scholarly journals Genetic Diversity of Babesia bovis MSA-1, MSA-2b and MSA-2c in China

Pathogens ◽  
2020 ◽  
Vol 9 (6) ◽  
pp. 473
Author(s):  
Jinming Wang ◽  
Jifei Yang ◽  
Shandian Gao ◽  
Xiaoxing Wang ◽  
Hao Sun ◽  
...  
Keyword(s):  
Genetic Diversity ◽  
Immune Responses ◽  
Nested Pcr ◽  
Phylogenetic Analyses ◽  
Apicomplexan Parasite ◽  
Amino Acid Sequences ◽  
Babesia Bovis ◽  
Chain Reaction ◽  
Merozoite Surface Antigen ◽  
Polymerase Chain

The apicomplexan parasite Babesia bovis is a tick-borne intracellular hemoprotozoan parasite that is widespread across China. Genetic diversity is an important strategy used by parasites to escape the immune responses of their hosts. In our present study, 575 blood samples, collected from cattle in 10 provinces, were initially screened using a nested PCR (polymerase chain reaction) for detection of B. bovis infection. To perform genetic diversity analyses, positive samples were further amplified to obtain sequences of three B. bovis merozoite surface antigen genes (MSA-1, MSA-2b, MSA-2c). The results of the nested PCR approach showed that an average of 8.9% (51/575) of cattle were positive for B. bovis infection. Phylogenetic analyses of the predicted amino acid sequences revealed that unique antigen variants were formed only by Chinese isolates. Our findings provide vital information for understanding the genetic diversity of B. bovis in China.

scholarly journals Occurrence and Genetic Diversity of Grapevine berry inner necrosis virus from Grapevines in China

Plant Disease ◽  
2017 ◽  
Vol 101 (1) ◽  
pp. 144-149 ◽  
Author(s):  
X. D. Fan ◽  
Z. P. Zhang ◽  
F. Ren ◽  
G. J. Hu ◽  
J. Zhou ◽  
...  
Keyword(s):  
Genetic Diversity ◽  
Movement Protein ◽  
Phylogenetic Analyses ◽  
Amino Acid Sequences ◽  
Chinese Isolate ◽  
Necrosis Virus ◽  
Grapevine Berry ◽  
Group 3 ◽  
Polymerase Chain ◽  
Ring Spot

To investigate the prevalence and genetic diversity of Grapevine berry inner necrosis virus (GINV) in China, 195 grapevine samples from 15 Chinese provinces and regions were tested using reverse-transcription polymerase chain reaction. The samples included symptomatic and asymptomatic cultivars, with 35.9% (70 of 195) of samples testing positive for GINV. Seventeen samples had obvious ring spot symptoms, and 94.1% (16 of 17) tested positive for GINV, suggesting that GINV may be highly associated with the ring spot symptom. The genetic diversity of GINV isolates was analyzed based on the partial nucleotide and amino acid sequences of the coat protein (CP) and movement protein (MP) genes. Phylogenetic analyses of the MP and CP gene sequences divided the GINV isolates into three groups. The majority of the Chinese isolates were in groups 1 and 2, and only one Chinese isolate, along with a previously reported Japanese isolate, was in group 3. This is the first report on the genetic diversity of GINV isolates and their prevalence and distribution in China.

scholarly journals Genetic Diversity Among Banana streak virus Isolates from Australia

2000 ◽  
Vol 90 (8) ◽  
pp. 921-927 ◽  
Author(s):  
A. D. W. Geering ◽  
L. A. McMichael ◽  
R. G. Dietzgen ◽  
J. E. Thomas
Keyword(s):  
Phylogenetic Analyses ◽  
Amino Acid Sequences ◽  
Ribonuclease H ◽  
Streak Virus ◽  
Specific Detection ◽  
Chain Reaction ◽  
Banana Streak Virus ◽  
Reading Frame ◽  
Polymerase Chain ◽  
Virus Isolates

Banana streak virus (BSV) is an important pathogen of bananas and plantains (Musa spp.) throughout the world. We have cloned and sequenced part of the genomes of four isolates of BSV from Australia, designated BSV-RD, BSV-Cav, BSV-Mys, and BSV-GF. These isolates originated from banana cvs. Red Dacca, Williams, Mysore, and Goldfinger, respectively. All clones contained a sequence covering part of open reading frame III and the intergenic region of the badnavirus genome. The sequences were compared with those of other badnaviruses, including BSV-Onne, a previously characterized isolate from Nigeria. The BSV-RD sequence was virtually identical to that of BSV-Onne, differing by only two nucleotides over 1,292 bp. However, BSV-Cav, -Mys, and -GF were divergent in nucleotide sequence. Phylogenetic analyses using conserved sequences in the ribonuclease H domain revealed that all BSV isolates were more closely related to each other than to any other badnavirus. BSV-Cav was most closely related to BSV-Onne, and there was 95.1% identity between the two amino acid sequences. Other relationships between the BSV isolates were less similar, with sequence identities ranging from 66.4 to 78.2%, which is a magnitude comparable to the distance between some of the recognized badnavirus species. Immunocapture-polymerase chain reaction assays have been developed, allowing specific detection and differentiation of the four isolates of BSV.

scholarly journals A PCR-based Assay for Distinguishing between A1 and A2 Mating Types of Phytophthora capsici

2017 ◽  
Vol 142 (4) ◽  
pp. 260-264
Author(s):  
Ping Li ◽  
Dong Liu ◽  
Min Guo ◽  
Yuemin Pan ◽  
Fangxin Chen ◽  
...  
Keyword(s):  
Genetic Diversity ◽  
Mating Type ◽  
Phytophthora Capsici ◽  
Mating Types ◽  
Sequence Repeat ◽  
Chain Reaction ◽  
Plant Parasite ◽  
Dna Fragment ◽  
Polymerase Chain ◽  
Microsatellite Diversity

Sexual reproduction in the plant parasite Phytophthora capsici Leonian requires the interaction of two distinct mating types, A1 and A2. Co-occurrence of these mating types can enhance the genetic diversity of P. capsici and alter its virulence or resistance characteristics. Using an intersimple sequence repeat (ISSR) screen of microsatellite diversity, we identified, cloned, and sequenced a novel 1121-base pair (bp) fragment specific to the A1 mating type of P. capsici. Primers Pcap-1 and Pcap-2 were designed from this DNA fragment to specifically detect the A1 mating type. Polymerase chain reaction (PCR) using these primers amplified an expected 997-bp fragment from known A1 mating types, but yielded a 508-bp fragment from known A2 mating types. This PCR-based assay could be adapted to accurately and rapidly detect the co-occurrence of A1 and A2 P. capsici mating types from field material.

scholarly journals Structurally and functionally conserved regions of cytochrome P-450 reductase as targets for DNA amplification by the polymerase chain reaction. Cloning and nucleotide sequence of the Schizosaccharomyces pombe cDNA

1992 ◽  
Vol 287 (1) ◽  
pp. 195-200 ◽  
Author(s):  
J S Miles
Keyword(s):  
Polymerase Chain Reaction ◽  
Schizosaccharomyces Pombe ◽  
Binding Domain ◽  
Amino Acid Sequences ◽  
Chain Reaction ◽  
Conserved Regions ◽  
Reductase Gene ◽  
Degenerate Oligonucleotide ◽  
Polymerase Chain ◽  
Cytochrome P 450

1. Alignments of the available cytochrome P-450 reductase amino acid sequences, and comparison with the crystal structure of ferredoxin-NADP reductase, indicate that two highly conserved regions are of functional importance. 2. Degenerate oligonucleotide primers, based on these sequences, were used in the polymerase chain reaction to amplify a 309 bp fragment of the cytochrome P-450 reductase gene from Schizosaccharomyces pombe for use as an homologous probe. 3. A 2.6 kb cDNA was cloned from a lambda library, and sequencing revealed an open-reading frame of 2034 bp encoding a protein of M(r) 76774. This protein shares 38-41% identity with other eukaryotic cytochrome P-450 reductases, and 30% identity with that of Bacillus megaterium. 4. Comparison of the N-terminal FMN-binding domain with flavodoxin, and the C-terminal FAD- and NADP-binding domain with ferredoxin-NADP reductase, indicates the presence of several functionally conserved regions. 5. The Sc. pombe cytochrome P-450 reductase gene was shown to contain no introns.

scholarly journals Molecular diagnosis and biochemical studies of tick-borne diseases (anaplasmosis and babesiosis) in Aberdeen Angus Cattle in New Valley, Egypt

2020 ◽  
Vol 13 (9) ◽  
pp. 1884-1891
Author(s):  
Nani Nasreldin ◽  
Rania M. Ewida ◽  
Hatem Hamdon ◽  
Yasser F. Elnaker
Keyword(s):  
Oxidative Stress ◽  
Polymerase Chain Reaction ◽  
Molecular Techniques ◽  
Babesia Bovis ◽  
Blood Parasites ◽  
Chain Reaction ◽  
Infected Female ◽  
Angus Cattle ◽  
Polymerase Chain ◽  
Female Cattle

Background and Aim: Anaplasmosis and babesiosis are tick-borne diseases that threaten livestock production with subsequent considerable economic losses. This study was conducted to diagnose Anaplasma and Babesia infection using molecular techniques in imported Aberdeen Angus cattle imported from Uruguay to El-Kharga Oasis in New Valley, Egypt, and to investigate the effects of disease on some serum biochemical and oxidative stress parameters. Materials and Methods: Blood samples were collected from 31 cattle, 21 diseased and ten apparently normal, of varying ages and sex. The blood was used for the preparation of blood smears, polymerase chain reaction assay, and separation of serum for biochemical investigation. The experimental production farm at the Faculty of Agriculture, New Valley University, was infested with ticks and variable clinical manifestations during the period from December 2017 to March 2018. One calf died of a suspected blood parasite infection. Results: The blood film examination revealed infection by blood parasites in 21 samples. Anaplasma marginale and Babesia bovis were identified in 12 and 14 samples, respectively. A total of 14 samples were examined by polymerase chain reaction (PCR) to make these identifications. Biochemical parameters showed significantly elevated serum alanine aminotransferase, aspartate aminotransferase, total bilirubin (T. Bil), and urea in blood from parasite-infected female cattle and male calves compared with controls. Increased serum total protein, globulin, and creatinine were recorded only in infected female cattle. The blood glucose level was significantly decreased in infected female cattle and male calves compared with controls. Furthermore, albumin and albumin/globulin ratio was significantly reduced in the infected female cattle. Oxidative stress profiles of infected animals showed a significant increase in serum nitric oxide and malondialdehyde, and both total antioxidant capacity and reduced glutathione (GSH) were significantly reduced in comparison with control animals. Conclusion: The incidence of A. marginale and B. bovis infection is high in imported Aberdeen Angus cattle in New Valley Province. PCR methods provide a short-term assessment of disease. An extensive epidemiological survey, employing serology together with molecular genetic methods, monitoring of abundance and distribution of tick vectors, availability of vaccination programs, and tracking of animal transport is also needed for control of blood parasites.

Evaluation of genetic variations at glutenin loci in Korean wheat landraces using allele-specific DNA markers

2014 ◽  
Vol 12 (3) ◽  
pp. 353-356 ◽  
Author(s):  
Jeong Hwan Ahn ◽  
Soo-Kyung Lee ◽  
Chul Soo Park
Keyword(s):  
Genetic Diversity ◽  
Gluten Strength ◽  
Pcr Markers ◽  
Chain Reaction ◽  
Baking Quality ◽  
Allele Specific ◽  
Polymerase Chain ◽  
Wheat Landraces ◽  
Allelic Variations ◽  
Wheat Lines

The allelic variations at glutenin loci could significantly affect the bread baking quality, and specific glutenin alleles might be closely associated with greater gluten strength, which, in turn, is related to superior bread baking quality. In this study, allelic variations at Glu-1, Glu-A3 and Glu-B3 loci were evaluated in 222 Korean wheat landraces using gene-specific polymerase chain reaction (PCR) markers. Ten alleles were identified at Glu-1 loci. Glu-A1c, Glu-B1b, and Glu-D1a or Glu-D1f alleles were predominantly found at the respective loci and their frequencies were 86.5, 87.8 and 96.9 %, respectively. Seven Korean wheat landraces carried the Glu-D1d allele, and only one Korean wheat landrace (IT173162) achieved 10 points for the Glu-1 score. Fifteen alleles were identified at Glu-A3 and Glu-B3 loci; Glu-A3c and Glu-B3d or Glu-B3i alleles were commonly found at the respective loci and their frequencies were 77.0, 33.3 and 37.8 %, respectively. Glu-B3 alleles exhibited the highest genetic diversity than other alleles, while Glu-B1 and Glu-A1 alleles exhibited the lowest genetic diversity than other alleles. Twenty Korean wheat landraces had the Glu-A3d and Glu-B3b, Glu-B3d, Glu-B3f, Glu-B3g or Glu-B3i alleles, which were correlated with superior bread baking quality. Among these wheat lines, two (IT59787 and IT236544) carried the Glu-D1d allele.

Epidemiology of Epizootic Lymphangitis of cart horses in Northern Ethiopia Using Conventional diagnostic Methods and Nested Polymerase Chain Reaction

2019 ◽  
Author(s):  
Birhanu Hadush Abera ◽  
Molla Michaelay ◽  
Habtamu Taddele ◽  
Nigus Abebe ◽  
Abrha Tesfay ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Clinical Examination ◽  
Roc Curve ◽  
Nested Pcr ◽  
Control Measure ◽  
Diagnostic Methods ◽  
Northern Ethiopia ◽  
Chain Reaction ◽  
Nested Polymerase Chain Reaction ◽  
Polymerase Chain

Abstract Background: Epizootic lymphangitis (EL), caused by Histoplasma capsulatum variety farciminosum (HCF) is a contagious chronic disease of equines characterized by development of nodular lesions in the lymph nodes, lymphatic vessels and skin. This disease is the most important diseases of equines in Ethiopia causing a significant economic loss, particularly cart pulling equines. Todate there is no sound diagnostic nor control measure implemented in the country. Furthermore, there is a shortage of data on the epidemiology of the disease in different regions of the country including northern Ethiopia. This study was conducted to investigate the epidemiology of EL in northern Ethiopia using the conventional methods and the nested polymerase chain reaction (PCR). Methods: A total of 191 cart-horses were enrolled and used as sources of pus and blood samples. The blood was used for the extraction of the DNA of HCF from buffy coat for nested PCR while the pus samples were cultured on Sabourauds Dextrose Agar for isolation. Statistical Package for Social Sciences (SPSS) version 21 was used for data analysis by applying logistic regression, receiver operating characteristic (ROC) curve and Cohen’s kappa coefficient test. In addition, the level of agreement between the clinical examination and the nested PCR was evaluated. Results: Infection with HCF was confirmed in 44% (84/191) of the horses using nested PCR. Subclinical infection was observed in 18.18% (22/121) of the apparently healthy horses. Considering nested PCR as a gold standard, the sensitivity and specificity of the clinical examination were 74% and 95%, respectively while the area under the ROC curve (AUR) was 0.83 (95% confidence interval, 0.77, 0.896). Moreover, a moderate (k=0.675) agreement was observed between the nested PCR and clinical examination.Conclusions: The findings of the present study showed the wide spread occurrence of EL in northern Ethiopia and the advantage of the nested PCR in detecting of the infection of HCF even before the clinical symptoms are apparent.

scholarly journals In SituReverse Transcriptase-Nested Polymerase Chain Reaction to Identify Intracellular Nucleic Acids without the Necessity of Dnase Pretreatment and Hybridisation

2001 ◽  
Vol 22 (3) ◽  
pp. 151-158 ◽  
Author(s):  
Mario Menschikowski ◽  
Margot Vogel ◽  
Rolf Eckey ◽  
Gerd Dinnebier ◽  
Werner Jaross
Keyword(s):  
Polymerase Chain Reaction ◽  
Nucleic Acids ◽  
Nested Pcr ◽  
In Situ Hybridisation ◽  
Target Sequence ◽  
Chain Reaction ◽  
Multiple Control ◽  
Nested Polymerase Chain Reaction ◽  
Polymerase Chain

In the present study a protocol of in situ reverse transcriptase‐nested polymerase chain reaction (in situ RT‐nested PCR) was examined based on the following modifications. (i) To exclude false positive signals caused by “DNA repair mechanisms” and “endogenous priming”, a two‐step PCR was applied after reverse transcription. The first step was performed in the presence of extrinsic primers and unlabeled nucleotides with a maximum of PCR cycles possible without destroying the cell morphology. The second step consisted of only one annealing/elongation reaction, the target sequence marked by addition of digoxigenin‐labeled nucleotides and intrinsic primers. (ii) In order to prevent amplifications of genomic DNA nested primer pairs were applied crossing intron sequences. (iii) To minimize the diffusion of PCR products in cells, the extrinsic primers were extended with complementary 5′‐tails. This approach results in the generation of high molecular weight concatamers during PCR cycles. By applying this protocol, immunostainings specific for phospholipase A2 of type IIA mRNA were exclusively detectable in the cytoplasm of HepG2 hepatoma cells, which were used as a model system, whereas the nuclei were unstained. Multiple control experiments yielded completely negative results. These data suggest that the in situ RT‐nested PCR, which in comparison to the method of in situ RT‐PCR‐in situ‐hybridisation is simpler and less time‐consuming, can be used as an alternative approach to identify intracellular nucleic acids.

scholarly journals Broad Diversity of Ralstonia solanacearum Strains in Cameroon

Plant Disease ◽  
2009 ◽  
Vol 93 (11) ◽  
pp. 1123-1130 ◽  
Author(s):  
Gabriel Mahbou Somo Toukam ◽  
Gilles Cellier ◽  
Emmanuel Wicker ◽  
Caroline Guilbaud ◽  
Rémi Kahane ◽  
...  
Keyword(s):  
Genetic Diversity ◽  
Ralstonia Solanacearum ◽  
Central Africa ◽  
Sequence Analyses ◽  
Chain Reaction ◽  
Specific Pcr ◽  
Economic Threat ◽  
The Indian Ocean ◽  
Polymerase Chain ◽  
Reference Strains

In 2005, an extensive survey of bacterial wilt in Cameroon collected 110 strains of Ralstonia solanacearum from wilting tomato, potato, pepper, huckleberry (Solanum scabrum), sesame, and amaranth. The genetic diversity and phylogeny of selected strains from Cameroon were assessed by multiplex–polymerase chain reaction (PCR), race 3/biovar 2–specific PCR, and sequence analyses of the mutS and egl genes. These data were compared with those from 33 reference strains covering the known diversity within the R. solanacearum species complex. Strains isolated in Cameroon clustered into three of the four known phylotypes: I (Asian), II (American), and III (African). Lowland tomato strains belonged to phylotype I and were quite homogeneous. The strains belonging to phylotype II were genetically diverse, and partitioned into subclusters IIA and IIB (sequevar 1, race 3/biovar 2). Cameroon strains in the African phylotype III were distinct from reference strains from Zimbabwe or the Indian Ocean, highlighting the genetic diversity present within this phylotype. Strains from potatoes growing in the highlands of West Cameroon fell into both phylotypes II (race 3/biovar 2) and III. These phylotype II and III highland strains attacked both potato and tomato and could therefore pose an economic threat to potato and tomato crops throughout Central Africa. This is the first comprehensive report on the genetic diversity of R. solanacearum strains in Cameroon.

scholarly journals Genetic diversity of Ehrlichia ruminantium strains in Cameroon

2014 ◽  
Vol 81 (1) ◽  
Author(s):  
Seraphine N. Esemu ◽  
Roland N. Ndip ◽  
Lucy M. Ndip
Keyword(s):  
Genetic Diversity ◽  
Deoxyribonucleic Acid ◽  
Amino Acid Level ◽  
Gene Products ◽  
Chain Reaction ◽  
Nested Polymerase Chain Reaction ◽  
Ehrlichia Ruminantium ◽  
Polymerase Chain ◽  
Study Sites ◽  
Partial Fragment

In order to investigate the extent of genetic diversity among Ehrlichia ruminantium strains in Cameroon, a partial fragment (800 bp) of the E. ruminantium map1 gene was amplified by nested polymerase chain reaction in 121 of 156 E. ruminantium pCS20-positive DNA samples extracted from ticks and cattle collected from two ranches. Deoxyribonucleic acid sequencing of the map1 gene products indicated the presence of at least 21 genotypes at the nucleotide level and 16 genotypes at the amino acid level circulating within the study sites. Some of the genotypes were identical to Antigua (U50830), Blaaukrans (AF368000) or UmBanein (U50835), whilst the others were new genotypes. Twenty-four representative sequences were deposited in GenBank and given accession numbers JX477663 – JX477674 (for sequences of tick origin) and JX486788 – JX486799 (for sequences of cattle origin). Knowledge of E. ruminantium strain diversity could be important in understanding the epidemiology of heartwater

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