Investigations on First Confirmed Outbreak of Ovine Theileriosis (Theileria luwenshuni) from Maharashtra State, India

2020 ◽  
Author(s):  
V.S. Dhaygude ◽  
K. Kundu ◽  
B.P. Kamdi ◽  
U.R. Bagal ◽  
S.B. Bhosale ◽  
...  
Keyword(s):  
18S Rrna ◽  
Clinical Signs ◽  
Specific Treatment ◽  
Small Ruminants ◽  
18S Rrna Gene ◽  
Pcr Detection ◽  
Nucleotide Sequencing ◽  
Rrna Gene ◽  
Blood Samples ◽  
Theileria Luwenshuni

Background: Clinical theileriosis of small ruminants is tick-borne disease caused by Theileria lestoquardi, Theileria uilenbergi and Theileria luwenshuni. Theileria annulata, the causative agent of bovine tropical theileriosis in cattle, can also infect sheep but does not cause any significant illness. It is one of the economically important diseases. There are no reports of ovine clinical theileriosis from Maharashtra state and there is paucity of information on its epidemiology. This paper reports first confirmed outbreak of ovine theileriosis based on clinical signs, microscopic examination, PCR and sequencing in the Maharashtra State of India. Methods: Whole blood samples from 22 ailing sheep were collected and subjected to hematological examination. Blood smears stained with Leishman’s stain were examined under 100X objective of the microscope. The blood samples from sheep found positive by microscopic method were subjected to PCR detection of 18S rRNA gene of hemoprotozoa and then for nucleotide sequencing and sequence analysis.Conclusion: Samples from 14 out of 22 sheep were found positive for piroplasms of Theileria spp by light microscopy. All positive samples were further confirmed by PCR detection of 18S rRNA gene of hemoprotozoa. PCR amplification yielded expected product of 1750 bp for all samples. BLAST and phylogenetic analysis of one sample revealed high sequence homology with T. luwenshuni reported from India and other countries. Characteristic clinical signs like fever, progressive anaemia, laboured breathing, lymphadenopathy, debility and non-responsiveness to antibiotic therapy were recorded. The animals responded to specific treatment against theileriosis. It is the first ever confirmed report of ovine theileriosis in Maharashtra state of India and hence reported.

scholarly journals Analytical validation of a real-time hydrolysis probe PCR assay for quantifying Plasmodium falciparum parasites in experimentally infected human adults

2021 ◽  
Vol 20 (1) ◽  
Author(s):  
Claire Y. T. Wang ◽  
Emma L. Ballard ◽  
Zuleima Pava ◽  
Louise Marquart ◽  
Jane Gaydon ◽  
...  
Keyword(s):  
Clinical Trials ◽  
Plasmodium Falciparum ◽  
18S Rrna ◽  
Drug Efficacy ◽  
18S Rrna Gene ◽  
Molecular Techniques ◽  
Qpcr Assay ◽  
Rrna Gene ◽  
Analytical Sensitivity ◽  
Blood Samples

Abstract Background Volunteer infection studies have become a standard model for evaluating drug efficacy against Plasmodium infections. Molecular techniques such as qPCR are used in these studies due to their ability to provide robust and accurate estimates of parasitaemia at increased sensitivity compared to microscopy. The validity and reliability of assays need to be ensured when used to evaluate the efficacy of candidate drugs in clinical trials. Methods A previously described 18S rRNA gene qPCR assay for quantifying Plasmodium falciparum in blood samples was evaluated. Assay performance characteristics including analytical sensitivity, reportable range, precision, accuracy and specificity were assessed using experimental data and data compiled from phase 1 volunteer infection studies conducted between 2013 and 2019. Guidelines for validation of laboratory-developed molecular assays were followed. Results The reportable range was 1.50 to 6.50 log10 parasites/mL with a limit of detection of 2.045 log10 parasites/mL of whole blood based on a parasite diluted standard series over this range. The assay was highly reproducible with minimal intra-assay (SD = 0.456 quantification cycle (Cq) units [0.137 log10 parasites/mL] over 21 replicates) and inter-assay (SD = 0.604 Cq units [0.182 log10 parasites/mL] over 786 qPCR runs) variability. Through an external quality assurance program, the QIMR assay was shown to generate accurate results (quantitative bias + 0.019 log10 parasites/mL against nominal values). Specificity was 100% after assessing 164 parasite-free human blood samples. Conclusions The 18S rRNA gene qPCR assay is specific and highly reproducible and can provide reliable and accurate parasite quantification. The assay is considered fit for use in evaluating drug efficacy in malaria clinical trials.

scholarly journals Emergence of Babesia Conradae Infection in Coyote-hunting Greyhounds in Oklahoma, USA

2021 ◽  
Author(s):  
Erin M Stayton ◽  
Megan Lineberry ◽  
Jennifer Thomas ◽  
Tina Bass ◽  
Kelly Allen ◽  
...  
Keyword(s):  
18S Rrna ◽  
Clinical Signs ◽  
18S Rrna Gene ◽  
Post Treatment ◽  
Rrna Gene ◽  
Babesia Gibsoni ◽  
Wide Range ◽  
Pcr Products ◽  
Life Threatening ◽  
Tick Vectors

Abstract Background: Babesia species are intraerythrocytic Apicomplexan parasites that infect a wide range of vertebrate hosts. These pathogens are typically transmitted either by tick vectors or by direct blood-to-blood contact, and may cause life-threatening clinical disease such as thrombocytopenia, hemolytic anemia, and acute renal failure in canine hosts. While Babesia vogeli and Babesia gibsoni infections have both been reported in Oklahoma, reports of B. conradae infections have been limited to California. Methods: Whole blood samples were collected in EDTA tubes from all dogs in four separate kennels in Oklahoma. DNA was extracted from each blood sample and a nested PCR was performed using general Apicomplexan primers for the partial 18S rRNA gene. PCR products were electrophoresed in agarose matrix and appropriately sized amplicons were sequenced. Sequences were compared to reference 18S rRNA sequences available in GenBank, and samples with >98% homology to B. conradae (GenBank MK256976) were considered positive. B. conradae positive dogs were then treated with atovaquone (13.5 mg/kg TID) and azithromycin (10 mg/kg SID) for 10 days and retested at 30 and 60 days post treatment by PCR. Results: Fifteen of 40 dogs tested positive for B. conradae with 98–100% sequence homology to B. conradae from California. All positive cases were coyote-hunting Greyhounds. Treatment of clinically ill dogs with atovaquone and azithromycin resulted in complete clinical recovery in clinically ill dogs and all treated dogs had negative follow-up PCR at 30 and 60 days post treatment. Conclusions: Collectively, this study (i) documents the occurrence of B. conradae in Oklahoma, (ii) highlights this pathogen as a differential to be considered when clinical signs are present, and (iii) supports the use of atovaquone and azithromycin as effective treatment in these cases.

scholarly journals Molecular differentiation of Thysaniezia (Helictometra) giardi and Moniezia species based on 18s rRNA gene in small ruminants

2021 ◽  
Vol 35 (1) ◽  
pp. 105-108
Author(s):  
Monyer A. Alfatlawi ◽  
Yahia Kh. Ismail ◽  
Mansour J. Ali ◽  
Azhar C. Karawan ◽  
Israa N. Ibadi
Keyword(s):  
18S Rrna ◽  
Small Ruminants ◽  
18S Rrna Gene ◽  
Rrna Gene ◽  
Molecular Differentiation

scholarly journals Survey and Molecular Study of Babesia gibsoni in Dogs of Baghdad Province, Iraq

2020 ◽  
Vol 44 ((E0)) ◽  
pp. 34-41
Author(s):  
Naseir M. Badawi ◽  
Afaf A. Yousif
Keyword(s):  
Infection Rate ◽  
18S Rrna ◽  
Sequence Data ◽  
Phylogenetic Analyses ◽  
Clinical Signs ◽  
Molecular Study ◽  
18S Rrna Gene ◽  
Universal Primers ◽  
Rrna Gene ◽  
Babesia Gibsoni

This study aimed to detect Babesia gibsoni (B. gibsoni) in dogs of different ages, sex and breeds in Baghdad province by microscopic and molecular investigations using polymerase chain reaction (PCR), sequencing, and phylogenetic analyses. The present study was investigated B. gibsoni in 310 blood samples of dogs for the period December 2018 to September 2019 in Baghdad province, Iraq. The molecular study was carried out by using universal primers of Babesia spp. (PIRO-A and PIRO-B) and specific primers of B. gibsoni (BAGIF and BAGIR) products size of 410 bp and 488 bp fragments of 18S rRNA gene respectively. The clinical signs revealed higher percentage and specific clinical signs of B. gibsoni as depression, anorexia, fever, pale mucus membrane, and ticks infestation, however icterus, and dead were low in which only occurred in two dogs out of infected dogs. The PCR assay and microscopic diagnosis revealed the infection rate of B. gibsoni 9 out of 310 (2.9%) in dogs. The sequence data analyses of nine DNA products were 98-100% similar to sequences of 18S rRNA gene of B. gibsoni data available in Gene bank. According to breed, age, and sex, the results revealed a significantly high-risk factor of infection in Husky dogs; B. gibsoni detected in females which was increased non-significantly than males; while the highest occurrence of disease was in young dogs aged three years or less in addition to the above, the infection rate of B. gibsoni was high in the spring season. In conclusion, this study was considered the first molecular record of B. gibsoni in Baghdad, Iraq documented no differences in diagnosis by blood smear and conventional PCR to amplify of 18S rRNA gene and partial sequencing of B. gibsoni with low-cost method and easily done.

scholarly journals Molecular Detection of Tick-Borne Agents in Cats from Southeastern and Northern Brazil

Pathogens ◽  
2022 ◽  
Vol 11 (1) ◽  
pp. 106
Author(s):  
Marcos Rogério André ◽  
Ana Cláudia Calchi ◽  
Maria Eduarda Chiaradia Furquim ◽  
Isabela de Andrade ◽  
Paulo Vitor Cadina Arantes ◽  
...  
Keyword(s):  
18S Rrna ◽  
Clinical Signs ◽  
18S Rrna Gene ◽  
Molecular Techniques ◽  
Rrna Gene ◽  
Molecular Identity ◽  
Northern Brazil ◽  
Rrna Gene Sequencing ◽  
First Time ◽  
Theileria Spp

Even though the epidemiology of tick-borne agents (TBA) in dogs has been extensively investigated around the world, the occurrence, vectors involved, and molecular identity of these agents in cats remains elusive in many regions. Among TBA, Ehrlichia, Anaplasma, Babesia, Cytauxzoon, and Hepatozoon are responsible for diseases with non-specific clinical signs in cats, making essential the use of molecular techniques for accurate diagnosis and proper treatment. The present work aimed to investigate the occurrence and molecular identity of tick-borne agents (Ehrlichia, Anaplasma, Babesia/Theileria, Cytauxzoon, and Hepatozoon) in cats from southeastern (states of São Paulo (SP) and Minas Gerais (MG)) and northern (state of Rondônia (RO)) Brazil. For this purpose, 390 blood samples were collected from domiciled cats in MG (n = 155), SP (n = 151), and RO(n = 84) states, submitted to DNA extraction and PCR assays for Ehrlichia spp. (dsb gene), Anaplasma spp. (rrs gene), piroplasmids (18S rRNA gene), and Hepatozoon spp. (18S rRNA gene), sequencing, and phylogenetic inferences. The overall positivity for Anaplasma spp., Ehrlichia spp., Babesia/Theileria spp., Cytauxzoon spp., and Hepatozoon spp. were 7.4% (12.3% (MG) and 6.6% (SP)), 2% (4.5% (MG) and 0.6% (SP)), 0.7% (0.6% (MG), 0.6% (SP) and 1.2% (RO)), 27.2% (41.9% (MG), 24.5% (SP) and 4.8% (RO), and 0%, respectively. The phylogenetic analysis grouped the obtained sequences with ‘Candidatus Anaplasma amazonensis’, A. platys, B. vogeli, and Cytauxzoon sp. previously detected in wild felids from Brazil. qPCR specific for E. canis based on the dsb gene confirmed the molecular identity of the detected ehrlichial agent. The present study expanded the list and geographical distribution of hemoparasites in cats. ‘Candidatus Anaplasma amazonensis’, recently detected in sloths from northern Brazil, was described for the first time in cats. This is the first report of piroplasmids infecting cats in northern Brazil. Coinfection by Cytauxzoon and other TBA (Ehrlichia, Anaplasma, and B. vogeli) reported in the present study raises the need for veterinary practitioners’ awareness of cats parasitized by multiple TBA.

scholarly journals Molecular Identification and Genotyping of Babesia canis in Dogs from Mesh-kin Shahr County, Northwestern Iran

2021 ◽  
Author(s):  
Majid Khanmohammadi ◽  
Reza Z Olfaghari-Emameh ◽  
Mehdi Arshadi ◽  
Elham Razmjou ◽  
Poorya Karimi
Keyword(s):  
18S Rrna ◽  
Gene Sequencing ◽  
18S Rrna Gene ◽  
Rrna Gene ◽  
Babesia Species ◽  
Canine Babesiosis ◽  
Sequence Alignments ◽  
Blood Samples ◽  
Standard Methods ◽  
Northwestern Iran

Background: Canine babesiosis is one of the mainly worldwide-distributed tick-borne haemoprotozoan parasitic dis-eases in dogs. Methods: A total of 43 blood samples were randomly collected from naturally infected dogs in seven villages from different geographical areas of Meshkin Shahr, Ardabil Province, Iran. The presence of Babesia species detected with standard methods including parasitological and gene sequencing techniques targeting the 18S rRNA gene. Results: Our results revealed that four dogs 9.3% (4/43) including one female and three male dogs were infected with Babesia. All four Babesia-infected dogs were confirmed B. canis by the molecular-based method. Sequence alignments comparison of the B. canis genotypes A and B, it was revealed that all B. canis isolates belonged to genotype B. Conclusion: This study provides essential data for subsequently define the critical importance of the molecular studies in management and prevention of the canine babesiosis in Iran.  

scholarly journals First molecular isolation of Mycoplasma ovis from small ruminants in North Africa

2015 ◽  
Vol 82 (1) ◽  
Author(s):  
Mohamed R. Rjeibi ◽  
Mohamed A. Darghouth ◽  
Houda Omri ◽  
Khemaïs Souidi ◽  
Mourad Rekik ◽  
...  
Keyword(s):  
North Africa ◽  
Small Ruminant ◽  
Acute Infection ◽  
Clinical Signs ◽  
Small Ruminants ◽  
Molecular Study ◽  
Rrna Gene ◽  
North African ◽  
Blood Samples ◽  
Central Tunisia

Eperythrozoonosis is a small ruminant disease caused by the bacterium Mycoplasma ovis (formerly known as Eperythrozoon ovis). Whilst acute infection in sheep may result in an anaemia and ill thrift syndrome, most animals do not develop clinical signs. Molecular methods were used to compare and evaluate the prevalence of infection with M. ovis in sheep and goats in Tunisia. A total of 739 whole blood samples from 573 sheep and 166 goats were tested for the M. ovis 16S rRNA gene using PCR. The overall prevalence was 6.28% ± 0.019 (36/573). Only sheep were infected with M. ovis (p < 0.001), and the prevalence was significantly higher in central Tunisia (29.2%) compared with other regions (p < 0.05). The prevalence revealed significant differences according to breed and bioclimatic zones (p < 0.001). Furthermore, the prevalence in young sheep (35/330; 10.6%) was higher than in adults (1/243; 0.41%) (p < 0.001). Only sheep of the Barbarine breed were infected, with a prevalence of 11.8% (p < 0.001). This is the first molecular study and genetic characterisation of M. ovis in North African sheep breeds.

scholarly journals 16S rRNA gene and 18S rRNA gene diversity in microbial mat communities in meltwater ponds on the McMurdo Ice Shelf, Antarctica

Polar Biology ◽  
2021 ◽  
Author(s):  
Eleanor E. Jackson ◽  
Ian Hawes ◽  
Anne D. Jungblut
Keyword(s):  
16S Rrna ◽  
18S Rrna ◽  
High Throughput Sequencing ◽  
Microbial Mats ◽  
Gene Diversity ◽  
Salinity Gradient ◽  
18S Rrna Gene ◽  
Rrna Gene ◽  
Ice Shelf ◽  
The Impact

AbstractThe undulating ice of the McMurdo Ice Shelf, Southern Victoria Land, supports one of the largest networks of ice-based, multiyear meltwater pond habitats in Antarctica, where microbial mats are abundant and contribute most of the biomass and biodiversity. We used 16S rRNA and 18S rRNA gene high-throughput sequencing to compare variance of the community structure in microbial mats within and between ponds with different salinities and pH. Proteobacteria and Cyanobacteria were the most abundant phyla, and composition at OTU level was highly specific for the meltwater ponds with strong community sorting along the salinity gradient. Our study provides the first detailed evaluation of eukaryote communities for the McMurdo Ice Shelf using the 18S rRNA gene. They were dominated by Ochrophyta, Chlorophyta and Ciliophora, consistent with previous microscopic analyses, but many OTUs belonging to less well-described heterotrophic protists from Antarctic ice shelves were also identified including Amoebozoa, Rhizaria and Labyrinthulea. Comparison of 16S and 18S rRNA gene communities showed that the Eukaryotes had lower richness and greater similarity between ponds in comparison with Bacteria and Archaea communities on the McMurdo Ice shelf. While there was a weak correlation between community dissimilarity and geographic distance, the congruity of microbial assemblages within ponds, especially for Bacteria and Archaea, implies strong habitat filtering in ice shelf meltwater pond ecosystems, especially due to salinity. These findings help to understand processes that are important in sustaining biodiversity and the impact of climate change on ice-based aquatic habitats in Antarctica.

scholarly journals 18S rRNA gene sequences of leptocephalus gut contents, particulate organic matter, and biological oceanographic conditions in the western North Pacific

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Tsuyoshi Watanabe ◽  
Satoshi Nagai ◽  
Yoko Kawakami ◽  
Taiga Asakura ◽  
Jun Kikuchi ◽  
...  
Keyword(s):  
Organic Matter ◽  
Particulate Organic Matter ◽  
North Pacific ◽  
Western North Pacific ◽  
18S Rrna ◽  
18S Rrna Gene ◽  
Gut Contents ◽  
Rrna Gene ◽  
Marine Snow ◽  
Chlorophyll Maximum

AbstractEel larvae apparently feed on marine snow, but many aspects of their feeding ecology remain unknown. The eukaryotic 18S rRNA gene sequence compositions in the gut contents of four taxa of anguilliform eel larvae were compared with the sequence compositions of vertically sampled seawater particulate organic matter (POM) in the oligotrophic western North Pacific Ocean. Both gut contents and POM were mainly composed of dinoflagellates as well as other phytoplankton (cryptophytes and diatoms) and zooplankton (ciliophoran and copepod) sequences. Gut contents also contained cryptophyte and ciliophoran genera and a few other taxa. Dinoflagellates (family Gymnodiniaceae) may be an important food source and these phytoplankton were predominant in gut contents and POM as evidenced by DNA analysis and phytoplankton cell counting. The compositions of the gut contents were not specific to the species of eel larvae or the different sampling areas, and they were most similar to POM at the chlorophyll maximum in the upper part of the thermocline (mean depth: 112 m). Our results are consistent with eel larvae feeding on marine snow at a low trophic level, and feeding may frequently occur in the chlorophyll maximum in the western North Pacific.

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