scholarly journals Cross-Genera PCR Amplification of DNA from Apicomplexan Parasites

2018 ◽  
Author(s):  
Philippe Gil de Mendonça
Keyword(s):  
Pathogenic Bacteria ◽  
Hypervariable Region ◽  
Pcr Amplification ◽  
18S Rrna Gene ◽  
Rrna Gene ◽  
Apicomplexan Parasites ◽  
Diagnostic Samples ◽  
Successful Amplification ◽  
Babesia Spp ◽  
Theileria Spp

Background: The discovery of an unexpected genetic sequence raised doubts about the specificity of a primer pair targeting Babesia spp. and Theileria spp. This study aimed to check the specificity of this primer pair. Methods: Conventional end-point PCR and real-time PCR protocols using primers targeting the 18S rRNA gene V4 hypervariable region of Babesia spp. and Theileria spp. were tested for potential cross-genera amplification using DNA from a palette of parasitic protists and pathogenic bacteria as a template. These investigations took place at the Ludwig Maximilian University of Munich (Germany) in 2010 as part of the EDEN project. Results: Successful amplification was obtained with DNA from five apicomplexan genera: Babesia, Theileria, Hepa­tozoon, Toxoplasma, and Hammondia. No amplicons were obtained when DNA from Leishmania infantum or bacte­ria within the genera Borrelia, Leptospira or Anaplasma was used as a template. Conclusion: This cross-genera amplification ability is useful for the quick exclusion of many parasite species from PCR negative diagnostic samples. Accurate species identification from PCR positive samples requires genetic se­quencing of the amplicon.

scholarly journals Phylogenetic Analysis of the Hypervariable Region of the 18S rRNA Gene of Cryptosporidium Oocysts in Feces of Canada Geese (Branta canadensis): Evidence for Five Novel Genotypes

2004 ◽  
Vol 70 (1) ◽  
pp. 452-458 ◽  
Author(s):  
Kristen L. Jellison ◽  
Daniel L. Distel ◽  
Harold F. Hemond ◽  
David B. Schauer
Keyword(s):  
Genetic Diversity ◽  
Phylogenetic Analysis ◽  
18S Rrna ◽  
Hypervariable Region ◽  
Pcr Amplification ◽  
The United States ◽  
18S Rrna Gene ◽  
Rrna Gene ◽  
Canada Geese ◽  
Pcr Products

ABSTRACT To assess genetic diversity in Cryptosporidium oocysts from Canada geese, 161 fecal samples from Canada geese in the United States were analyzed. Eleven (6.8%) were positive for Cryptosporidium spp. following nested PCR amplification of the hypervariable region of the 18S rRNA gene. Nine PCR products from geese were cloned and sequenced, and all nine diverged from previously reported Cryptosporidium 18S rRNA gene sequences. Five sequences were very similar or identical to each other but genetically distinct from that of Cryptosporidium baileyi; two were most closely related to, but genetically distinct from, the first five; and two were distinct from any other sequence analyzed. One additional sequence in the hypervariable region of the 18S rRNA gene isolated from a cormorant was identical to that of C. baileyi. Phylogenetic analysis provided evidence for new genotypes of Cryptosporidium species in Canada geese. Results of this study suggest that the taxonomy of Cryptosporidium species in geese is complex and that a more complete understanding of genetic diversity among these parasites will facilitate our understanding of oocyst sources and species in the environment.

Theileria ovis (Piroplasmida: Theileriidae) Detected in Melophagus ovinus (Diptera: Hippoboscoidea) and Ornithodoros lahorensis (Ixodida: Argasidae) Removed From Sheep in Xinjiang, China

2019 ◽  
Vol 57 (2) ◽  
pp. 631-635 ◽  
Author(s):  
Li Zhao ◽  
Jinling Wang ◽  
Yulin Ding ◽  
Kairui Li ◽  
Bo He ◽  
...  
Keyword(s):  
18S Rrna ◽  
18S Rrna Gene ◽  
Economic Losses ◽  
Rrna Gene ◽  
Protozoan Parasites ◽  
Autonomous Region ◽  
Apicomplexan Parasites ◽  
Wide Range ◽  
Melophagus Ovinus ◽  
Theileria Spp

Abstract Theileria spp. are tick-transmitted, intracellular apicomplexan protozoan parasites that infect a wide range of animals and, as such, can cause significant economic losses. The aim of the present study was to detect and analyze apicomplexan parasites from two different ectoparasites that were collected from Xinjiang Uygur Autonomous Region, China. The PCR-based detection of 18S rRNA indicated that Ornithodoros lahorensis specimens from Kashgar, Xinjiang, and Aksu were positive for Theileria spp., as were Melophagus ovinus specimens from Aksu. Meanwhile, phylogenetic analysis, based on the 18S rRNA gene sequences, revealed that the four amplified Theileria sequences could be attributed to T. ovis. To the best of our knowledge, this is the first study to report the detection of T. ovis DNA in M. ovinus and the first molecular identification study to confirm the detection of T. ovis in O. lahorensis in China. Accordingly, the present study extends the known distribution of T. ovis.

scholarly journals Prevalence of Theileria and Babesia species in Tunisian sheep

2016 ◽  
Vol 83 (1) ◽  
Author(s):  
Mohamed R. Rjeibi ◽  
Mohamed A. Darghouth ◽  
Mohamed Gharbi
Keyword(s):  
18S Rrna Gene ◽  
Rrna Gene ◽  
Infection Prevalence ◽  
Northern Tunisia ◽  
Blood Smears ◽  
Blood Smear Examination ◽  
Polymerase Chain ◽  
Babesia Ovis ◽  
Babesia Spp ◽  
Theileria Spp

In this study, the prevalence of Theileria and Babesia species in sheep was assessed with Giemsastained blood smear examination and polymerase chain reaction to identify the different piroplasms in 270 sheep from three Tunisian bioclimatic zones (north, centre, and south). The overall infection prevalence by Babesia spp. and Theileria spp. in Giemsa-stained blood smears was 2.9% (8/270) and 4.8% (13/270) respectively. The molecular results showed that sheep were more often infected by Theileria ovis than Babesia ovis with an overall prevalence of 16.3% (44/270) and 7.8% (21/270) respectively (p = 0.01). The molecular prevalence by Babesia ovis was significantly higher in females than in males (p < 0.05). According to localities B. ovis was found exclusively in sheep from the centre of Tunisia (Kairouan) whereas Theileria ovis was found in all regions. Infections with T. ovis and B. ovis were confirmed by sequencing. The sequence of T. ovis in this study (accession numbers KM924442) falls into the same clade as T. ovis deposited in GenBank. The T. ovis amplicons (KM924442) showed 99%–100% identities with GenBank sequences. Moreover, comparison of the partial sequences of 18S rRNA gene of B. ovis described in this study (KP670199) revealed 99.4% similarity with B. ovis recently reported in northern Tunisia from sheep and goats. Three nucleotides were different at positions 73 (A/T), 417 (A/T), and 420 (G/T). It also had 99% identity with B. ovis from Spain, Turkey and Iraq. The results suggest a high T. ovis prevalence in Tunisia with a decreasing north-south gradient. This could be correlated to the vector tick distribution.

scholarly journals Genetic Diversity of Bovine Hemoprotozoa in South Korea

Pathogens ◽  
2020 ◽  
Vol 9 (9) ◽  
pp. 768
Author(s):  
Dongmi Kwak ◽  
Min-Goo Seo
Keyword(s):  
South Korea ◽  
Clinical Symptoms ◽  
Surface Protein ◽  
18S Rrna Gene ◽  
Rrna Gene ◽  
Theileria Orientalis ◽  
Bovine Theileriosis ◽  
Major Piroplasm Surface Protein ◽  
Theileria Spp ◽  
Epidemiological Information

Tick-borne pathogens cause economically significant diseases in cattle. Theileria spp. are parasitic protozoa and the causative agent of bovine theileriosis. Here we report the distribution and risk factors of bovine Theileria using blood samples taken between 2018 and 2019. Of 737 tested cattle, nine animals (1.2%) were positive for Theileria orientalis infection by 18S rRNA gene amplification. Further analysis of the infected samples using the T. orientalis major piroplasm surface protein (MPSP) gene revealed five different genotypes circulating in the population: Types 1, 2, 3, 7, and N3. To the best of our knowledge, this is the first research to describe the existence of the T. orientalis MPSP genotype N3 in South Korea. Although the prevalence of bovine T. orientalis was low, our study offers data on the geographical distribution and prevalence of bovine Theileria spp. in South Korea. Further studies are warranted to determine the correlation of clinical symptoms with parasite MPSP genotypes. Our data provide epidemiological information to help control bovine theileriosis in South Korea.

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.

scholarly journals Legionella Species Diversity in an Acidic Biofilm Community in Yellowstone National Park

2005 ◽  
Vol 71 (1) ◽  
pp. 507-511 ◽  
Author(s):  
Kathy B. Sheehan ◽  
Joan M. Henson ◽  
Michael J. Ferris
Keyword(s):  
Yellowstone National Park ◽  
National Park ◽  
Pcr Amplification ◽  
18S Rrna Gene ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Rrna Gene ◽  
Thermal Gradients ◽  
Legionella Species ◽  
Legionella Micdadei

ABSTRACT Legionella species are frequently detected in aquatic environments, but their occurrence in extreme, acidic, geothermal habitats has not been explored with cultivation-independent methods. We investigated a predominately eukaryotic algal mat community in a pH 2.7 geothermal stream in Yellowstone National Park for the presence of Legionella and potential host amoebae. Our analyses, using PCR amplification with Legionella-specific primers targeting 16S rRNA genes, detected four known Legionella species, as well as Legionella sequences from species that are not represented in sequence databases, in mat samples and cultivated isolates. The nonrandom occurrence of sequences detected at lower (30�C) and higher (35 to 38�C) temperatures suggests that natural thermal gradients in the stream influence Legionella species distributions in this mat community. We detected only one sequence, Legionella micdadei, from cultivated isolates. We cultured and sequenced partial 18S rRNA gene regions from two potential hosts, Acanthamoeba and Euglena species.

scholarly journals MixedTheileriainfections in free-ranging buffalo herds: implications for diagnosingTheileria parvainfections in Cape buffalo (Syncerus caffer)

Parasitology ◽  
2011 ◽  
Vol 138 (7) ◽  
pp. 884-895 ◽  
Author(s):  
RONEL PIENAAR ◽  
FRED T. POTGIETER ◽  
ABDALLA A. LATIF ◽  
ORIEL M. M. THEKISOE ◽  
BEN J. MANS
Keyword(s):  
South Africa ◽  
Hypervariable Region ◽  
18S Rrna Gene ◽  
Control Measures ◽  
Locked Nucleic Acid ◽  
Rrna Gene ◽  
Mixed Infections ◽  
Endemic Region ◽  
Signal Suppression

SUMMARYBuffalo-adaptedTheileria parvacauses Corridor disease in cattle. Strict control measures therefore apply to the movement of buffalo in South Africa and include mandatory testing of buffalo for the presence ofT. parva. The official test is a real-time hybridization PCR assay that amplifies the V4 hypervariable region of the 18S rRNA gene ofT. parva, T.sp. (buffalo) andT.sp. (bougasvlei). The effect that mixedT. parvaandT.sp. (buffalo)-like infections have on accurateT. parvadiagnosis was investigated in this study.In vitromixed infection simulations indicated PCR signal suppression at 100 to 1000-foldT.sp. (buffalo) excess at lowT. parvaparasitaemia. Suppression of PCR signal was found in field buffalo with mixed infections. TheT. parva-positive status of these cases was confirmed by selective suppression ofT.sp. (buffalo) amplification using a locked nucleic acid clamp and independent assays based on the p67, p104 andTprgenes. The incidence of mixed infections in the Corridor disease endemic region of South Africa is significant, while the prevalence in buffalo outside the endemic area is currently low. A predicted increase ofT.sp. (buffalo)-like infections can affect future diagnoses where mixed infections occur, prompting the need for improvements in current diagnostics.

scholarly journals Molecular Assay Proves the Presence of Theileria annulata Infection in Camels in Al-Diwaniyah Province, Iraq

2021 ◽  
Author(s):  
Noaman N. A'aiz ◽  
Hayder N. Ayyez ◽  
Ahmed J. Neamah
Keyword(s):  
Dna Amplification ◽  
18S Rrna Gene ◽  
Rrna Gene ◽  
Molecular Assay ◽  
Conventional Pcr ◽  
Molecular Investigation ◽  
Causative Agents ◽  
The One ◽  
Specific Species ◽  
Theileria Spp

Background: Theileria camelensis and T. dromedarii are parasitic protozoans reported by several studies as specific species that infect the one-humped camel (Camelus dromedarius). However, other findings casted significant doubts on the true identity of the causative species of theileriosis in camels. Therefore, the present study was conducted to investigate of T. camelensis and T. dromedarii in one humped camels in Iraq during Apr-Oct 2017. Methods: Blood samples for DNA extraction were obtained from 181 slaughtered camels. Molecular investigation was performed following the amplification of 18S rRNA gene by conventional PCR technique. DNA sequencing was then utilized only for the positive samples to confirm the infection with the Theileria species. Results: Nine (4.97%) out of 181 examined samples showed a positive result to infection with Theileria spp., and all these appeared as a T. annulata when subjected to DNA amplification and sequencing techniques. There was a complete absence of any new sequence outside the known species. Conclusion: Most of Theileria infection in camels in the study area is caused by T. annulata and no other causative agents like T. camelensis or T. dromedarii.  

scholarly journals Short-term responses to ocean acidification: effects on relative abundance of eukaryotic plankton from the tropical Timor Sea

2020 ◽  
Author(s):  
Janina Rahlff ◽  
Sahar Khodami ◽  
Lisa Voskuhl ◽  
Matthew P. Humphreys ◽  
Christian Stolle ◽  
...  
Keyword(s):  
Ocean Acidification ◽  
Community Composition ◽  
Hypervariable Region ◽  
18S Rrna Gene ◽  
Marine Biodiversity ◽  
Rrna Gene ◽  
Tropical Regions ◽  
Timor Sea ◽  
Decreasing Ph ◽  
Species Specific

ABSTRACTAnthropogenic carbon dioxide (CO2) emissions drive climate change and pose one of the major challenges of our century. The effects of increased CO2 in the form of ocean acidification (OA) on the communities of marine planktonic eukaryotes in tropical regions such as the Timor Sea are barely understood. Here, we show the effects of high CO2 (pCO2=1823±161 μatm, pHT=7.46±0.05) versus in situ CO2 (pCO2=504±42 μatm, pHT=7.95±0.04) seawater on the community composition of marine planktonic eukaryotes immediately and after 48 hours of treatment exposure in a shipboard microcosm experiment. Illumina sequencing of the V9 hypervariable region of 18S rRNA (gene) was used to study the eukaryotic community composition. Down-regulation of extracellular carbonic anhydrase occurred faster in the high CO2 treatment. Increased CO2 significantly suppressed the relative abundances of different eukaryotic operational taxonomic units (OTUs), including important primary producers. These effects were consistent between abundant (DNA-based) and active (cDNA-based) taxa after 48 hours, e.g., for the diatoms Trieres chinensis and Stephanopyxis turris. Effects were also very species-specific among different diatoms. Planktonic eukaryotes showed adaptation to the CO2 treatment over time, but many OTUs were adversely affected by decreasing pH. OA effects might fundamentally impact the base of marine biodiversity, suggesting profound outcomes for food web functioning in the future ocean.

scholarly journals Isolation of Clostridium Perfringens type C from a Neonatal Yangtze Finless Porpoise (Neophocaena Asiaeorientalis Asiaeorientalis)

2022 ◽  
Author(s):  
Jia Li ◽  
Richard William McLaughlin ◽  
Yingli Liu ◽  
Junying Zhou ◽  
Xueying Hu ◽  
...  
Keyword(s):  
Clostridium Perfringens ◽  
Pathogenic Bacteria ◽  
Pcr Amplification ◽  
Penicillin G ◽  
Rrna Gene ◽  
Finless Porpoise ◽  
Fecal Samples ◽  
Yangtze Finless Porpoise ◽  
Type C ◽  
Neophocaena Asiaeorientalis Asiaeorientalis

Abstract The aim of this study was to culture pathogenic bacteria from the blowhole, lung, stomach and fecal samples of a neonatal crucially endangered Yangtze finless porpoise (Neophocaena asiaeorientalis asiaeorientalis) that died 27 days after birth. Bacteria were inoculated and representative isolates were identified through 16S rRNA gene sequence analysis. A total of three Clostridium perfringens type C strains from the fecal samples were isolated. Toxin genes, including cpa, cpb and cpb2, were detected by PCR amplification, while the etx, iap and cpe genes were absent. Biofilm formation of the three strains was examined. Only one strain was able to form a biofilm. In addition, isolates showed strong resistance against the antibiotics amikacin (3/3), erythromycin (1/3), gentamicin (3/3), streptomycin (3/3), and trimethoprim (3/3), while sensitivity to ampicillin (3/3), bacitracin (3/3), erythromycin (2/3), penicillin G (3/3), and tetracycline (3/3). The results suggested C. perfringens type C could have contributed to the death of this neonatal porpoise.

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