scholarly journals Molecular prevalence of Cryptosporidium species among household cats and pet shop kittens in Japan

2017 ◽  
Vol 3 (2) ◽  
pp. 205511691773071 ◽  
Author(s):  
Yoichi Ito ◽  
Naoyuki Itoh ◽  
Yuko Iijima ◽  
Yuya Kimura
Keyword(s):  
Clinical Signs ◽  
18S Rrna Gene ◽  
Cryptosporidium Species ◽  
Molecular Characteristics ◽  
Published Data ◽  
Rrna Gene ◽  
Pcr Assay ◽  
Base Pairs ◽  
Faecal Samples ◽  
Significant Difference

Objectives To address the lack of up-to-date published data, the present study evaluates the PCR-based prevalence of Cryptosporidium species infection and molecular characteristics of isolates among household cats and pet shop kittens in Japan. Methods A total of 357 and 329 fresh faecal samples were collected from household cats and pet shop kittens, respectively, with or without clinical signs of infection. A nested PCR assay targeting the 18S rRNA gene was employed for the detection of Cryptosporidium species. After specific DNA fragments (approximately 826 base pairs) were confirmed, the amplicons were sequenced to determine species. Results Seven (2.0%) household cats and one (0.3%) pet shop kitten tested positive for the presence of Cryptosporidium species. In household cats, there was a significant difference in prevalence between cats aged <1 year (4.6%) and those aged ⩾1 year (0.4%). No significantly different prevalence was observed with regard to faecal condition in either household cats or pet shop kittens. A total of eight Cryptosporidium species isolates, seven from household cats and one from a pet shop kitten, were identified as Cryptosporidium felis. Conclusions and relevance The present study demonstrates the risk of zoonotic transmission of Cryptosporidium species from household cats and pet shop kittens to humans is low in Japan.

Prevalence and molecular characterisation of Cryptosporidium spp. in goat kids

2018 ◽  
Author(s):  
A. K. Dixit ◽  
Pooja Dixit ◽  
M.L.V. Rao ◽  
Rohita Gupta ◽  
P. C. Shukla
Keyword(s):  
Cryptosporidium Parvum ◽  
Cryptosporidium Species ◽  
Field Conditions ◽  
Rrna Gene ◽  
Molecular Characterisation ◽  
Ssu Rrna ◽  
Cryptosporidium Spp ◽  
Faecal Samples ◽  
Significant Difference ◽  
Pcr Rflp

Prevalence and molecular characterisation of Cryptosporidium species was done in kids belonging to organised and non-organised goat farms at Jabalpur. The overall prevalence of Cryptosporidium was 14.63%. The prevalence was non-significantly higher in male kids (16.16%) as compared to that of female kids (13.21%). Age wise prevalence was higher in kids up to one month age (16.13%) than that of kids upto 3 months age (13.99%). No significant difference was found in prevalence among different breeds and in kids kept in farm or field conditions. The prevalence was non-significantly higher in non-diarrhoeic kids than diarrhoeic kids. Most of the infections were of one score (76.6%). Molecular characterisation by PCR-RFLP of 18S SSU rRNA gene revealed presence of Cryptosporidium parvum species in positive faecal samples.

scholarly journals Cryptosporidium myocastoris n. sp. (Apicomplexa: Cryptosporidiidae), the Species Adapted to the Nutria (Myocastor coypus)

2021 ◽  
Vol 9 (4) ◽  
pp. 813
Author(s):  
Jana Ježková ◽  
Zlata Limpouchová ◽  
Jitka Prediger ◽  
Nikola Holubová ◽  
Bohumil Sak ◽  
...  
Keyword(s):  
Phylogenetic Analyses ◽  
Clinical Signs ◽  
Prepatent Period ◽  
Cryptosporidium Species ◽  
The Novel ◽  
Myocastor Coypus ◽  
The Czech Republic ◽  
Molecular Analyses ◽  
Cryptosporidium Spp ◽  
Faecal Samples

Cryptosporidium spp., common parasites of vertebrates, remain poorly studied in wildlife. This study describes the novel Cryptosporidium species adapted to nutrias (Myocastor coypus). A total of 150 faecal samples of feral nutria were collected from locations in the Czech Republic and Slovakia and examined for Cryptosporidium spp. oocysts and specific DNA at the SSU, actin, HSP70, and gp60 loci. Molecular analyses revealed the presence of C. parvum (n = 1), C. ubiquitum subtype family XIId (n = 5) and Cryptosporidium myocastoris n. sp. XXIIa (n = 2), and XXIIb (n = 3). Only nutrias positive for C. myocastoris shed microscopically detectable oocysts, which measured 4.8–5.2 × 4.7–5.0 µm, and oocysts were infectious for experimentally infected nutrias with a prepatent period of 5–6 days, although not for mice, gerbils, or chickens. The infection was localised in jejunum and ileum without observable macroscopic changes. The microvilli adjacent to attached stages responded by elongating. Clinical signs were not observed in naturally or experimentally infected nutrias. Phylogenetic analyses at SSU, actin, and HSP70 loci demonstrated that C. myocastoris n. sp. is distinct from other valid Cryptosporidium species.

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 Isolation, screening and identification of Lipase producing fungi from cotton seed soapstock

2020 ◽  
Vol 13 (36) ◽  
pp. 3762-3771
Author(s):  
Gayatriben B Patel ◽  
Keyword(s):  
Aspergillus Niger ◽  
Fusarium Solani ◽  
Cotton Seed ◽  
Quantitative Estimation ◽  
Chemical Constituents ◽  
18S Rrna Gene ◽  
Lipase Production ◽  
Molecular Characteristics ◽  
Rrna Gene ◽  
Plate Assay

Background/Objectives: The present study was focused to exploit the indigenous strains of fungi isolated from cotton seed soapstock for the production of the extracellular lipase through submerged fermentation technique. Methods/ Statistical analysis: Cotton seed soapstock samples used in the study contains gelatinous oil richer chemical constituents. In addition, their enrichment and diluted materials were used for the isolation of lipase producing microorganisms on tributyrin agar plates. All isolates were lipase positive confirmed by a qualitative plate assay. Quantitative estimation of Lipase production activity was measured spectrophotometrically using pnitrophenyl palmitate (p-NPP) as a substrate. In order to exploit the isolated fungal strain for industrial usage, other cellulase and protease enzymes were tested by plate assay. Morphological and molecular characteristics of selected isolates were studied. Findings: From enriched soapstock samples, a total of 49 cultures were isolated among them 19 fungal isolates were screened for lipase, cellulase, and protease activity qualitatively by plate assay. Out of 19, six fungi were selected based on their lipase activity. Highly potent Fusarium solani 7F had the ability to produce 5.95 U/mL/min. crude lipase whereas Aspergillus niger 13F has 4.2 U/mL/min. after 4days incubation at 30oC. Potent fungi culture was identified by morphological, cultural, and molecular characteristics (18s rRNA gene sequence and phylogenetic analysis) revealed them as Penicillium griseofulvum 5F, Aspergillus flavus 6F, Fusarium solani 7F, Aspergillus niger 12F, Aspergillus niger 13F, and Aspergillus terreus 17F. Novelty: Fungi was the first time reported and isolated from cotton seed soapstock materials. In future studies, this enzyme will be used in the degradation of soapstock and also in the production of biosurfactant from soapstock. Keywords: Cotton seed soapstock; Fusarium solani 7F; p-nitrophenyl palmitate; Tributyrin agar plates; degradation

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 Molecular characterization of Eurytrema coelomaticum in cattle from Paraná, Brazil

2014 ◽  
Vol 23 (3) ◽  
pp. 383-386 ◽  
Author(s):  
Gustavo Freire Figueira ◽  
Victor Henrique Silva de Oliveira ◽  
Alessandra Taroda ◽  
Amauri Alcindo Alfieri ◽  
Selwyn Arlington Headley
Keyword(s):  
Molecular Characterization ◽  
Gene Sequence ◽  
Domestic Animals ◽  
18S Rrna Gene ◽  
Rrna Gene ◽  
Rrna Gene Sequence ◽  
Sequence Analyses ◽  
Pcr Assay ◽  
Bovine Pancreas

This study investigated the occurrence of Eurytremaspp. in cattle by analysis of the partial 18S rRNA gene sequence. Trematodes from 44 bovine pancreas were collected and classified based on typical morphological features. PCR assay and sequence analyses of amplified products confirmed that the trematodes classified as Eurytrema coelomaticum were phylogenetically distinct from those identified as E. pancreaticum. The results of this study represent the first molecular characterization of E. coelomaticum within the Americas, and provide an efficient method to differentiate digenean trematodes of domestic animals.

scholarly journals Metagenetic Analysis for Microbial Characterization of Focaccia Doughs Obtained by Using Two Different Starters: Traditional Baker’s Yeast and a Selected Leuconostoc citreum Strain

Foods ◽  
2021 ◽  
Vol 10 (6) ◽  
pp. 1189
Author(s):  
Massimo Ferrara ◽  
Angelo Sisto ◽  
Giuseppina Mulè ◽  
Paola Lavermicocca ◽  
Palmira De Bellis
Keyword(s):  
Hypervariable Region ◽  
18S Rrna Gene ◽  
Biochemical Properties ◽  
Baker’S Yeast ◽  
Rrna Gene ◽  
Baker's Yeast ◽  
Hypervariable Regions ◽  
Leuconostoc Citreum ◽  
Significant Difference ◽  
Microbial Environment

Lactic acid bacteria (LAB) decisively influence the technological, nutritional, organoleptic and preservation properties of bakery products. Therefore, their use has long been considered an excellent strategy to improve the characteristics of those goods. The aim of this study was the evaluation of microbial diversity in different doughs used for the production of a typical Apulian flatbread, named focaccia. Leavening of the analyzed doughs was obtained with baker’s yeast or by applying an innovative “yeast-free” protocol based on a liquid sourdough obtained by using Leuconostoc citreum strain C2.27 as a starter. The microbial populations of the doughs were studied by both a culture-dependent approach and metagenetic analyses. The flours used for dough preparation were also subjected to the same analyses. The metagenetic analyses were performed by sequencing the V5–V6 hypervariable regions of the 16S rRNA gene and the V9 hypervariable region of the 18S rRNA gene. The results indicate that these hypervariable regions were suitable for studying the microbiota of doughs, highlighting a significant difference between the microbial community of focaccia dough with baker’s yeast and that of the dough inoculated with the bacterial starter. In particular, the dough made with baker’s yeast contained a microbiota with a high abundance of Proteobacteria (82% of the bacterial population), known to be negatively correlated with the biochemical properties of the doughs, while the Proteobacteria in dough produced with the L. citreum starter were about 43.5% lower than those in flour and dough prepared using baker’s yeast. Moreover, the results show that the L. citreum C2.27 starter was able to dominate the microbial environment and also reveal the absence of the genus Saccharomyces in the dough used for the production of the “yeast-free” focaccia. This result is particularly important because it highlights the suitability of the starter strain for obtaining an innovative “yeast-free” product.

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 Development and validation of a multiplex real-time qPCR assay using GMP-grade reagents for leprosy diagnosis.

2021 ◽  
Author(s):  
Fernanda Saloum de Neves Manta ◽  
Thiago Jacomasso ◽  
Rita de Cássia Pontello Rampazzo ◽  
Suelen Justo Maria Moreira ◽  
Najua M Zahra ◽  
...  
Keyword(s):  
Internal Control ◽  
Pathogen Detection ◽  
Limit Of Detection ◽  
Mycobacterium Leprae ◽  
18S Rrna Gene ◽  
Cross Reactivity ◽  
Rrna Gene ◽  
Adequate Care ◽  
Independent Panel ◽  
Significant Difference

Leprosy is a chronic dermato-neurological disease caused by Mycobacterium leprae, an obligate intracellular bacterium. Timely detection is a challenge in leprosy diagnosis, relying on clinical examination and trained health professionals. Furthermore, adequate care and transmission control depend on early and reliable pathogen detection. Here, we describe a qPCR test for routine diagnosis of leprosy-suspected patients. The reaction simultaneously amplifies two specific Mycobacterium leprae targets (16S rRNA and RLEP), and the human 18S rRNA gene as internal control. The limit of detection was estimated to be 2.29 copies of M. leprae genome. Analytical specificity was evaluated using a panel of 20 other skin pathogenic microorganisms and Mycobacteria, showing no cross-reactivity. Intra- and inter-operator Cp variation was evaluated using dilution curves of M. leprae DNA or a synthetic gene, and no significant difference was observed between three operators in two different laboratories. The multiplex assay was evaluated using 97 patient samples with clinical and histopathological leprosy confirmation, displaying high diagnostic sensitivity (91%) and specificity (100%). Validation tests in an independent panel of 50 samples confirmed sensitivity and specificity of 97% and 98%, respectively. Importantly, assay performance remained stable for at least five months. Our results show that the newly developed multiplex qPCR effectively and specifically detects M. leprae DNA in skin samples, contributing to an efficient diagnosis that expedites the appropriate treatment.

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