scholarly journals Real-time PCRs for detection of Trichomonas vaginalis β-tubulin and 18S rRNA genes in female genital specimens

2007 ◽  
Vol 56 (6) ◽  
pp. 772-777 ◽  
Author(s):  
Paul Simpson ◽  
Geoff Higgins ◽  
Ming Qiao ◽  
Russell Waddell ◽  
Tuckweng Kok
Keyword(s):  
Real Time ◽  
Trichomonas Vaginalis ◽  
18S Rrna ◽  
Clinical Manifestations ◽  
Developed Countries ◽  
Rrna Genes ◽  
Rrna Gene ◽  
Wet Mount ◽  
Increased Risk ◽  
18S Rrna Genes

Trichomonas vaginalis is the cause of one of the most common types of vaginitis, trichomoniasis. The incidence of trichomoniasis in developed countries has decreased substantially during the past decade, but high prevalence of this disease can still be found in rural and remote areas of Australia. Clinical manifestations of symptomatic women are generally non-specific, but include vaginal discharge, vaginitis and irritation. T. vaginalis infection has also been linked to the increased risk of human immunodeficiency virus transmission. Current diagnosis of T. vaginalis relies on the visualization of motile organisms in a wet-mount preparation. Culture is used mainly in reference laboratories. The latter two methods require viable organisms and would not be suitable for use where transportation of specimens can be delayed. Two real-time fluorescence resonance energy transfer (FRET) hybridization probe PCR assays were used in this study to test for T. vaginalis DNA, targeting the β-tubulin and 18S rRNA genes. We tested 500 randomly selected female patients, in an STD setting, for T. vaginalis DNA. The FRET PCRs targeting the β-tubulin gene and the 18S rRNA gene detected 96 % (85/89) and 100 % (89/89) , respectively, of the positive specimens (first-void urine sample or genital swabs). Wet-mount microscopy was performed on 76 of these PCR-positive specimens and showed a sensitivity of 38 % (29/76). The prevalence, by PCR, of trichomoniasis was 18 % in this study. The two real-time PCRs developed in this study, targeting different genetic regions of the organism, provide a rapid, sensitive and specific diagnosis of T. vaginalis infection.

scholarly journals Crenosoma striatum in lungs of European hedgehogs (Erinaceus europeus) from Portugal

2020 ◽  
Vol 57 (2) ◽  
pp. 179-184
Author(s):  
P. F. Barradas ◽  
A. R. Flores ◽  
T. L. Mateus ◽  
F. Carvalho ◽  
F. Gärtner ◽  
...  
Keyword(s):  
Molecular Characterization ◽  
18S Rrna ◽  
Population Sample ◽  
Genetic Data ◽  
Rrna Genes ◽  
12S Rrna ◽  
Rrna Gene ◽  
Mitochondrial 12S Rrna ◽  
18S Rrna Genes

SummaryCrenosoma striatum is a host-specifi c metastrongiloid nematode causing respiratory tract disease in hedgehogs (Erinaceus europaeus). Since few studies have reported C. striatum in hedgehogs and little genetic data is available concerning this lungworm, this study aimed to determine the occurrence of C. striatum in a population sample of hedgehogs from Portugal, additionally providing morphological, histological and molecular data. From 2017 to 2018 a survey of infection was carried out in 11 necropsied hedgehogs. Worms were extracted from fresh lung tissues and microscopically evaluated. Molecular characterization of partial mitochondrial (12S rRNA) and nuclear (18S rRNA) genes was performed. The presence of lungworms in pulmonary tissues of five hedgehogs (45.5%) was detected. Morphological and histopathological analyses evidenced adult forms of nematodes consistent with C. striatum. Molecular characterization of 18S rRNA genes confirmed the classifi cation as C. striatum. Also, novel genetic data characterizing the mitochondrial (12S rRNA) gene of C. striatum is presented.This is the first report of C. striatum infection in hedgehogs of Portugal. The findings here reported provide new insights regarding the geographic distribution and the molecular identification of this lungworm species.

scholarly journals Design and Validation of Four New Primers for Next-Generation Sequencing To Target the 18S rRNA Genes of Gastrointestinal Ciliate Protozoa

2014 ◽  
Vol 80 (17) ◽  
pp. 5515-5521 ◽  
Author(s):  
Suzanne L. Ishaq ◽  
André-Denis G. Wright
Keyword(s):  
18S Rrna ◽  
18S Rrna Gene ◽  
Rrna Genes ◽  
Rrna Gene ◽  
Content Type ◽  
Hypervariable Regions ◽  
Blast Database ◽  
Primer Sets ◽  
18S Rrna Genes ◽  
Ciliate Protozoa

ABSTRACTFour new primers and one published primer were used to PCR amplify hypervariable regions within the protozoal 18S rRNA gene to determine which primer pair provided the best identification and statistical analysis. PCR amplicons of 394 to 498 bases were generated from three primer sets, sequenced using Roche 454 pyrosequencing with Titanium, and analyzed using the BLAST database (NCBI) and MOTHUR version 1.29. The protozoal diversity of rumen contents from moose in Alaska was assessed. In the present study, primer set 1, P-SSU-316F and GIC758R (amplicon of 482 bases), gave the best representation of diversity using BLAST classification, and the set amplifiedEntodinium simplexandOstracodiniumspp., which were not amplified by the other two primer sets. Primer set 2, GIC1080F and GIC1578R (amplicon of 498 bases), had similar BLAST results and a slightly higher percentage of sequences that were identified with a higher sequence identity. Primer sets 1 and 2 are recommended for use in ruminants. However, primer set 1 may be inadequate to determine protozoal diversity in nonruminants. The amplicons created by primer set 1 were indistinguishable for certain species within the generaBandia,Blepharocorys,Polycosta, andTetratoxumand betweenHemiprorodon gymnoprosthiumandProrodonopsiscoli, none of which are normally found in the rumen.

Molecular identification ofSarcocystisspp. helped to define the origin of green pythons (Morelia viridis) confiscated in Germany

Parasitology ◽  
2013 ◽  
Vol 141 (5) ◽  
pp. 646-651 ◽  
Author(s):  
GASTÓN MORÉ ◽  
NIKOLA PANTCHEV ◽  
DALAND C. HERRMANN ◽  
MAJDA GLOBOKAR VRHOVEC ◽  
SABINE ÖFNER ◽  
...  
Keyword(s):  
18S Rrna ◽  
Sequence Data ◽  
18S Rrna Gene ◽  
Rrna Genes ◽  
Rrna Gene ◽  
Intermediate Hosts ◽  
Apicomplexan Parasites ◽  
Large Numbers ◽  
Wide Range ◽  
18S Rrna Genes

SUMMARYSarcocystisspp. represent apicomplexan parasites. They usually have a heteroxenous life cycle. Around 200 species have been described, affecting a wide range of animals worldwide, including reptiles. In recent years, large numbers of reptiles have been imported into Europe as pets and, as a consequence, animal welfare and species protection issues emerged. A sample of pooled feces from four confiscated green pythons (Morelia viridis) containingSarcocystisspp. sporocysts was investigated. These snakes were imported for the pet trade and declared as being captive-bred. Full length 18S rRNA genes were amplified, cloned into plasmids and sequenced. Two differentSarcocystisspp. sequences were identified and registered asSarcocystissp. fromM. viridisin GenBank. Both showed a 95–97% sequence identity with the 18S rRNA gene ofSarcocystis singaporensis.Phylogenetic analysis positioned these sequences together with otherSarcocystisspp. from snakes and rodents as definitive and intermediate hosts (IH), respectively. Sequence data and also the results of clinical and parasitological examinations suggest that the snakes were definitive hosts forSarcocystisspp. that circulate in wild IH. Thus, it seems unlikely that the infected snakes had been legally bred. Our research shows that information on the infection of snakes withSarcocystisspp. may be used to assess compliance with regulations on the trade with wildlife species.

Dogs are reservoir hosts for possible transmission of human strongyloidiasis in Thailand: molecular identification and genetic diversity of causative parasite species

2019 ◽  
Vol 94 ◽  
Author(s):  
O. Sanpool ◽  
P.M. Intapan ◽  
R. Rodpai ◽  
P. Laoraksawong ◽  
L. Sadaow ◽  
...  
Keyword(s):  
18S Rrna ◽  
Gastrointestinal Disease ◽  
Close Association ◽  
Lao Pdr ◽  
Strongyloides Stercoralis ◽  
18S Rrna Gene ◽  
Rrna Genes ◽  
Rrna Gene ◽  
North East ◽  
18S Rrna Genes

Abstract Human strongyloidiasis is a deleterious gastrointestinal disease mainly caused by Strongyloides stercoralis infection. We aimed to study the possible transmission of S. stercoralis between humans and pet animals. We isolated Strongyloides from humans and domestic dogs in the same rural community in north-east Thailand and compared the nucleotide sequences of derived worms using portions of the mitochondrial cytochrome c oxidase subunit 1 (cox1) and 18S ribosomal RNA (18S rRNA) genes. Twenty-eight sequences from the 18S rRNA gene were obtained from worms derived from humans (n = 23) and dogs (n = 5), and were identical with S. stercoralis sequences (from Thailand, Cambodia, Lao PDR and Myanmar) published in the GenBank database. The 28 cox1 sequences from humans and dogs showed high similarity to each other. The available published cox1 sequences (n = 150), in combination with our 28 sequences, represented 68 haplotypes distributed among four clusters. The 28 samples from the present study represented eight haplotypes including four new haplotypes. Dogs and humans shared the same haplotypes, suggesting the possibility of zoonotic transmission from pet dogs to humans. This is of concern since dogs and humans live in close association with each other.

scholarly journals 18S rRNA Gene Variation among Common Airborne Fungi, and Development of Specific Oligonucleotide Probes for the Detection of Fungal Isolates

2003 ◽  
Vol 69 (9) ◽  
pp. 5389-5397 ◽  
Author(s):  
Zhihong Wu ◽  
Yoshihiko Tsumura ◽  
Göran Blomquist ◽  
Xiao-Ru Wang
Keyword(s):  
18S Rrna ◽  
Airborne Fungi ◽  
18S Rrna Gene ◽  
Rrna Genes ◽  
Molecular Diagnostic ◽  
Rrna Gene ◽  
Oligonucleotide Probes ◽  
Gene Variation ◽  
Species Specific ◽  
18S Rrna Genes

ABSTRACT In this study, we sequenced 18S rRNA genes (rDNA) from 49 fungal strains representing 31 species from 15 genera. Most of these species are common airborne fungi and pathogens that may cause various public health concerns. Sequence analysis revealed distinct divergence between Zygomycota and Ascomycota. Within Ascomycota, several strongly supported clades were identified that facilitate the taxonomic placement of several little-studied fungi. Wallemia appeared as the group most diverged from all the other Ascomycota species. Based on the 18S rDNA sequence variation, 108 oligonucleotide probes were designed for each genus and species included in this study. After homology searches and DNA hybridization evaluations, 33 probes were verified as genus or species specific. The optimal hybridization temperatures to achieve the best specificity for these 33 probes were determined. These new probes can contribute to the molecular diagnostic research for environmental monitoring.

Molecular identification and phylogenetic characterisation of Thelohanellus kitauei — short communication

2013 ◽  
Vol 61 (1) ◽  
pp. 30-35 ◽  
Author(s):  
Sang Shin ◽  
Ji Kim ◽  
Casiano Choresca ◽  
Jee Han ◽  
Jin Jun ◽  
...  
Keyword(s):  
18S Rrna ◽  
Short Communication ◽  
Genetic Relationships ◽  
18S Rrna Gene ◽  
Rrna Genes ◽  
Rrna Gene ◽  
Pcr Product ◽  
Cyprinus Carpio Haematopterus ◽  
18S Rrna Genes ◽  
Rrna Sequences

Thelohanellus kitauei was isolated from the koi Cyprinus carpio haematopterus, and the 18S rRNA gene of T. kitauei was amplified by optimised nested-PCR. The PCR product was sequenced and compared with other 18S rRNA genes of Thelohanellus species to investigate the relationships between their host specificities and infection sites. Based on the 18S rRNA sequences, T. kitauei is most closely related to T. hovorkai (which can infect the intestine). Phylogenetic analysis revealed that T. kitauei was clustered with other Thelohanellus spp. infecting Cyprininae. The present study suggests that the infection site and the host specificity (subfamily level) are reflected in the genetic relationships among Thelohanellus species.

scholarly journals Culturing and environmental DNA sequencing uncover hidden kinetoplastid biodiversity and a major marine clade within ancestrally freshwater Neobodo designis

2005 ◽  
Vol 55 (6) ◽  
pp. 2605-2621 ◽  
Author(s):  
Sophie von der Heyden ◽  
Thomas Cavalier-Smith
Keyword(s):  
Genetic Diversity ◽  
Dna Sequences ◽  
18S Rrna ◽  
Environmental Dna ◽  
Pcr Primers ◽  
Rrna Genes ◽  
Rrna Gene ◽  
Free Living ◽  
Marine Habitats ◽  
18S Rrna Genes

Bodonid flagellates (class Kinetoplastea) are abundant, free-living protozoa in freshwater, soil and marine habitats, with undersampled global biodiversity. To investigate overall bodonid diversity, kinetoplastid-specific PCR primers were used to amplify and sequence 18S rRNA genes from DNA extracted from 16 diverse environmental samples; of 39 different kinetoplastid sequences, 35 belong to the subclass Metakinetoplastina, where most group with the genus Neobodo or the species Bodo saltans, whilst four group with the subclass Prokinetoplastina (Ichthyobodo). To study divergence between freshwater and marine members of the genus Neobodo, 26 new Neobodo designis strains were cultured and their 18S rRNA genes were sequenced. It is shown that the morphospecies N. designis is a remarkably ancient species complex with a major marine clade nested among older freshwater clades, suggesting that these lineages were constrained physiologically from moving between these environments for most of their long history. Other major bodonid clades show less-deep separation between marine and freshwater strains, but have extensive genetic diversity within all lineages and an apparently biogeographically distinct distribution of B. saltans subclades. Clade-specific 18S rRNA gene primers were used for two N. designis subclades to test their global distribution and genetic diversity. The non-overlap between environmental DNA sequences and those from cultures suggests that there are hundreds, possibly thousands, of different rRNA gene sequences of free-living bodonids globally.

Phylogenetic relationships of Scelimeninae genera (Orthoptera: Tetrigoidea) based on COI, 16S rRNA and 18S rRNA gene sequences

Zootaxa ◽  
2018 ◽  
Vol 4482 (2) ◽  
pp. 392
Author(s):  
YA-ZHEN CHEN ◽  
WEI-AN DENG ◽  
JIA-MIN WANG ◽  
LI-LIANG LIN ◽  
SHAN-YI ZHOU
Keyword(s):  
16S Rrna ◽  
Genetic Distance ◽  
Phylogenetic Relationships ◽  
18S Rrna ◽  
18S Rrna Gene ◽  
Rrna Genes ◽  
Rrna Gene ◽  
Average Genetic Distance ◽  
Molecular Phylogenetic ◽  
18S Rrna Genes

Scelimeninae is an important subfamily of Tetrigoidea; however, the phylogenetic relationships within Scelimeninae are poorly understood, and its generic classification has remained unstable. In this study, the COI, 16S rRNA and 18S rRNA genes from 24 species in 9 genera within Scelimeninae were amplified and sequenced, the base composition and inter-species genetic distance of the combined sequence of COI, 16S rRNA and 18S rRNA genes were analyzed, and the molecular phylogenetic relationships were reconstructed using Maximum Likelihood (ML) and Bayesian inference (BI) methods. The results of sequence analysis showed that the total length of the combined COI, 16S rRNA and 18S rRNA gene sequence was 3507 bp, including 2345 conservative sites, 1144 variable sites and 901 parsimony-informative sites. The average A+T content was 63.5% and 78.1% in the COI, 16S rRNA sequences, respectively, indicating A+T bias. The average genetic distance between all species was 0.134, and the average genetic distance in the inner group (Scelimeninae) was 0.126. A phylogenetic tree based on the combined sequences of the COI, 16S rRNA and 18S rRNA genes showed that the phylogenetic relationships among 9 Scelimeninae genera were as follows: Criotettix + (((Zhengitettix + Hebarditettix) + (Falconius + (Scelimena + Paragavialidium))) + ((Eucriotettix + Thoradonta) + Loxilobus)). The molecular phylogenetic results generally support the morphological taxonomy; at the genus level, Criotettix, Scelimena, Paragavialidium, Thoradonta and Eucriotettix are monophyletic groups, Scelimena and Paragavialidium form sister groups, and Thoradonta and Eucriotettix also form sister groups, but the relationship between Hebarditettix and Zhengitettix needs further study. At the species level, synonyms may exist between Thoradonta spiculoba and Thoradonta transpicula and Thoradonta nodulosa and Thoradonta obtusilobata, but more studies are required to confirm this inference. 

scholarly journals Detection Moleculer Of Putative 18S rRNA Gen Protozoa Trichodina sp. Infected Larvae Gurami (Osphronemus gouramy L) in Balai Benih Ikan Kutasari Purbalingga Central Java

2021 ◽  
Vol 3 (1) ◽  
pp. 26
Author(s):  
Rokhmani Rokhmani ◽  
Daniel Joko Wahyono ◽  
Lilis Mulyani
Keyword(s):  
18S Rrna ◽  
18S Rrna Gene ◽  
Fish Growth ◽  
Rrna Genes ◽  
Rrna Gene ◽  
Gene Detection ◽  
Central Java ◽  
Polymerase Chain ◽  
18S Rrna Genes

Trichodina spp. are ectoparasitic pathogens of ciliata group that commonly infect both freshwater and marine fish, including gouramy fish. As a result of infection of Trichodina spp. this will lead to inhibition of fish growth and decreased fish production, resulting in low fish selling value. The rate of occurrence of Trichodina spp. that infects gurami can reach 100%. Research has been conducted to determine which one Trichodina spp. Protozoa that infects the gouramy seeds of BBI (Fish Seed Center) Kutasari Purbalingga following detection of 18S RNA gene. Gene detection method used in this research is Polymerase Chain Reaction (PCR) is a technique of DNA synthesis and amplification in vitro. This research is done following these methodes: (1) sampling of Gurami fish with purposive sampling which obtained from BBI Kutasari Purbalingga, (2) isolation of Trichodina spp., (3). Preparation of Trichodina spp. sample and its identification, and (4). Molecular character obervation following detection of 18S rRNA gene. This study obtained 10% percentage of detection of 18S rRNA genes of the species of Trichodina paraheterodentata that infect on the gouramy fish of Purbalingga. The percentage rate of detection of these genes is low when compared with the results of the detection of 18S rRNA Trichodina paraheterodentata gene that infects gouramy fish in Banjarnegara.

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