  
Author(s):  
N. A. Al-Saady ◽  
A. M. Al-Subhi ◽  
A. Al-Nabhani ◽  
A. J. Khan
Keyword(s):  
Nested Pcr ◽  
Animal Feed ◽  
Restriction Enzymes ◽  
Universal Primers ◽  
Polymorphism Analysis ◽  
Disease Symptoms ◽  
First Report ◽  
Pcr Products ◽  
Polymerase Chain ◽  
Group 2

Chickpea (Cicer arietinum), locally known as “Dungo”, is grown for legume and animal feed mainly in the interior region of Oman. During February 2006, survey samples of chickpea leaves from plants showing yellows disease symptoms that included phyllody and little leaf were collected from the Nizwa Region (175 km south of Muscat). Total nucleic acid was extracted from asymptomatic and symptomatic chickpea leaves using a cetyltrimethylammoniumbromide method with modifications (3). All leaf samples from eight symptomatic plants consistently tested positive using a polymerase chain reaction assay (PCR) with phytoplasma universal primers (P1/P7) that amplify a 1.8-kb phytoplasma rDNA product and followed by nested PCR with R16F2n/R16R2 primers yielding a product of 1.2 kb (2). No PCR products were evident when DNA extracted from healthy plants was used as template. Restriction fragment length polymorphism analysis of nested PCR products by separate digestion with Tru9I, HaeIII, HpaII, AluI, TaqI, HhaI, and RsaI restriction enzymes revealed that a phytoplasma belonging to group 16SrII peanut witches'-broom group (2) was associated with chickpea phyllody and little leaf disease in Oman. Restriction profiles of chickpea phytoplasma were identical with those of alfalfa witches'-broom phytoplasma, a known subgroup 16SrII-B strain (3). To our knowledge, this is the first report of phytoplasma infecting chickpea crops in Oman. References: (1) A. J. Khan et al. Phytopathology, 92:1038, 2002. (2). I.-M. Lee et al. Int. J. Syst. Bacteriol. 48:1153, 1998 (3) M. A. Saghai-Maroof et al. Proc. Natl. Acad. Sci. USA. 81:8014, 1984.

Application of polymerase chain reaction for detection of goats' milk adulteration by milk of cow

2001 ◽  
Vol 68 (2) ◽  
pp. 333-336 ◽  
Author(s):  
JACEK BANIA ◽  
MACIEJ UGORSKI ◽  
ANTONI POLANOWSKI ◽  
ERYK ADAMCZYK
Keyword(s):  
Polymerase Chain Reaction ◽  
Dna Analysis ◽  
Marker Gene ◽  
Meat Products ◽  
Universal Primers ◽  
Animal Origin ◽  
Chain Reaction ◽  
Specific Primers ◽  
Single Stranded Conformational Polymorphism ◽  
Polymerase Chain

Numerous methods based on DNA analysis have been employed in the food industry to monitor adulterations of food products of animal origin. Among them the most frequently used are: polymerase chain reaction (PCR) amplification of a marker gene fragment(s) with universal primers, or amplification of DNA with species-specific primers. PCR-products of different origin can be discriminated by size, restriction fragment length polymorphism (RFLP) or single stranded conformational polymorphism (SSCP) analysis. These methods have been used for identification, and differentiation between, the animal origins of raw or heat-treated meat and meat products (Chikuni et al. 1994; Meyer et al. 1994, 1995; Zehner et al. 1998; Behrens et al. 1999; Guoli et al. 1999; Hopwood et al. 1999; Matsunaga et al. 1999; Wolf et al. 1999). These approaches are also applicable to the analysis of dairy products. However, adulterations of goats' milk and its products are traditionally tested by immunological and/or electrophoretic methods (Amigo et al. 1992; Levieux & Venien, 1994; Mimmo & Pagani, 1998). So far, only a few DNA-based techniques designed to detect the presence of bovine DNA in goats' milk have been described (Plath et al. 1997; Branciari et al. 2000). This paper presents a one-step PCR procedure for detection of adulteration of goats' milk with cows' milk. The method, employing bovine-specific primers for amplification of a 274 bp fragment of cytochrome b DNA, seems to be simple, fast, specific and sensitive.

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.

Molecular and morphological discrimination betweenTylospora fibrillosaandTylospora asterophoramycorrhizae

1999 ◽  
Vol 77 (1) ◽  
pp. 11-21 ◽  
Author(s):  
Ursula Eberhardt ◽  
Lutz Walter ◽  
Ingrid Kottke
Keyword(s):  
Fragment Length Polymorphism ◽  
Pcr Primers ◽  
Morphological Identification ◽  
Chain Reaction ◽  
Specific Primers ◽  
Specific Pcr ◽  
Pcr Rflp ◽  
Polymerase Chain ◽  
Species Specific ◽  
First Time

Among the mycorrhizal types of spruce, Tylospora-type mycorrhizae are the most constant and abundant. Two species of the genus Tylospora occur in Europe, Tylospora fibrillosa and Tylospora asterophora. Mycorrhizae of T. asterophora are described in detail for the first time. Sequences of the internal transcribed spacer (ITS) of the ribosomal genes were obtained from T. fibrillosa and T. asterophora mycorrhizae, sporocarps, and cultured mycelium. Discrimination and identification of the two species by ITS polymerase chain reaction - restriction fragment length polymorphism (PCR-RFLP) are discussed in the light of inter- and intra-specific variability. Species-specific PCR primers were designed to distinguish both species. Molecular screening of Tylospora-type mycorrhizae from field material led to unambiguous results, whereas morphological identification is likely to fail because of great similarity even at the microscopic level.Key words: Tylospora asterophora, Tylospora fibrillosa, ectomycorrhizae, taxon specific primers (TSOPs), ITS sequences.

scholarly journals Detection of Gaeumannomyces graminis Varieties Using Polymerase Chain Reaction with Variety-Specific Primers

Plant Disease ◽  
2000 ◽  
Vol 84 (9) ◽  
pp. 947-951 ◽  
Author(s):  
H. M. Fouly ◽  
H. T. Wilkinson
Keyword(s):  
Polymerase Chain Reaction ◽  
Fungal Pathogens ◽  
Gaeumannomyces Graminis ◽  
Universal Primers ◽  
Middle Region ◽  
Chain Reaction ◽  
Specific Primers ◽  
Grass Roots ◽  
Polymerase Chain ◽  
Take All

The polymerase chain reaction (PCR) was used for detection of Gaeumannomyces graminis, the causal agent of take-all disease in wheat, oats, and turfgrass. NS5 and NS6 universal primers amplified the middle region of 18S ribosomal DNA of Gaeumannomyces species and varieties. Primers GGT-RP (5′ TGCAATGGCTTCGTGAA 3′) and GGA-RP (5′ TTTGTGTGTGAC CATAC 3′) were developed by sequence analysis of cloned NS5-NS6 fragments. The primer pair NS5:GGT-RP amplified a single 410-bp fragment from isolates of G. graminis var. tritici, a single 300-bp fragment from isolates of G. graminis var. avenae, and no amplification products from isolates of G. graminis var. graminis or other species of Gaeumannomyces. The primer pair NS5:GGA-RP amplified a single 400-bp fragment from isolates of varieties tritici and avenae. Two sets of primer pairs (NS5:GGT-RP and NS5:GGA-RP) were used in PCR reactions to detect and identify the varieties tritici and avenae either colonizing wheat, oats, or grass roots, or in culture. No amplification products were observed using DNA extracted from plants infected with eight other soilborne fungal pathogens or from uninoculated plants.

scholarly journals Identification of Eutypa lata by PCR-RFLP

Plant Disease ◽  
2004 ◽  
Vol 88 (9) ◽  
pp. 925-929 ◽  
Author(s):  
P. E. Rolshausen ◽  
F. Trouillas ◽  
W. D. Gubler
Keyword(s):  
Universal Primers ◽  
Phaeomoniella Chlamydospora ◽  
Phomopsis Viticola ◽  
Eutypa Lata ◽  
Incorrect Diagnosis ◽  
Pcr Rflp ◽  
Pcr Method ◽  
Polymerase Chain ◽  
Rflp Assay

Eutypa lata is a vascular canker pathogen of woody plants commonly diagnosed by isolating the pathogen from infected tissue. Related fungi from the same family, the Diatrypaceae, also have been found in association with grapevine in Californian vineyards. An in situ polymerase chain reaction (PCR) method has been developed for detection of E. lata in infected wood tissue. However, our results indicate that this method also would amplify other Diatrypaceous fungi, which could potentially lead to an incorrect diagnosis. Therefore, we developed a PCR-restriction fragment length polymorphism (PCR-RFLP) assay. The internal transcribed spacer (ITS)1/5.8S/ITS2 ribosomal DNA region was amplified by PCR using universal primers, and RFLP patterns were determined after digestion with AluI. The restriction profiles obtained served to distinguish E. lata from wood trunk pathogens of grapevine (Phomopsis viticola, Botryodiplodia sp., Phaeoacremonium aleophilum, and Phaeomoniella chlamydospora), Diatrypaceous fungi (Diatrype sp., Diatrypella sp., Eutypella vitis, and Eutypa leptoplaca), and Cryptovalsa sp. found on dead wood of grapevine, and other Eutypa spp. (E.petrakii var. hederae, E. astroidea, E. crustata, and E. lejoplaca), with the exception of E. armeniacae, which we regard as a synonym for E. lata, and E. laevata, which has not been found on grapevine.

scholarly journals Survey of mutations in prolificacy genes in Santa Ines and Morada Nova sheep

2017 ◽  
Vol 69 (4) ◽  
pp. 1047-1053
Author(s):  
G.M.L. Holanda ◽  
J.C. Oliveira ◽  
D.M.F. Silva ◽  
S.S.N. Rocha ◽  
V. Pandolfi ◽  
...  
Keyword(s):  
Restriction Site ◽  
Fragment Length Polymorphism ◽  
Nucleotide Sequencing ◽  
Chain Reaction ◽  
Specific Primers ◽  
Pcr Rflp ◽  
Pcr Products ◽  
Polymerase Chain ◽  
Santa Inês ◽  
Restriction Fragment Length

ABSTRACT Polymorphisms in the BMP-15 gene related to Galway (FecXG) and Inverdale (FecXI) and in the BMPR-1B gene known as Booroola (FecB) mutations were investigated using the Polymerase Chain Reaction - Restriction Fragment Length Polymorphism (PCR-RFLP) method, on sheep from the breeds Santa Inês (n= 574) and Morada Nova (n=282). DNA was extracted and amplified through PCR with specific primers that introduced a restriction site in association with the mutation. The PCR products were submitted to endonucleases. The experiment found no FecXG and FecXI mutations. Six samples of animals with multiple offspring/birth history presented polymorphism for FecB similar to control samples, but this pattern was not confirmed by nucleotide sequencing. Although the absence of these mutations in the studied breeds, other factors related to prolificacy should be investigated to explain the inherent prolificity mechanisms.

scholarly journals Disease Severity and Molecular Identification of Banana bunchy top virus, Infecting Local Banana in Bali Island

2020 ◽  
Vol 24 (1) ◽  
pp. 11
Author(s):  
Gusti Ngurah Alit Susanta Wirya ◽  
I Putu Sudiarta ◽  
Dewa Gede Wiryangga Selangga
Keyword(s):  
Sequence Analysis ◽  
Disease Severity ◽  
Specific Primers ◽  
Banana Bunchy Top Virus ◽  
Research Activities ◽  
Field Samples ◽  
Close Relationship ◽  
Dna Fragment ◽  
Polymerase Chain ◽  
Relationship Of

Bunchy top symptoms on banana has been reported in Bali Island since early 2011. Symptoms variation were observed in the field similar to infection of Banana bunchy top virus (BBTV). The identity of the BBTV in Bali on the basis of DNA-S nucleotide sequence has not been studied, therefore research was conducted to identify the species of BBTV infecting local banana in Bali based on sequence analysis. Research activities were initiated by collecting field samples from several local banana growing areas in Bali Island. Incidence of bunchy top disease in all locations reached 8% to 44% with disease severity ranged from 2.6% to 30%. Identification of BBTV from field samples were done by polymerase chain reaction using specific primers for BBTV (CPF/CPR) followed by sequence analysis of amplified DNA target. Specific BBTV DNA fragment was successfully amplified from 10 field samples; sequence analysis of DNA fragments showed their highest homology with BBTV. In addition the phylogenetic analysis confirmed the close relationship of  BBTV  isolates from Bali with various BBTV isolates from Indonesia and other isolates from the Asian group in GeneBank.

scholarly journals Phytoplasmas as Causal Agents of Celosia Disease in Israel

HortScience ◽  
2000 ◽  
Vol 35 (6) ◽  
pp. 1103-1106 ◽  
Author(s):  
E. Tanne ◽  
L. Kuznetsova ◽  
J. Cohen ◽  
S. Alexandrova ◽  
A. Gera
Keyword(s):  
Rflp Analysis ◽  
Restriction Pattern ◽  
Crop Failure ◽  
Specific Primers ◽  
Aster Yellows ◽  
Pcr Rflp ◽  
Different Types ◽  
Phytoplasma Infection ◽  
Total Crop ◽  
Polymerase Chain

Recently, yellows diseases have become more common in Israel, and phytoplasmas have been detected in some of these diseased crops. Commercial fields of two celosia species (Celosia plumosa L. and C. cristata L.) also have exhibited yellows symptoms and total crop failure. Typical mycoplasma-like bodies were observed in infected but not in healthy plants. The same plants were analyzed for the presence of phytoplasma by polymerase chain reaction (PCR), using the universal oligonucleotide pair r16SF2/r16SR2, followed by nested PCR using group-specific primers. Restriction analyses performed with these products indicated that two different types of phytoplasmas are infecting celosia. PCR-RFLP analysis of one type revealed a restriction pattern typical of aster yellows. Similar analysis of the second type indicated possible relatedness, though not identity, to the pattern of phytoplasmas of the Western-X group. This is, to our knowledge, the first report of phytoplasma infection in celosia.

scholarly journals RAPID PCR–BASED DETECTION OPTIMIZATION OF PORCINE DNA IN GELATIN CAPSULE SHELL

2018 ◽  
Vol 10 (6) ◽  
pp. 217
Author(s):  
Amalia Sitti Khayyira ◽  
Viktoria Mardhika Estepane ◽  
Amarila Malik
Keyword(s):  
Molecular Detection ◽  
Universal Primers ◽  
Gelatin Capsule ◽  
Duplex Pcr ◽  
Rapid Pcr ◽  
Capsule Shell ◽  
Pcr Rflp ◽  
Pcr Method ◽  
Polymerase Chain ◽  
Bovine Species

Objective: This study was conducted to optimize the genomic deoxyribonucleic acid (DNA) based molecular detection of gelatin derived from porcine by performing polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) and duplex PCR method employing cyt B gene.Methods: Optimization was carried out for DNA extraction, PCR conditions, and the sensitivity of the PCR-RFLP method. Due to the very low DNA trace in gelatin after the various manufacturing process, the extraction was optimized to obtain sufficient DNA which was visible on the agarose gel. PCR-RFLP was carried out using universal primers and BsaJI restriction enzyme, and duplex PCR was carried out using two sets of porcine-specific primers. Porcine and bovine DNA were mixed in various concentration to confirm sensitivity of both methods, i.e. 100%, 50%, 10%, 1%, 0.5%, 0.1%, 0.05%, and 0.01%Results: Both methods, PCR-RFLP, and Duplex PCR, were able to detect as low as 0.01% porcine DNA, indicated by the presence of porcine DNA amplicon bands (131 bp and 228 bp for PCR-RFLP, 212 bp and 398 bp for duplex PCR). Although DNA bands presented in low intensity, identification of porcine and bovine species and estimation of DNA quantities were possible.Conclusion: Both conventional PCR methods, i.e. PCR-RFLP and Duplex PCR, were sensitive, specific, and suitable as a rapid initial detection method for molecular detection of porcine in gelatin capsule shells.

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