scholarly journals Molecular characterization of Anopheles fluviatilis species complex in the Islamic Republic of Iran

2021 ◽  
Vol 9 (3) ◽  
pp. 257-265
Author(s):  
S. R. Naddaf ◽  
M. A. Oshaghi ◽  
H. Vatandoost ◽  
M. Assmar
Keyword(s):  
Sequence Data ◽  
Islamic Republic ◽  
Anopheles Fluviatilis ◽  
As Species ◽  
Pcr Product ◽  
Islamic Republic Of Iran ◽  
Polymerase Chain ◽  
Second Internal Transcribed Spacer ◽  
Species Specific

A species-specific polymerase chain reaction [PCR] assay was used to identify the species composition of the Anopheles fluviatilis complex in the Islamic Republic of Iran. All the amplified DNA samples from specimens collected from different areas yielded a fragment of 450 bp size, a PCR product corresponding to that of the species denoted as Y. The sequence data from 21 ITS2 [second internal transcribed spacer] regions were compared with those publicly available in the GenBank database and confirmed that the specimens were 100% identical to species Y of India. Species Y is presumably the same as species T that has no role in transmission of malaria in India, whereas An. fluviatilis is known as a secondary vector of malaria in the Islamic Republic of Iran

scholarly journals Characterization of Leishmania parasites isolated from provinces of the Islamic Republic of Iran

2002 ◽  
Vol 8 (2-3) ◽  
pp. 338-344
Author(s):  
H. Motazedian ◽  
B. Noamanpoor ◽  
S. Ardehali
Keyword(s):  
Random Amplified Polymorphic Dna ◽  
Cutaneous Lesion ◽  
Recent Change ◽  
Islamic Republic ◽  
Cutaneous Lesions ◽  
Rapd Pcr ◽  
Islamic Republic Of Iran ◽  
Polymerase Chain ◽  
Leishmania Parasites

Leishmania parasites isolated in the Islamic Republic of Iran were studied by a random amplified polymorphic DNA polymerase chain reaction [RAPD-PCR]. Of 82 isolates, 80 were from cutaneous lesions, 1 from a human throat lesion and 1 from a dog. Of these, 42 isolates were L. tropica, 36 were L. major and 2 were L. infantum. There were 2 unidentified isolates [from the throat lesion and a cutaneous lesion] and these demonstrated 52% and 48% similarity with L. tropica and L. infantum. Both L. tropica and L. major were isolated from four provinces indicating a recent change in the epidemiology of cutaneous leishmaniasis. L. tropica was isolated from three provinces; L. major from one province. L. infantum was isolated from a human cutaneous lesion and from a dog in Bushehr province.

scholarly journals Identification and Characterization of Benomyl-Resistant and -Sensitive Populations of Colletotrichum gloeosporioides from Statice (Limonium spp.)

2006 ◽  
Vol 96 (5) ◽  
pp. 542-548 ◽  
Author(s):  
Marcel Maymon ◽  
Aida Zveibil ◽  
Shimon Pivonia ◽  
Dror Minz ◽  
Stanley Freeman
Keyword(s):  
Colletotrichum Gloeosporioides ◽  
Sequence Analyses ◽  
Specific Primers ◽  
Tubulin Genes ◽  
Colletotrichum Spp ◽  
Polymerase Chain ◽  
Species Specific ◽  
Identification And Characterization ◽  
Propagation Material

Sixty-four isolates of Colletotrichum gloeosporioides were isolated from infected Limonium spp. cultivated in 12 different locations in Israel. All isolates were identified as belonging to the C. gloeosporioides complex by species-specific primers. Of these isolates, 46 were resistant to benomyl at 10 μg/ml and 18 were sensitive to this concentration of fungicide. Based on arbitrarily primed polymerase chain reaction of all isolates and internal transcribed spacer-1 sequence analyses of 12 selected isolates, the benomyl-resistant and -sensitive populations belong to two distinct genotypes. Sequence analyses of the β-tubulin genes, TUB1 and TUB2, of five sensitive and five resistant representative isolates of C. gloeosporioides from Limonium spp. revealed that the benomyl-resistant isolates had an alanine substitute instead of a glutamic acid at position 198 in TUB2. All data suggest that the resistant and sensitive genotypes are two independent and separate populations. Because all Limonium plant propagation material is imported from various geographic regions worldwide, and benomyl is not applied to this crop or for the control of Colletotrichum spp. in Israel, it is presumed that plants are bearing quiescent infections from the points of origin prior to arrival.

scholarly journals An Assessment of the Molecular Diversity of Ticks and Tick-Borne Microorganisms of Small Ruminants in Pakistan

2020 ◽  
Vol 8 (9) ◽  
pp. 1428 ◽  
Author(s):  
Abdul Ghafar ◽  
Adil Khan ◽  
Alejandro Cabezas-Cruz ◽  
Charles G. Gauci ◽  
Sadaf Niaz ◽  
...  
Keyword(s):  
Sequence Data ◽  
Phylogenetic Analyses ◽  
Strong Support ◽  
Small Ruminants ◽  
Ixodid Ticks ◽  
Nucleotide Sequence Data ◽  
Health Significance ◽  
Second Internal Transcribed Spacer ◽  
Theileria Spp

This study investigated ticks and tick-borne microorganisms of small ruminants from five districts of the Federally Administered Tribal Area (FATA) of Pakistan. Morphological (n = 104) and molecular (n = 54) characterization of the ticks revealed the presence of six ixodid ticks: Rhipicephalus (Rh.) haemaphysaloides, Rh. microplus, Rh. turanicus, Haemaphysalis (Hs.) punctata, Hs. sulcata and Hyalomma anatolicum. Phylogenetic analyses of nucleotide sequence data for two mitochondrial (16S and cytochrome c oxidase 1) and one nuclear (second internal transcribed spacer) DNA regions provided strong support for the grouping of the six tick species identified in this study. Microfluidic real-time PCR, employing multiple pre-validated nuclear and mitochondrial genetic markers, detected 11 potential pathogens and endosymbionts in 72.2% of the ticks (n = 54) tested. Rickettsia (R.) massiliae was the most common pathogen found (42.6% of ticks) followed by Theileria spp. (33.3%), Anaplasma (A.) ovis and R. slovaca (25.9% each). Anaplasma centrale, A. marginale, Ehrlichia spp., R. aeschlimannii, R. conorii and endosymbionts (Francisella- and Coxiella-like) were detected at much lower rates (1.9–22.2%) in ticks. Ticks from goats (83.9%) carried significantly higher microorganisms than those from sheep (56.5%). This study demonstrates that ticks of small ruminants from the FATA are carrying multiple microorganisms of veterinary and medical health significance and provides the basis for future investigations of ticks and tick-borne diseases of animals and humans in this and neighboring regions.

scholarly journals Molecular characterization of a Toxocara variant from cats in Kuala Lumpur, Malaysia

Parasitology ◽  
1998 ◽  
Vol 117 (2) ◽  
pp. 155-164 ◽  
Author(s):  
X. Q. ZHU ◽  
D. E. JACOBS ◽  
N. B. CHILTON ◽  
R. A. SANI ◽  
N. A. B. Y. CHENG ◽  
...  
Keyword(s):  
Internal Transcribed Spacer ◽  
Sequence Data ◽  
Distinct Species ◽  
Single Strand Conformation Polymorphism ◽  
Toxocara Canis ◽  
Nuclear Ribosomal Dna ◽  
Kuala Lumpur ◽  
Second Internal Transcribed Spacer ◽  
Conformation Polymorphism

The ascaridoid nematode of cats from Kuala Lumpur, Malaysia, previously identified morphologically as Toxocara canis, was characterized using a molecular approach. The nuclear ribosomal DNA (rDNA) region spanning the first internal transcribed spacer (ITS-1), the 5·8S gene and the second internal transcribed spacer (ITS-2) was amplified and sequenced. The sequences for the parasite from Malaysian cats were compared with those for T. canis and T. cati. The sequence data showed that this taxon was genetically more similar to T. cati than to T. canis in the ITS-1, 5·8S and ITS-2. Differences in the ITS-1 and ITS-2 sequences between the taxa (9·4–26·1%) were markedly higher than variation between samples within T. canis and T. cati (0–2·9%). The sequence data demonstrate that the parasite from Malaysian cats is neither T. canis nor T. cati and indicate that it is a distinct species. Based on these data, PCR-linked restriction fragment length polymorphism (RFLP) and single-strand conformation polymorphism (SSCP) methods were employed for the unequivocal differentiation of the Toxocara variant from T. canis and T. cati. These methods should provide valuable tools for studying the life-cycle, transmission pattern(s) and zoonotic potential of this parasite.

scholarly journals Assessment of metagenomic sequencing and qPCR for detection of influenza D virus in bovine respiratory tract samples

2020 ◽  
Author(s):  
Maodong Zhang ◽  
Yanyun Huang ◽  
Dale L. Godson ◽  
Champika Fernando ◽  
Trevor W. Alexander ◽  
...  
Keyword(s):  
Respiratory Tract ◽  
High Throughput Sequencing ◽  
Sequence Data ◽  
Metagenomic Sequencing ◽  
Nanopore Sequencing ◽  
Sequence Detection ◽  
Nasal Swabs ◽  
Sequencing Platforms ◽  
Species Specific

AbstractHigh throughput sequencing is currently revolutionizing the genomics field and providing new approaches to the detection and characterization of microorganisms. The objective of this study was to assess the detection of influenza D virus (IDV) in bovine respiratory tract samples using two sequencing platforms (MiSeq and Nanopore (GridION)), and species-specific qPCR. An IDV-specific qPCR was performed on 232 samples (116 nasal swabs and 116 tracheal washes) that had been previously subject to virome sequencing using MiSeq. Nanopore sequencing was performed on 19 samples positive for IDV by either MiSeq or qPCR. Nanopore sequence data was analyzed by two bioinformatics methods: What’s In My Pot (WIMP, on the EPI2ME platform), and an in-house developed analysis pipeline. The agreement of IDV detection between qPCR and MiSeq was 82.3%, between qPCR and Nanopore was 57.9% (in-house) and 84.2% (WIMP), and between MiSeq and Nanopore was 89.5% (in-house) and 73.7% (WIMP). IDV was detected by MiSeq in 14 of 17 IDV qPCR-positive samples with Cq (cycle quantification) values below 31, despite multiplexing 50 samples for sequencing. When qPCR was regarded as the gold standard, the sensitivity and specificity of MiSeq sequence detection were 28.3% and 98.9%, respectively. We conclude that both MiSeq and Nanopore sequencing are capable of detecting IDV in clinical specimens with a range of Cq values. Sensitivity may be further improved by optimizing sequence data analysis, improving virus enrichment, or reducing the degree of multiplexing.

scholarly journals HOT PHENOL EXTRACTION OF TOTAL RNA FROM Thermoascus aurantiacus AND CHARACTERIZATION OF ITS THERMOSTABLE XYLANASE GENE

2016 ◽  
Vol 1 (1) ◽  
pp. 55-58
Author(s):  
Ang Chung Huap ◽  
Awang Ahmad Sallehin Awang Husaini ◽  
Hairul Azman Roslan
Keyword(s):  
Reverse Transcriptase ◽  
28S Rrna ◽  
Thermoascus Aurantiacus ◽  
Xylanase Gene ◽  
Base Pairs ◽  
Total Rna ◽  
Phenol Extraction ◽  
Pcr Product ◽  
Polymerase Chain

Total RNA was successfully isolated using hot phenol extraction method. Three bands representing the 18S, 5.8S and 28S rRNA was observed. No heavy smearing was observed in the RNA band patterns, indicating low levels of polysaccharide contamination, when subjected to 1% agarose gel electrophoresis. Genomic DNA was eliminated using DNase I digestion and lithium chloride (LiCl) precipitation. Two-steps reverse transcriptase polymerase chain reaction (RT-PCR) using M-MuLV Reverse Transcriptase and sequence specific primers for xylanase gene, XynA(F) and XynA(R), successfully generated the target amplicon of 500 base pairs (bp). Sequence analysis of the PCR product indicated as partial sequence of Thermoascus aurantiacus xylanase gene (XynA) deposited in the NCBI GenBank with accession number: AF127529.1 and AJ132635.1. Hot phenol extraction is useful for extracting large quantities of total RNA sufficient for complementary DNA (cDNA) synthesis in shorter period of time.

scholarly journals Species-Specific Polymerase Chain Reaction Amplification of Camel (Camelus) DNA Extracts

2005 ◽  
Vol 88 (5) ◽  
pp. 1394-1398 ◽  
Author(s):  
Ying Chen ◽  
Ya-Jun Wu ◽  
Bao-Liang Xu ◽  
Jing Wan ◽  
Zeng-Ming Qian
Keyword(s):  
Polymerase Chain Reaction ◽  
Polymerase Chain Reaction Amplification ◽  
Chain Reaction ◽  
Pcr Product ◽  
Mitochondrial Cytochrome B ◽  
Base Pair Fragment ◽  
Sensitive Polymerase Chain Reaction ◽  
Pcr Method ◽  
Polymerase Chain ◽  
Species Specific

Abstract A sensitive polymerase chain reaction (PCR) method based on amplification of a specific DNA fragment was established for the identification of camel (Camelus) materials. The species-specific primer pair L183/H372 was designed based on the nucleotide sequence of the mitochondrial cytochrome b gene, and its specificity was confirmed by amplification of 3 camel (domestic double-humped camel, wild double-humped camel, wild one-humped camel) samples and 11 non-Camelus animal (sheep, goat, pig, chicken, cattle, fish, dog, horse, donkey, deer, and rabbit) materials. An expected 208 base pair fragment was amplified from camel materials; no cross-reactive or additional fragments were generated from other animal materials. Taq I restriction endonuclease digestion of the unpurified PCR product can be used routinely to confirm the camel origin of the amplified sequence.

scholarly journals A Multiplex PCR System for the Specific Detection of Cylindrocarpon liriodendri, C. macrodidymum, and C. pauciseptatum from Grapevine

Plant Disease ◽  
2009 ◽  
Vol 93 (8) ◽  
pp. 821-825 ◽  
Author(s):  
Sandra Alaniz ◽  
Josep Armengol ◽  
José García-Jiménez ◽  
Paloma Abad-Campos ◽  
Maela León
Keyword(s):  
Multiplex Pcr ◽  
Extraction Methods ◽  
Specific Pcr ◽  
Pcr Product ◽  
Total Dna ◽  
Polymerase Chain ◽  
Foot Disease ◽  
Species Specific ◽  
Black Foot ◽  
Black Foot Disease

Cylindrocarpon liriodendri and C. macrodidymum are the causal agents of grapevine black foot disease. Recently, a third species, C. pauciseptatum, has been isolated from roots of grapevine showing decline symptoms. Currently, reliable identification of isolates of these species through phenotypical characteristics has not been possible. The polymerase chain reaction (PCR)-based method developed in this study allows a quick and easy detection of Cylindrocarpon spp. associated with grapevine. Three primer pairs annealing to variable, partly species-specific sites of the internal transcribed spacer regions amplified species-specific PCR fragments of different sizes in C. liriodendri, C. macrodidymum, and C. pauciseptatum in a multiplex assay with DNA obtained with both quick and traditional extraction methods. They did not generate any PCR product in other fungal trunk pathogens or contaminants commonly associated with grapevines. When universal fungal ITS primers were used in a nested multiplex PCR, the three primer pairs also detected C. liriodendri, C. macrodidymum, and C. pauciseptatum in total DNA extracted from roots of inoculated grapevines. The designed methods can be used for the diagnosis of these fungi from pure culture or infected grapevines.

scholarly journals Assessment of Metagenomic Sequencing and qPCR for Detection of Influenza D Virus in Bovine Respiratory Tract Samples

Viruses ◽  
2020 ◽  
Vol 12 (8) ◽  
pp. 814 ◽  
Author(s):  
Maodong Zhang ◽  
Yanyun Huang ◽  
Dale L. Godson ◽  
Champika Fernando ◽  
Trevor W. Alexander ◽  
...  
Keyword(s):  
Respiratory Tract ◽  
High Throughput Sequencing ◽  
Sequence Data ◽  
Metagenomic Sequencing ◽  
Nanopore Sequencing ◽  
Sequence Detection ◽  
Nasal Swabs ◽  
Sequencing Platforms ◽  
Species Specific

High throughput sequencing is currently revolutionizing the genomics field and providing new approaches to the detection and characterization of microorganisms. The objective of this study was to assess the detection of influenza D virus (IDV) in bovine respiratory tract samples using two sequencing platforms (MiSeq and Nanopore (GridION)), and species-specific qPCR. An IDV-specific qPCR was performed on 232 samples (116 nasal swabs and 116 tracheal washes) that had been previously subject to virome sequencing using MiSeq. Nanopore sequencing was performed on 19 samples positive for IDV by either MiSeq or qPCR. Nanopore sequence data was analyzed by two bioinformatics methods: What’s In My Pot (WIMP, on the EPI2ME platform), and an in-house developed analysis pipeline. The agreement of IDV detection between qPCR and MiSeq was 82.3%, between qPCR and Nanopore was 57.9% (in-house) and 84.2% (WIMP), and between MiSeq and Nanopore was 89.5% (in-house) and 73.7% (WIMP). IDV was detected by MiSeq in 14 of 17 IDV qPCR-positive samples with Cq (cycle quantification) values below 31, despite multiplexing 50 samples for sequencing. When qPCR was regarded as the gold standard, the sensitivity and specificity of MiSeq sequence detection were 28.3% and 98.9%, respectively. We conclude that both MiSeq and Nanopore sequencing are capable of detecting IDV in clinical specimens with a range of Cq values. Sensitivity may be further improved by optimizing sequence data analysis, improving virus enrichment, or reducing the degree of multiplexing.

Cultural and molecular characterization of photobionts of Peltigera Membranacea

1997 ◽  
Vol 29 (6) ◽  
pp. 571-586 ◽  
Author(s):  
V.P.W. Mioa ◽  
A. Rabenau ◽  
A. Lee
Keyword(s):  
16S Rdna ◽  
Ribosomal Rna ◽  
Molecular Study ◽  
16S Ribosomal Rna ◽  
Chain Reaction ◽  
Ribosomal Rna Gene ◽  
Pcr Product ◽  
16S Ribosomal Rna Gene ◽  
Polymerase Chain

AbstractA molecular study was undertaken to clarify the identity of the photobiont in colourmorphs of the lichen, Peltigera membranacea. Two strains of cyanobacteria, identified as Nosroc sp. by morphology, were cultivated from each of two lichen specimens. Prokaryotic (16S) ribosomal RNA gene fragments were amplified by the polymerase chain reaction (PCR) from DNA extracted from the isolated strains and the lichens, and sequenced directly. Sequences were 98 1% identical between lichen specimens, TDI#AR94 and TDI#AR95, and highly similar to sequences published, or generated in this study from a type culture, for Nostoc. The 16S ribosomal RNA gene sequences (‘ 16S rDNA’) of all four lichen-derived cyanobacteria appeared the same, even though the lichen specimens from which they originated had different sequences. The 16S rDXA from strains 9A and 9B were different from that of specimen TDI#AR94, the thallus from which they were isolated, and instead were the same as that of strains 10A and 10B, and their source, specimen TDI#AR95. When primers selective for the strain 9A sequence were used, however, a small amount of PCR product corresponding to the 16S rDNA of strain 9A was obtained from lichen TDI#AR94. The results confirm that the photobionts of P. membranacea belong to Nostoc, and suggest that genetic differences in the photobiont may be a factor in the occurrence of colourmorphs among cyanolichens.

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