scholarly journals Occurrence of Maize rayado fino virus in Maize in Argentina

Plant Disease ◽  
2000 ◽  
Vol 84 (9) ◽  
pp. 1046-1046 ◽  
Author(s):  
M. P. Giménez-Pecci ◽  
E. Oliveira ◽  
R. Resende ◽  
C. Borgogno ◽  
C. F. Nome ◽  
...  
Keyword(s):  
Enzyme Linked Immunosorbent Assay ◽  
Plant Tissues ◽  
Rt Pcr ◽  
Xylem Parenchyma ◽  
Chain Reaction ◽  
Polymerase Chain ◽  
Maize Rayado Fino Virus ◽  
Size Relation ◽  
Pcr Conditions ◽  
Das Elisa

Symptoms of fine chlorotic stipple-striping of the veins, chlorosis, numerous dots and stripes, formation of holes in the leaf blade, and ears reduced in size, bearing few grains, were observed in maize crops in Tafí del Valle (Tucumán Province), Orán, El Galpón (Salta Province), Tilcara and Yaví (Jujuy Province), the subtropical area of northwest Argentina where the leafhopper vector Dalbulus maidis (DeLong & Wolcott) is present. Maize rayado fino virus (MRFV) was detected in these samples by a positive reaction in double antibody sandwich-enzyme-linked immunosorbent assay (DAS-ELISA) using an AGDIA kit. Electron microscopy revealed abundant isometric particles about 30 nm in diameter in the cytoplasm and vacuoles of phloem cells and xylem parenchyma cells. The virus was also detected by reverse transcription polymerase chain reaction (RT-PCR) using a primer pair MRFV-09/MRFV-10. Primers and PCR conditions were as previously described (1). Virus amplification was observed only in samples from symptomatic plants. In 1981 (2), the presence of MRFV in Argentina was revealed by serological assay in plants from temperate central areas. No further reports were released since then. This is the first evidence of MRFV in subtropical areas of Argentina and identification of the virus by combining DAS-ELISA, particle size, relation with plant tissues, and RTPCR. References: (1) R. W. Hammond et al. J. Gen. Virol. 78:3153, 1997. (2) S. F. Nome et al. RIA XIX:257, 1984.

scholarly journals Evaluation of Sandwich Enzyme-Linked Immunosorbent Assay and Reverse Transcription Polymerase Chain Reaction for the Diagnosis of Foot-and-Mouth Disease

Intervirology ◽  
2021 ◽  
pp. 1-6
Author(s):  
Salman Khan ◽  
Syed Asad Ali Shah ◽  
Syed Muhammad Jamal
Keyword(s):  
Polymerase Chain Reaction ◽  
Viral Genome ◽  
Foot And Mouth Disease ◽  
Enzyme Linked Immunosorbent Assay ◽  
Rt Pcr ◽  
Chain Reaction ◽  
Kappa Value ◽  
Polymerase Chain ◽  
Pcr Assays ◽  
Level Of Agreement

<b><i>Background:</i></b> Foot-and-mouth disease (FMD) is an infectious and highly contagious disease of cloven-hoofed domestic and wild animals, causing heavy economic losses to the livestock industry. Rapid and reliable diagnosis of the disease is essential for the implementation of effective control measures. This study compared sandwich enzyme-linked immunosorbent assay (S-ELISA) and conventional reverse transcription polymerase chain reaction (RT-PCR) for the diagnosis of FMD. <b><i>Methods:</i></b> A total of 60 epithelial samples from suspected cases of FMD were tested using both S-ELISA and RT-PCR assays. The level of agreement between the assays was assessed by calculating the Kappa value. <b><i>Results:</i></b> S-ELISA detected 38 (63%) samples positive for FMD virus (FMDV). Being predominant, serotype O was detected in 22 (57.9%) of the total samples tested positive, whereas 9 (23.7%) and 7 (18.4%) samples were found positive for serotypes A and Asia-1, respectively. RT-PCR detected viral genome in 51 (85%) of the samples using pan-FMDV primers set, 1F/1R. Thirty-six samples were found positive and 7 negative by both the tests. The level of agreement between the tests was assessed by calculating the Kappa value, which was found to be fair (Kappa value = 0.303 and 95% CI = 0.089; 0.517) and significant (<i>p</i> = 0.009). However, 2 samples, which were found positive on S-ELISA tested negative on RT-PCR. This may be attributed to the presence of nucleotide mismatch(es) in the primer-binding sites that may have resulted in failure of amplification of the viral genome. The serotype-specific RT-PCR assays not only confirmed serotyping results of S-ELISA but were also able to establish serotype in 9 S-ELISA-negative but pan-FMDV RT-PCR-positive samples. <b><i>Conclusions:</i></b> The RT-PCR assay contributes significantly to establishing a quick, sensitive, and definitive diagnosis of FMD in resource-constrained countries. Samples giving negative results in S-ELISA should be tested in RT-PCR for the disease detection and virus typing.

Detection of small round structured viruses in clinical and environmental samples by polymerase chain reaction

1995 ◽  
Vol 31 (5-6) ◽  
pp. 375-382 ◽  
Author(s):  
M. Wolfaardt ◽  
C. L. Moe ◽  
W. O. K. Grabow
Keyword(s):  
Polymerase Chain Reaction ◽  
Tap Water ◽  
Practical Method ◽  
Glass Wool ◽  
Polluted Water ◽  
Enzyme Linked Immunosorbent Assay ◽  
Rt Pcr ◽  
Chain Reaction ◽  
Polymerase Chain ◽  
Elution Method

Norwalk (NV) and other small round structured viruses (SRSVs) have been identified as common causes of gastroenteritis. Outbreaks of Norwalk gastroenteritis have been associated with contaminated drinking water and food such as oysters and salads. The cloning and sequencing of the NV genome has made it possible to detect NV and related viruses by the reverse transcription-polymerase chain reaction (RT-PCR). We applied RT-PCR to detect SRSVs in faecal specimens from two gastroenteritis outbreaks in South Africa, designated “Christmas” and “Grootbrak” and were able to detect SRSVs in all of the three specimens from the Christmas outbreak and in two of 16 specimens from the Grootbrak outbreak. The RT-PCR procedure used appeared to be more sensitive for the detection of SRSVs in patient stool specimens than immune electron microscopy and NV antigen detection by enzyme linked immunosorbent assay. The RT-PCR procedure proved suitable for the detection of SRSVs in seeded samples of sewage, sewage sludge, river water, and tap water. However, sensitivity was lower for seeded samples of sewage and sludge than for tap water, which indicates interference by high levels of organic matter. The RT-PCR procedure was also used to show that small numbers of SRSVs can successfully be recovered from large volumes of water by means of a glass wool adsorption-elution method. Since no practical method is available for quantitation of the small numbers of SRSVs concerned, it was not possible to evaluate the efficiency of recovery. Although no SRSVs have been detected by direct testing of sewage and sludge samples, the results obtained in this study show that RT-PCR detection of SRSVs in sewage and polluted water environments is feasible, and that small numbers of the viruses can, like many other enteric viruses, successfully be recovered by means of a glass wool adsorption-elution method.

scholarly journals RT-PCR Method for Detecting Cowpea Mottle Carmovirus in Vigna Germ Plasm

Plant Disease ◽  
1999 ◽  
Vol 83 (7) ◽  
pp. 639-643 ◽  
Author(s):  
A. G. Gillaspie ◽  
S. E. Mitchell ◽  
G. W. Stuart ◽  
R. F. Bozarth
Keyword(s):  
Germ Plasm ◽  
Enzyme Linked Immunosorbent Assay ◽  
Mild Mosaic ◽  
Rt Pcr ◽  
Chain Reaction ◽  
Viral Pathogens ◽  
Highly Sensitive ◽  
Pcr Method ◽  
The Common ◽  
Polymerase Chain

A highly sensitive reverse transcription-polymerase chain reaction (RT-PCR) method was developed to detect cowpea mottle carmovirus (CPMoV) in newly acquired germ plasm of Vigna spp. It detected virus in tissues diluted up to 10-9. The preferred primers were designed from the RNA replicase cDNA sequence of CPMoV. These primers were able to detect CPMoV in plants infected with 10 different isolates of the virus. There were no cross-reactions with either bean mild mosaic or melon necrotic spot carmoviruses or any of the common cowpea viral pathogens tested. The RT-PCR method was up to 105 times more sensitive than direct antigen coating enzyme-linked immunosorbent assay (DAC-ELISA) in detecting CPMoV. The RT-PCR method gave no false positive reaction as is sometimes seen with ELISA.

scholarly journals Circulation of bluetongue virus in goats in the Karamoja region of Uganda

2013 ◽  
Vol 84 (1) ◽  
Author(s):  
Elijah N. Mulabbi ◽  
Chrisostom Ayebazibwe ◽  
Samuel Majalija ◽  
Carrie A. Batten ◽  
Christopher A.L. Oura
Keyword(s):  
Real Time ◽  
Whole Blood ◽  
Bluetongue Virus ◽  
Rna Virus ◽  
Enzyme Linked Immunosorbent Assay ◽  
Rt Pcr ◽  
High Seroprevalence ◽  
Chain Reaction ◽  
Polymerase Chain ◽  
Karamoja Region

The presence of bluetongue virus (BTV) in indigenous goats from the Karamoja region of northern Uganda was investigated. A total of 300 goats were sampled (serum and whole blood) from five districts within the Karamoja region. The samples were analysed for the presence of bluetongue (BT) antibodies using a commercial Enzyme-linked immunosorbent assay (ELISA) and for the presence of BTV viral RNA by real-time Reverse transcription polymerase chain reaction (RT-PCR), because BTV is an RNA virus. Of the 300 goats tested, 269 (90%) were positive for BTV antibodies, indicating high levels of BTV circulation within the region. Out of the 150 whole blood samples tested for the presence of the virus by real-time RT-PCR, 84 (56%) were positive for BTV RNA. This study, which is the first of its kind in Uganda, showed a high seroprevalence of BT antibodies and active circulation of BTV in a high proportion of goats in the Karamoja region.

scholarly journals The occurrence of the Tulip breaking virus in tulips in the northern part of Turkey

2019 ◽  
Vol 31 (2) ◽  
pp. 263-268
Author(s):  
Ilyas Deligoz ◽  
Mehmet Ali Sevik
Keyword(s):  
Viral Disease ◽  
Cut Flower ◽  
Rt Pcr ◽  
Chain Reaction ◽  
Black Sea Region ◽  
Tulip Breaking Virus ◽  
Polymerase Chain ◽  
Pcr Assays ◽  
Garden Plant ◽  
Das Elisa

AbstractThe tulip (Tulipa sp.) is one of the most important ornamental bulbous plants, which has been cultivated as a cut-flower, potted, and garden plant, and used for landscaping in Turkey. This study investigated the occurrence of a viral disease in the tulip cultivars Strong Gold, Pretty Woman and Purple Prince that causes striping of the leaves, flames of different colours on the petals and mosaic patterns on the leaves, in Samsun province of Turkey. Surveys of virus-infected tulip plants were carried out in the Middle Black Sea Region of Turkey in 2015-2016. A total of 212 samples were collected from four locations and checked by biological, serological and molecular methods for the presence of the Tulip breaking virus (TBV). TBV was detected in the leaves and flowers by double-antibody sandwich enzyme-linked immunosorbent assays (DAS-ELISA) in the tulip cultivars (15.5%) tested from Samsun province. TBV infection was found at the highest rate in the cultivar Strong Gold (19.7%), followed by Pretty Woman (14.1%) and Purple Prince (12.8%). The presence of TBV in samples was further confirmed by reverse transcription polymerase chain reaction (RT-PCR) assays. This is the first report on TBV naturally infecting tulips in Samsun province, Turkey.

Reverse-Transcription Polymerase Chain Reaction Detection of the Enteroviruses

1999 ◽  
Vol 123 (12) ◽  
pp. 1161-1169 ◽  
Author(s):  
JoséR. Romero
Keyword(s):  
Polymerase Chain Reaction ◽  
Sensitivity And Specificity ◽  
Reverse Transcription ◽  
Health Care Cost ◽  
Cost Savings ◽  
Rt Pcr ◽  
Chain Reaction ◽  
Reverse Transcription Pcr ◽  
Polymerase Chain ◽  
Pcr Conditions

Abstract Objective.—This review focuses on commercial and in-house–developed reverse-transcription polymerase chain reaction (RT-PCR) assays used for the detection of enteroviral infections. In addition to providing details on the performance of RT-PCR, its specificity, and sensitivity, the clinical utility of this diagnostic method with specific reference to its impact on hospitalization and cost savings is addressed. Data Sources.—MEDLINE was searched for reports relating to RT-PCR detection of the enteroviruses in adults and children. The search was restricted to studies reported in English language journals. Study Selection.—Reports documenting detailed information regarding the RT-PCR conditions, primers, sensitivity, specificity and, if relevant, clinical impact were selected for analysis. Data Extraction.—Details regarding method of extraction of the enteroviral genome, the primers used, RT-PCR conditions, and sensitivity and specificity of the assay were extracted from the literature. For reports detailing the use of RT-PCR in the clinical management of enteroviral infections in children, the reduction in duration of hospitalization and health care cost savings were recorded. Data Synthesis.—Reverse-transcription PCR can increase the yield of detection of enteroviruses from cerebrospinal fluid by a mean of approximately 20% over tissue culture. Reverse-transcription PCR of cerebrospinal fluid has been shown to exhibit sensitivity and specificity values of 86% to 100% and 92% to 100%, respectively. Reductions of 1 to 3 days of hospitalization per patient are predicted if RT-PCR is used to diagnose enteroviral meningitis in children. Conclusions.—Reverse-transcription PCR detection of enteroviral infections is an extremely rapid, sensitive, and specific diagnostic modality. Both commercial assays and assays developed in-house appear to be equivalent with regard to sensitivity and specificity. Reverse-transcription PCR diagnosis of enteroviral infections in children could reduce the length of hospitalization and result in significant health care cost savings.

Interlaboratory Evaluation of a New Reverse Transcriptase Polymerase Chain Reaction–Based Enzyme-Linked Immunosorbent Assay for the Detection of Circulating Melanoma Cells: A Multicenter Study of the Dermatologic Cooperative Oncology Group

2001 ◽  
Vol 19 (6) ◽  
pp. 1723-1727 ◽  
Author(s):  
U. Reinhold ◽  
C. Berkin ◽  
A.-K. Bosserhoff ◽  
A. Deutschmann ◽  
C. Garbe ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Tumor Cells ◽  
High Sensitivity ◽  
Enzyme Linked Immunosorbent Assay ◽  
Rt Pcr ◽  
Chain Reaction ◽  
Blood Samples ◽  
Melanoma Patients ◽  
Polymerase Chain ◽  
Interlaboratory Reproducibility

PURPOSE: Reverse transcription-polymerase chain reaction (RT-PCR)–based detection of tyrosinase mRNA is the most frequently used laboratory method for the detection of circulating tumor cells in melanoma patients. However, previously published results showed considerable variability in the PCR positivity rates. MATERIALS AND METHODS: We designed a collaborative study to assess the sensitivity, specificity, and clinical relevance of a new standardized RT-PCR–based enzyme-linked immunosorbent assay (ELISA) for the detection of circulating melanoma cells. Blood samples of healthy donors mixed with cells of a melanoma cell line were prepared in a blinded fashion, and aliquots were sent to seven participating laboratories experienced in RT-PCR. RESULTS: The results demonstrate a high sensitivity (1 melanoma cell/mL blood) and specificity (no false-negatives and 7.4% [2 of 28] false-positives) of the assay and a satisfactory rate of interlaboratory reproducibility. The analysis of aliquots of blinded samples derived from 60 melanoma patients identified tyrosinase mRNA in 17 of 60 (28.3%): three (20%) of 15 stage I patients, two (13.3%) of 15 stage II patients, five (35.7%) of 14 stage III patients, and seven (43.8%) of 16 stage IV patients. The interlaboratory reproducibility of positive samples, however, was extremely low and indicates the presence of low amounts of target mRNA. CONCLUSION: Reverse transcriptase-PCR ELISA has a high sensitivity and specificity for the detection of tyrosinase mRNA in peripheral blood cells. The low interlaboratory reproducibility for the detection of tumor cells in blood samples of melanoma patients, however, raises the question of relevance of this assay for clinical use.

scholarly journals Sensitive and specific cytokeratin 18 reverse transcription-polymerase chain reaction that excludes amplification of processed pseudogenes from contaminating genomic DNA

1997 ◽  
Vol 43 (12) ◽  
pp. 2244-2250 ◽  
Author(s):  
Peter Tschentscher ◽  
Christoph Wagener ◽  
Michael Neumaier
Keyword(s):  
Polymerase Chain Reaction ◽  
Reverse Transcription ◽  
Genomic Dna ◽  
Rt Pcr ◽  
Chain Reaction ◽  
Cytokeratin 18 ◽  
Processed Pseudogenes ◽  
Polymerase Chain ◽  
Positive Results ◽  
Pcr Conditions

Abstract Processed pseudogenes of residual contaminating genomic DNA interfere with a sensitive detection of cytokeratin 18 (CK18) mRNA by reverse transcription and polymerase chain reaction (RT-PCR). This may cause false-positive results when CK18 mRNA is used as a marker for ectopic tumor cells in specimens from cancer patients. To establish a sensitive CK18 RT-PCR by excluding the amplification of processed pseudogenes, the following strategy was chosen: (a) CK18 pseudogene sequences were cloned from genomic DNA by PCR; (b) cDNA-specific primers were designed on the basis of mismatches between pseudogenes and cDNA; (c) PCR conditions were adjusted to reach maximum sensitivity and specificity. Epithelial cells (1–10) could be detected in 1 mL of blood. Among the numerous CK18 genes homologous to the transcribed gene, at least two different processed pseudogenes exist that are highly homologous to each other and to the exons of the transcribed CK18 gene.

scholarly journals Evaluation of peanut genotypes for resistance to Tomato spotted wilt virus by mechanical and thrips inoculation

2006 ◽  
Vol 41 (6) ◽  
pp. 937-942 ◽  
Author(s):  
Luciana Cordeiro do Nascimento ◽  
Viboon Pensuk ◽  
Nivânia Pereira da Costa ◽  
Francisco Miguel de Assis Filho ◽  
Gilvan Pio-Ribeiro ◽  
...  
Keyword(s):  
Tomato Spotted Wilt Virus ◽  
Enzyme Linked Immunosorbent Assay ◽  
Mechanical Inoculation ◽  
Rt Pcr ◽  
Chain Reaction ◽  
Breeding Lines ◽  
Tomato Spotted Wilt ◽  
Ic 10 ◽  
Polymerase Chain ◽  
Wilt Virus

The objective of this work was to evaluate the reactions of three peanut breeding lines (IC-10, IC-34, and ICGV 86388) to Tomato spotted wilt virus (TSWV) by mechanical and thrips inoculation, under greenhouse conditions, and compare them to the reactions of cultivars SunOleic, Georgia Green, and the breeding line C11-2-39. TSWV infection by mechanical inoculation was visually assessed using an index ranging from 0 (no symptoms) to 4 (apical death). Enzyme-linked immunosorbent assay was used to confirm TSWV infection from both mechanical and thrips inoculations. IC-10, IC-34, ICGV 86388, and C11-2-39 were more resistant than the cultivars SunOleic and Georgia Green based on mechanical inoculation. Upon thrips inoculation only IC-34 and ICGV-86388 were infected by TSWV, as demonstrated by reverse transcription polymerase chain reaction (RT-PCR), although no symptoms of infection were observed. The peanut breeding lines IC-10, IC-34, and ICGV 86388 show higher level of resistance to TSWV than cultivar Georgia Green considered a standard for TSWV resistance.

scholarly journals Interference Between D and M Types of Plum pox virus in Japanese Plum Assessed by Specific Monoclonal Antibodies and Quantitative Real-Time Reverse Transcription-Polymerase Chain Reaction

2006 ◽  
Vol 96 (3) ◽  
pp. 320-325 ◽  
Author(s):  
Nieves Capote ◽  
M. Teresa Gorris ◽  
M. Carmen Martínez ◽  
Margarita Asensio ◽  
Antonio Olmos ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Monoclonal Antibodies ◽  
Real Time ◽  
Reverse Transcription ◽  
Enzyme Linked Immunosorbent Assay ◽  
Plum Pox Virus ◽  
Rt Pcr ◽  
Chain Reaction ◽  
Japanese Plum ◽  
Polymerase Chain

The dynamics of virus interference between two isolates of Plum pox virus (PPV) belonging to the main PPV types, D and M, were analyzed in Japanese plum (Prunus salicina) by challenge inoculations. To assess the consequences of a PPV-M infection on plum already infected with PPV-D, and vice versa (predominance of one of the strains, recombination, synergism, symptoms aggravation, and so on), 30 Japanese plum trees were graft inoculated with PPV-D or PPV-M isolates in quarantine conditions. One year postinoculation, in the event that the inoculated isolates were detected in the whole plant, a second challenge inoculation (PPV-M or PPV-D, respectively) was performed by grafting. The presence of PPV-D, PPV-M, or both was monitored for 7 years by double-antibody sandwich indirect enzyme-linked immunosorbent assay using specific monoclonal antibodies. Reverse transcription-polymerase chain reaction (RT-PCR) with D- and M-specific primers confirmed the serological typing. Real-time RT-PCR assays were performed using D- and M-specific fluorescent 3′ minor groove binder-DNA probes, which were able to detect and quantify PPV populations in the inoculated plants with greater precision. The presence of PPV-D in Japanese plum did not cross-protect the trees against PPV-M infection. In PPV-D-infected plants, the PPV-M strain used as challenge inoculum behaved differently depending on the plum cultivar assayed. In cv. Black Diamond, PPV-M invaded the plant progressively, displacing the previous PPV-D population; whereas, in cv. Sun Gold, both PPV isolates coexisted in the plant. In contrast, the PPV-D isolate used was unable to infect plants of both cultivars in which a PPV-M population already was established. After 7 years, no synergism was observed and no recombination event between PPV-D and PPV-M genomes was detected.

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