scholarly journals Molecular Detection of Jujube Witches' Broom Phytoplasmas in Micropropagated Jujube Shoots

HortScience ◽  
2000 ◽  
Vol 35 (7) ◽  
pp. 1274-1275 ◽  
Author(s):  
J.B. Tian ◽  
H.P. Guo ◽  
A. Bertaccini ◽  
M. Martini ◽  
S. Paltrinieri ◽  
...  
Keyword(s):  
Restriction Enzymes ◽  
Shanxi Province ◽  
Nested Polymerase Chain Reaction ◽  
Group V ◽  
Pcr Products ◽  
Zizyphus Jujuba ◽  
Polymerase Chain ◽  
Positive Results ◽  
Important Disease ◽  
Apparently Healthy

Jujube (Zizyphus jujuba Mill.) witches' broom (JWB) is the most important disease in the areas of jujube cultivation in China, where it occurs every year. Micropropagated shoots of the three most important cultivars (`Lizao', `Junzao', and `Muzao') in the National Jujube Gene Pool, collected at the Pomology Institute of Shanxi province, were tested for the presence of phytoplasmas. Phytoplasma ribosomal (16Sr) general and specific primer pairs were used in direct or nested polymerase chain reaction (PCR). Positive results were obtained only from symptomatic micropropagated samples of `Lizao' and from phytoplasma controls. Restriction fragment length polymorphism (RFLP) analyses of PCR products with several restriction enzymes revealed that the phytoplasmas infecting the symptomatic plants belong to the 16SrRNA group V subgroup B. The positive correlation between symptoms and the presence of phytoplasmas was verified in tissue culture. Samples from apparently healthy shoots of `Junzao', `Muzao', and `Lizao' were free of phytoplasmas.

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.

RT-PCR analysis of Na+/H+ exchanger mRNAs in rat medullary thick ascending limb

1995 ◽  
Vol 268 (6) ◽  
pp. F1224-F1228 ◽  
Author(s):  
P. Borensztein ◽  
M. Froissart ◽  
K. Laghmani ◽  
M. Bichara ◽  
M. Paillard
Keyword(s):  
Rat Kidney ◽  
Pcr Amplification ◽  
Restriction Enzymes ◽  
Pcr Analysis ◽  
Thick Ascending Limb ◽  
Rt Pcr ◽  
Medullary Thick Ascending Limb ◽  
Pcr Products ◽  
Polymerase Chain ◽  
Nhe Isoforms

The thick ascending limb (TAL) of rat kidney absorbs bicarbonate secondary to proton secretion, but displays both basolateral and luminal Na+/H+ exchange (NHE) activity. Several NHE genes, including NHE-1, NHE-2, NHE-3, and NHE-4, are expressed in the kidney. To identify the NHE isoforms expressed in the rat medullary TAL (MTAL), we used the reverse transcription-polymerase chain reaction (RT-PCR) to detect the mRNAs for NHE in microdissected MTAL. RT-PCR amplification from total RNA was performed between two specific primers for each NHE isoform. In rat kidney homogenate, the four NHE isoform mRNAs were detected, and the identity of the PCR products was demonstrated by the sizes of the fragments, digestion with restriction enzymes, and Southern blot analysis. In microdissected rat MTAL, NHE-3 was strongly expressed and NHE-1 mRNA was also detected, whereas NHE-2 and NHE-4 mRNAs were not detected. Therefore, NHE-3 could be the apical Na+/H+ exchanger, and NHE-1 could be the basolateral isoform in the MTAL.

scholarly journals Occurrence of a 16SrIV Group Phytoplasma not Previously Associated with Palm Species in Yucatan, Mexico

Plant Disease ◽  
2011 ◽  
Vol 95 (3) ◽  
pp. 256-262 ◽  
Author(s):  
Roberto Vázquez-Euán ◽  
Nigel Harrison ◽  
María Narvaez ◽  
Carlos Oropeza
Keyword(s):  
Cocos Nucifera ◽  
Restriction Enzymes ◽  
Nested Polymerase Chain Reaction ◽  
Palm Trees ◽  
Palm Species ◽  
Leaf Yellowing ◽  
Sabal Mexicana ◽  
Polymerase Chain ◽  
First Time

The occurrence of 16SrIV group phytoplasmas in palm species Sabal mexicana and Pseudophoenix sargentii is reported here for the first time. Palm trees showed leaf decay and leaf yellowing syndromes, respectively. An amplification product (1.4 kb) was obtained in symptomatic S. mexicana (18 of 21) and symptomatic P. sargentii (1 of 1) palm trees sampled in different locations in Yucatan State, Mexico; five of the positive S. mexicana and the positive P. sargentii trees died. The identity of the phytoplasmas from these species was determined by restriction fragment length polymorphism profiling with restriction enzymes AluI and HinfI, showing there could be two phytoplasma strains of the 16SrIV group. In one S. mexicana palm, the profile was the same as observed with these enzymes for phytoplasmas of 16SrIV-A subgroup, previously associated with Cocos nucifera palm trees and, in the rest of the trees, including the P. sargentii palm, the profile was for phytoplasmas of the 16SrIV-D subgroup. These identities were supported by analyses of the amplicons obtained by nested polymerase chain reaction by nucleotide-nucleotide BLAST analysis. Geographical distribution of the association S. mexicana/16SrIV group phytoplasmas was found widely dispersed in Yucatan State. A potential role of S. mexicana palm trees as a permanent source of phytoplasma inoculum is suggested. In addition to P. sargentii, other palm species (Thrinax radiata and C. nucifera) coexisting with S. mexicana trees were also sampled and analyzed.

Nested PCR protocol for the rapid detection of Escherichia coli in potable water

1996 ◽  
Vol 42 (8) ◽  
pp. 862-866 ◽  
Author(s):  
David Juck ◽  
Jordan Ingram ◽  
Michèle Prévost ◽  
Josée Coallier ◽  
Charles Greer
Keyword(s):  
Escherichia Coli ◽  
Nested Pcr ◽  
Potable Water ◽  
Test Organism ◽  
Bacterial Cells ◽  
Nested Polymerase Chain Reaction ◽  
Fecal Indicator ◽  
E Coli ◽  
Pcr Products ◽  
Polymerase Chain

A rapid and sensitive method for the detection of low levels of bacteria in potable water was developed. The fecal indicator bacterium Escherichia coli was used as the test organism in a filtration concentration–nested polymerase chain reaction (PCR) protocol, combined with ethidium bromide visualization of PCR products. Two sets of primers were designed from the E. coli specific β-glucuronidase gene (uidA), the primary pair producing a 486-bp fragment that was used as template for the nested primer pair delineating a 186-bp fragment. This protocol can detect 1–10 bacterial cells/50 mL water sample within 6–8 h, in contrast to traditional culturing or Southern hybridization methods which require 2–3 days for results.Key words: nested PCR, sensitive, detection, potable water.

scholarly journals Detection of feline coronavirus spike gene mutations as a tool to diagnose feline infectious peritonitis

2016 ◽  
Vol 19 (4) ◽  
pp. 321-335 ◽  
Author(s):  
Sandra Felten ◽  
Karola Weider ◽  
Stephanie Doenges ◽  
Stefanie Gruendl ◽  
Kaspar Matiasek ◽  
...  
Keyword(s):  
Gene Mutations ◽  
Control Group ◽  
Nested Polymerase Chain Reaction ◽  
Feline Infectious Peritonitis ◽  
Spike Gene ◽  
S Gene ◽  
Pcr Products ◽  
Study Population ◽  
Polymerase Chain ◽  
Feline Coronavirus

Objectives Feline infectious peritonitis (FIP) is an important cause of death in the cat population worldwide. The ante-mortem diagnosis of FIP in clinical cases is still challenging. In cats without effusion, a definitive diagnosis can only be achieved post mortem or with invasive methods. The aim of this study was to evaluate the use of a combined reverse transcriptase nested polymerase chain reaction (RT-nPCR) and sequencing approach in the diagnosis of FIP, detecting mutations at two different nucleotide positions within the spike (S) gene. Methods The study population consisted of 64 cats with confirmed FIP and 63 cats in which FIP was initially suspected due to similar clinical or laboratory signs, but that were definitively diagnosed with another disease. Serum/plasma and/or effusion samples of these cats were examined for feline coronavirus (FCoV) RNA by RT-nPCR and, if positive, PCR products were sequenced for nucleotide transitions within the S gene. Results Specificity of RT-nPCR was 100% in all materials (95% confidence interval [CI] in serum/plasma 83.9–100.0; 95% CI in effusion 93.0–100.0). The specificity of the sequencing step could not be determined as none of the cats of the control group tested positive for FCoV RNA. Sensitivity of the ‘combined RT-nPCR and sequencing approach’ was 6.5% (95% CI 0.8–21.4) in serum/plasma and 65.3% (95% CI 50.4–78.3) in effusion. Conclusions and relevance A positive result is highly indicative of the presence of FIP, but as none of the control cats tested positive by RT-nPCR, it was not possible to confirm that the FCoV mutant described can only be found in cats with FIP. Further studies are necessary to evaluate the usefulness of the sequencing step including FCoV-RNA-positive cats with and without FIP. A negative result cannot be used to exclude the disease, especially when only serum/plasma samples are available.

scholarly journals Taxa of Leafhoppers Carrying Phytoplasmas at Sites of Ash Yellows Occurrence in New York State

Plant Disease ◽  
2000 ◽  
Vol 84 (2) ◽  
pp. 134-138 ◽  
Author(s):  
G. T. Hill ◽  
W. A. Sinclair
Keyword(s):  
New York ◽  
New York State ◽  
Pcr Amplification ◽  
Restriction Enzymes ◽  
Disease Group ◽  
York State ◽  
Aster Yellows ◽  
Pcr Products ◽  
Polymerase Chain ◽  
Abundant Genus

Leafhopper (Homoptera: Cicadellidae) populations were sampled and leafhopper carriers of ash yellows (AshY) phytoplasmas were identified as first steps toward vector identification. Nearly 5,000 leafhoppers were collected in malaise traps at two sites of high AshY incidence in New York state in 1996 and 1997. These insects comprised 33 taxa, including representatives of 13 genera known to contain phytoplasma vectors. The most abundant genus was Scaphoideus, with numbers about six times greater than any other genus. A total of 1,632 insects were assayed individually for phytoplasmas by polymerase chain reaction (PCR) amplification of phytoplasmal 16S rDNA and restriction fragment length polymorphism analyses of PCR products using restriction enzymes TaqI and RsaI separately. Phytoplasmas were detected in 35 insects, all but one in the subfamily Deltocephalinae. AshY phytoplasmas were detected in 19 of 812 individuals of Scaphoideus spp. and 1 of 87 of Colladonus clitellarius. Phytoplasmas of the Prunus X-disease group were detected in 1 Scaphoideus sp., 4 individuals of C. clitellarius, and 4 of 83 Scaphytopius acutus individuals. Phytoplasmas of the aster yellows group were detected in 1 of 68 individuals of Gyponana spp. and 5 of S. acutus. AshY phytoplasma carriers merit testing for possible vector ability.

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 Correlation Between Chlamydia pneumoniae Infection and Lipid Profile in Patients with Cardiovascular Diseases

2019 ◽  
pp. 2129-2135
Author(s):  
Mohammed M. F. Al. Halbosiy ◽  
May K. Ismael ◽  
Hadeel A. Nasser
Keyword(s):  
Cardiovascular Diseases ◽  
Significant Relationship ◽  
Chlamydia Pneumoniae ◽  
Age Groups ◽  
Conventional Polymerase Chain Reaction ◽  
Control Group ◽  
Blood Samples ◽  
Polymerase Chain ◽  
Positive Results ◽  
Apparently Healthy

      Cardiovascular disease (CVD) is a class of diseases that involve the blood vessels or heart. Chlamydia pneumoniae has been considered as the most reasonable; also, it is able to increase and persevere inside vascular cells and to make the chronic inflammation in atherosclerosis. In this study, blood samples were subjected for molecular detection of Chlamydia pneumoniae by using conventional polymerase chain reaction (PCR) depending on 16S rRNA. Seventy patients who suffer from cardiovascular diseases (angina, myocardial infarction and atherosclerosis) aged between 33-86 years have been investigated and compared to twenty of apparently healthy individuals were studied as a control group. Twenty-six samples (37.14) % revealed positive results for Chlamydia pneumoniae by PCR technique in blood samples of patients group, while all control samples were negative. No significant relationship was found among HDL,LDL, cholesterol, but the significant differences in the levels of triglyceride, VLDL between Chlamydia pneumoniae positive and negative within the patients' group were significant (P<0.05).The result of this study revealed that there was elevation of cholesterol and triglycerides level  in patients their ages less than 50 years  compared with other age groups although  there was no significant relationship  between  HDL VLDL,LDL and age ; but generally certain patients with more than 60 years  have the highest level of HDL VLDL, LDL.

scholarly journals Specific Polymerase Chain Reaction Primers for the Detection of Plasmodiophora brassicae in Soil and Water

1999 ◽  
Vol 89 (5) ◽  
pp. 392-397 ◽  
Author(s):  
R. Faggian ◽  
S. R. Bulman ◽  
A. C. Lawrie ◽  
I. J. Porter
Keyword(s):  
Polymerase Chain Reaction ◽  
Plasmodiophora Brassicae ◽  
Chain Reaction ◽  
Nested Polymerase Chain Reaction ◽  
Simple Method ◽  
Soil P ◽  
Pcr Products ◽  
Polymerase Chain ◽  
Polymerase Chain Reaction Primers ◽  
Soil And Water

The development of specific oligonucleotide primers for Plasmodiophora brassicae has led to a nested polymerase chain reaction (PCR) detection method for P. brassicae in soil and water. Initially, the PCR was used to amplify a section of the rDNA repeat. The PCR products were sequenced and the data used to design primers that were directed at the ribosomal RNA genes and internal transcribed spacer regions. Specificity was tested against more than 40 common soil organisms, host plants, and spore suspension contaminants, as well as P. brassicae isolates from around Australia and the world. Sensitivity was determined to be 0.1 fentograms (fg; 10-15 g) for pure template and as low as 1,000 spores per g of potting mix. In soil, P. brassicae was detected in all soils where the inoculum was sufficient to result in clubroot symptoms. Also outlined is a simple method of DNA extraction from soil.

scholarly journals PCR detection of Toxoplasma gondii B1 gene in women suffering from abortion

2019 ◽  
Vol 13 (1) ◽  
pp. 12-15
Author(s):  
Aseel S. Mahmood ◽  
Sabeeha A. Al-Sarray ◽  
Abdulkareem Al-Kazaz
Keyword(s):  
Toxoplasma Gondii ◽  
Complex Disease ◽  
Control Group ◽  
Chi Square ◽  
Nested Polymerase Chain Reaction ◽  
B1 Gene ◽  
History Of ◽  
Polymerase Chain ◽  
And Control ◽  
Apparently Healthy

Background: Primary infection of maternal with toxoplasmosis during gestation and this infection transmission to the fetus continue to be the cause complex disease in offspring. Objective: This study was conducted to test the utility of nested Polymerase Chain Reaction (nPCR) assay to detect recent infections with Toxoplasma in abortive women. Material and methods: Toxoplasma gondii DNA was detected by using B1 gene as a target for amplification which was highly specific for T. gondii and is well conserved among all of the tested strains. Blood from 60 abortive women and 25 apparently healthy pregnant women with no history of abortion (as control group) were taken in this current study. Results: The results revealed that nPCR was positive in 48(80%) subjects and negative in 12(20%), Chi-square- χ2 for patients and control was ( 13.82 , 15.75 ) respectively. Conclusion: It can be concluded that nPCR assay in blood has advantage in detection of recent and active toxoplasmosis.

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