scholarly journals Rapid, Simultaneous Detection ofClostridium sordelliiandClostridium perfringensin Archived Tissues by a Novel PCR-Based Microsphere Assay: Diagnostic Implications for Pregnancy-Associated Toxic Shock Syndrome Cases

2012 ◽  
Vol 2012 ◽  
pp. 1-8 ◽  
Author(s):  
Julu Bhatnagar ◽  
Marlene DeLeon-Carnes ◽  
Kathryn L. Kellar ◽  
Kakali Bandyopadhyay ◽  
Zoi-Anna Antoniadou ◽  
...  
Keyword(s):  
Clostridium Perfringens ◽  
Simultaneous Detection ◽  
Case Fatality ◽  
Cost Effective ◽  
Human Pathogens ◽  
Toxic Shock ◽  
Clostridium Sordellii ◽  
Formalin Fixed Paraffin Embedded ◽  
Ffpe Tissues ◽  
Species Specific

Clostridium sordelliiandClostridium perfringensare infrequent human pathogens; however, the case-fatality rates for the infections are very high, particularly in obstetricC. sordelliiinfections (>90%). Deaths fromClostridium sordelliiandClostridium perfringenstoxic shock (CTS) are sudden, and diagnosis is often challenging. Formalin-fixed, paraffin-embedded (FFPE) tissues usually are the only specimens available for sudden fatal cases, and immunohistochemistry (IHC) for Clostridia is generally performed but it cannot identify species. A clear need exists for a rapid, species-specific diagnostic assay for FFPE tissues. We developed a duplex PCR-based microsphere assay for simultaneous detection ofC. sordelliiandC. perfringensand evaluated DNA extracted from 42Clostridiumisolates and FFPE tissues of 28 patients with toxic shock/endometritis (20 CTS, 8 non-CTS, as confirmed by PCR and sequencing). The microsphere assay correctly identifiedC. sordelliiandC. perfringensin all known isolates and in all CTS patients (10C. sordellii, 8C. perfringens, 2 both) and showed 100% concordance with PCR and sequencing results. The microsphere assay is a rapid, specific, and cost-effective method for the diagnosis of CTS and offers the advantage of simultaneous testing forC. sordelliiandC. perfringensin FFPE tissues using a limited amount of DNA.

scholarly journals Clostridium perfringens–Associated Necrotic Enteritis-Like Disease in Coconut Lorikeets (Trichoglossus haematodus)

2020 ◽  
pp. 030098582097178
Author(s):  
Llorenç Grau-Roma ◽  
Mauricio Navarro ◽  
Sohvi Blatter ◽  
Christian Wenker ◽  
Sonja Kittl ◽  
...  
Keyword(s):  
Clostridium Perfringens ◽  
Toxin Gene ◽  
Necrotic Enteritis ◽  
Gene Encoding ◽  
Formalin Fixed Paraffin ◽  
Formalin Fixed Paraffin Embedded ◽  
Ffpe Tissues ◽  
Polymerase Chain ◽  
Beta Toxin ◽  
Formalin Fixed

Several outbreaks of necrotic enteritis-like disease in lorikeets, from which Clostridium perfringens was consistently isolated, are described. All lorikeets had acute, segmental, or multifocal fibrinonecrotizing inflammatory lesions in the small and/or the large intestine, with intralesional gram-positive rods. The gene encoding C. perfringens alpha toxin was detected by PCR (polymerase chain reaction) on formalin-fixed, paraffin-embedded (FFPE) tissues in 20 out of 24 affected lorikeets (83%), but it was not amplified from samples of any of 10 control lorikeets ( P < .0001). The second most prevalent C. perfringens toxin gene detected was the beta toxin gene, which was found in FFPE from 7 out of 24 affected lorikeets (29%). The other toxin genes were detected inconsistently and in a relatively low number of samples. These cases seem to be associated with C. perfringens, although the specific type involved could not be determined.

scholarly journals Development of a 16S metabarcoding assay for the environmental DNA (eDNA) detection of aquatic reptiles across northern Australia

2020 ◽  
Author(s):  
Katrina West ◽  
Matthew Heydenrych ◽  
Rose Lines ◽  
Tony Tucker ◽  
Sabrina Fossette ◽  
...  
Keyword(s):  
Simultaneous Detection ◽  
Environmental Dna ◽  
Cost Effective ◽  
Distribution Data ◽  
Freshwater Turtles ◽  
Northern Australia ◽  
Non Invasive ◽  
Sea Snakes ◽  
Reptile Species ◽  
Species Specific

AbstractA severe lack of distribution data for aquatic reptiles in northern Australia leaves many taxa vulnerable to extirpation and extinction. Environmental DNA (eDNA) technologies offer sensitive and non-invasive genetic alternatives to trapping and visual surveys and are increasingly employed for the detection of aquatic and semi-aquatic reptiles. However, at present, these studies have largely applied species-specific primers which do not provide a cost-effective avenue for the simultaneous detection of multiple reptilian taxa. Here, we present a 16S rRNA metabarcoding assay for the broad detection of aquatic and semi-aquatic reptile species. This assay is tested on water samples collected at multiple sampling sites at two tropical locations: 12 marine/estuarine sites in Roebuck Bay, Western Australia, and 4 estuarine sites in Cooktown, Queensland, Australia. A total of nine reptile taxa were detected from 10 of the 16 sampled sites, including marine and freshwater turtles, aquatic and semi-aquatic/terrestrial snakes, and terrestrial skinks. However, inconsistencies in the detection of previously observed aquatic reptiles at our sampled sites, such as saltwater crocodile and sea snakes, indicates that further research is required to assess the reliability, strengths and limitations of eDNA methods for aquatic reptile detection before it can be integrated as a broad-scale bioassessment tool.

Toxic Shock Associated With Clostridium sordellii and Clostridium perfringens After Medical and Spontaneous Abortion

2007 ◽  
Vol 110 (5) ◽  
pp. 1027-1033 ◽  
Author(s):  
Adam L. Cohen ◽  
Julu Bhatnagar ◽  
Sarah Reagan ◽  
Suzanne B. Zane ◽  
Marisa A. DʼAngeli ◽  
...  
Keyword(s):  
Clostridium Perfringens ◽  
Spontaneous Abortion ◽  
Toxic Shock ◽  
Clostridium Sordellii

Species Specific Immunoassays to Measure Blood Platelet and Coagulation Activation in the Rat

1996 ◽  
Vol 76 (06) ◽  
pp. 1090-1095 ◽  
Author(s):  
C Ravanat ◽  
M Freund ◽  
S Schuhler ◽  
P Grunert ◽  
L Meyer ◽  
...  
Keyword(s):  
Antithrombin Iii ◽  
Plasma Concentrations ◽  
Simultaneous Detection ◽  
Blood Platelet ◽  
Coagulation Activation ◽  
Platelet Factor ◽  
Prethrombotic State ◽  
Low Levels ◽  
Species Specific ◽  
Higher Sensitivity

SummaryThe purpose of this study was to develop specific and sensitive immunoassays to detect early indices of hypercoagulability in the rat. Rat platelet factor 4 (rPF4) and rat fibrinopeptide A (rFPA) assays, tools for the detection of activation of platelets and coagulation respectively, were designed using antibodies raised against purified rPF4 and against synthetic rFPA. The relevance of these new assays and of the commercially available ELISA kit for thrombin-antithrombin III (TAT) complexes was demonstrated in a rat model of a prethrombotic state induced by intravenous infusion of varying doses of thromboplastin (90 to 2400 μl/kg/h). In this model, the immunoassays allowed simultaneous detection of low levels of rFPA and rPF4 which were correlated with fibrinogen and platelet consumption and TAT generation and further proved to be of higher sensitivity than the classical methods of platelet count or measurement of fibrinogen levels. Plasma concentrations of rFPA, rPF4 and TAT were dependent on infusion time and thromboplastin dose, while hirudin (1 mg/kg) prevented their appearance. Thus the new specific immunoassays for rPF4 and rFPA and the commercial human TAT assay represent useful tools for pathophysiological studies or the screening of antithrombotic drugs in rats.

scholarly journals Conversion of sequence-characterized amplified region (SCAR) bands into high-throughput DNA markers based on RAPD technique for detection of the stem nematode Ditylenchus dipsaci in crucial plant hosts

2008 ◽  
Vol 53 (No. 3) ◽  
pp. 97-104 ◽  
Author(s):  
M. Zouhar ◽  
M. Marek ◽  
O. Douda ◽  
J. Mazáková ◽  
P. Ryšánek
Keyword(s):  
Cost Effective ◽  
Healthy Plant ◽  
Host Preferences ◽  
Sequence Characterized Amplified Region ◽  
Detection And Identification ◽  
Plant Hosts ◽  
Cost Effective Technique ◽  
Ditylenchus Dipsaci ◽  
Species Specific ◽  
Template Dna

<i>Ditylenchus dipsaci</i>, the stem nematode, is a migratory endoparasite of over 500 species of angiosperms. The main method of <i>D. dipsaci</i> control is crop rotation, but the presence of morphologically indistinguishable host races with different host preferences makes rotation generally ineffective. Therefore, a sensitive, rapid, reliable, as well as cost effective technique is needed for identification of <i>D. dipsaci</i> in biological samples. This study describes the development of species-specific pairs of PCR oligonucleotides for detection and identification of the <i>D. dipsaci</i> stem nematode in various plant hosts. Designed DIT-2 primer pair specifically amplified a fragment of 325 bp, while DIT-5 primer pair always produced a fragment of 245 bp in all <i>D. dipsaci</i> isolates. Two developed SCAR primer pairs were further tested using template DNA extracted from a collection of twelve healthy plant hosts; no amplification was however observed. The developed PCR protocol has proved to be quite sensitive and able to specifically detect <i>D. dipsaci</i> in artificially infested plant tissues.

scholarly journals A New Multiplex Real-Time RT-PCR for Simultaneous Detection and Differentiation of Avian Bornaviruses

Viruses ◽  
2021 ◽  
Vol 13 (7) ◽  
pp. 1358
Author(s):  
Brigitte Sigrist ◽  
Jessica Geers ◽  
Sarah Albini ◽  
Dennis Rubbenstroth ◽  
Nina Wolfrum
Keyword(s):  
Real Time ◽  
Simultaneous Detection ◽  
Epidemiological Studies ◽  
Virus Species ◽  
Rt Pcr ◽  
P Gene ◽  
Field Samples ◽  
Causative Agents ◽  
Species Specific ◽  
Proventricular Dilatation Disease

Avian bornaviruses were first described in 2008 as the causative agents of proventricular dilatation disease (PDD) in parrots and their relatives (Psittaciformes). To date, 15 genetically highly diverse avian bornaviruses covering at least five viral species have been discovered in different bird orders. Currently, the primary diagnostic tool is the detection of viral RNA by conventional or real-time RT-PCR (rRT-PCR). One of the drawbacks of this is the usage of either specific assays, allowing the detection of one particular virus, or of assays with a broad detection spectrum, which, however, do not allow for the simultaneous specification of the detected virus. To facilitate the simultaneous detection and specification of avian bornaviruses, a multiplex real-time RT-PCR assay was developed. Whole-genome sequences of various bornaviruses were aligned. Primers were designed to recognize conserved regions within the overlapping X/P gene and probes were selected to detect virus species-specific regions within the target region. The optimization of the assay resulted in the sensitive and specific detection of bornaviruses of Psittaciformes, Passeriformes, and aquatic birds. Finally, the new rRT-PCR was successfully employed to detect avian bornaviruses in field samples from various avian species. This assay will serve as powerful tool in epidemiological studies and will improve avian bornavirus detection.

scholarly journals Characterization of epididymal and testicular histologic lesions and use of immunohistochemistry and PCR on formalin-fixed tissues to detect Brucella canis in male dogs

2021 ◽  
pp. 104063872098688
Author(s):  
Andrea M. Camargo-Castañeda ◽  
Lauren W. Stranahan ◽  
John F. Edwards ◽  
Daniel G. Garcia-Gonzalez ◽  
Leonardo Roa ◽  
...  
Keyword(s):  
Accurate Diagnosis ◽  
Testicular Atrophy ◽  
Detection Techniques ◽  
Formalin Fixed Paraffin ◽  
Brucella Canis ◽  
Formalin Fixed Paraffin Embedded ◽  
Variable Sensitivity ◽  
Ffpe Tissues ◽  
Formalin Fixed

In male dogs, Brucella canis frequently causes epididymitis, ultimately resulting in testicular atrophy and infertility. Although B. canis predominantly affects the epididymis, the misleading term “orchitis” is still commonly used by clinicians. Of additional concern, diagnosis in dogs remains challenging because of variable sensitivity and specificity of serologic assays and fluctuations in bacteremia levels in infected dogs, reducing the sensitivity of blood culture. We describe here the histologic lesions in the scrotal contents of 8 dogs suspected of being infected with B. canis and clinically diagnosed with orchitis. We explored the possibility of using immunohistochemistry (IHC) and real-time PCR (rtPCR) in formalin-fixed, paraffin-embedded (FFPE) tissues to detect the presence of B. canis. Epididymitis of variable chronicity was identified in all 8 dogs, with only 3 also exhibiting orchitis. Using rtPCR, the presence of B. canis was identified in 4 of 8 dogs, with 3 of these 4 dogs also positive by IHC. These results suggest that rtPCR and IHC are promising techniques that can be used in FFPE tissues to detect B. canis when other detection techniques are unavailable. Additionally, accurate recognition of epididymitis rather than orchitis in suspect cases could aid in accurate diagnosis.

Duplex DNA barcoding allows accurate species determination of morphologically similar limpets (Patella spp.) from non-destructive sampling

2016 ◽  
Vol 97 (7) ◽  
pp. 1479-1482 ◽  
Author(s):  
Thomas J. Ashton ◽  
Meriem Kayoueche-Reeve ◽  
Andrew J. Blight ◽  
Jon Moore ◽  
David M. Paterson
Keyword(s):  
Dna Barcoding ◽  
Species Abundance ◽  
Microscopic Analysis ◽  
Cost Effective ◽  
Field Studies ◽  
Similar Species ◽  
Species Determination ◽  
Destructive Sampling ◽  
Species Specific ◽  
Non Destructive

Accurate discrimination of two morphologically similar species of Patella limpets has been facilitated by using qPCR amplification of species-specific mitochondrial genomic regions. Cost-effective and non-destructive sampling is achieved using a mucus swab and simple sample lysis and dilution to create a PCR template. Results show 100% concurrence with dissection and microscopic analysis, and the technique has been employed successfully in field studies. The use of highly sensitive DNA barcoding techniques such as this hold great potential for improving previously challenging field assessments of species abundance.

scholarly journals The Potential Role of Nonhuman Primate Models to Better Comprehend Early Life Immunity and Maternal Antibody Transfer

Vaccines ◽  
2021 ◽  
Vol 9 (4) ◽  
pp. 306
Author(s):  
Julie Sartoretti ◽  
Christiane S. Eberhardt
Keyword(s):  
Nonhuman Primate ◽  
Early Life ◽  
Cost Effective ◽  
Maternal Antibody ◽  
Maternal Antibodies ◽  
Immunological Methods ◽  
Human Pathogens ◽  
Childhood Immunization ◽  
Primate Model ◽  
The Impact

Early life immunity is a complex field of research and there are still gaps in knowledge regarding the detailed mechanism of maternal antibody transfer, the impact of maternal antibodies on infant vaccine responses and the ontogeny of human early life immunity. A comprehensive understanding is necessary to identify requirements for early life vaccines and to improve early childhood immunization. New immunological methods have facilitated performing research in the youngest, however, some questions can only be addressed in animal models. To date, mostly murine models are used to study neonatal and infant immunity since they are well-described, easy to use and cost effective. Given their limitations especially in the transfer biology of maternal antibodies and the lack of infectivity of numerous human pathogens, this opinion piece discusses the potential and prerequisites of the nonhuman primate model in studying early life immunity and maternal antibody transfer.

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