scholarly journals Seoul Virus Infection and Spread in United States Home-Based Ratteries: Rat and Human Testing Results From a Multistate Outbreak Investigation

2020 ◽  
Vol 222 (8) ◽  
pp. 1311-1319
Author(s):  
Barbara Knust ◽  
Shelley Brown ◽  
Annabelle de St. Maurice ◽  
Shannon Whitmer ◽  
Sarah E Koske ◽  
...  
Keyword(s):  
Diagnostic Testing ◽  
Pcr Primers ◽  
Outbreak Investigation ◽  
Rt Pcr ◽  
Recent Infection ◽  
Seoul Virus ◽  
Home Based ◽  
Health Community ◽  
Polymerase Chain ◽  
Ig G

Abstract Background During 2017, a multistate outbreak investigation occurred after the confirmation of Seoul virus (SEOV) infections in people and pet rats. A total of 147 humans and 897 rats were tested. Methods In addition to immunoglobulin (Ig)G and IgM serology and traditional reverse-transcription polymerase chain reaction (RT-PCR), novel quantitative RT-PCR primers/probe were developed, and whole genome sequencing was performed. Results Seventeen people had SEOV IgM, indicating recent infection; 7 reported symptoms and 3 were hospitalized. All patients recovered. Thirty-one facilities in 11 US states had SEOV infection, and among those with ≥10 rats tested, rat IgG prevalence ranged 2%–70% and SEOV RT-PCR positivity ranged 0%–70%. Human laboratory-confirmed cases were significantly associated with rat IgG positivity and RT-PCR positivity (P = .03 and P = .006, respectively). Genomic sequencing identified >99.5% homology between SEOV sequences in this outbreak, and these were >99% identical to SEOV associated with previous pet rat infections in England, the Netherlands, and France. Frequent trade of rats between home-based ratteries contributed to transmission of SEOV between facilities. Conclusions Pet rat owners, breeders, and the healthcare and public health community should be aware and take steps to prevent SEOV transmission in pet rats and to humans. Biosecurity measures and diagnostic testing can prevent further infections.

scholarly journals Utility of forensic detection of rabies virus in decomposed exhumed dog carcasses

2015 ◽  
Vol 86 (1) ◽  
Author(s):  
Wanda Markotter ◽  
Jessica Coertse ◽  
Kevin Le Roux ◽  
Joey Peens ◽  
Jacqueline Weyer ◽  
...  
Keyword(s):  
Rabies Virus ◽  
Gold Standard ◽  
Diagnostic Method ◽  
Diagnostic Testing ◽  
Domestic Dogs ◽  
Rt Pcr ◽  
Chain Reaction ◽  
Brain Material ◽  
Polymerase Chain ◽  
Genomic Material

This report describes four suspected rabies cases in domestic dogs that were involved inhuman exposures. In all these cases, the animals were buried for substantial times beforerabies testing was performed. Animal rabies is endemic in South Africa and domestic dogsare the main vector for transmission to humans. Diagnosis of rabies in humans is complicated,and diagnosis in the animal vector can provide circumstantial evidence to support clinicaldiagnosis of rabies in humans. The gold standard diagnostic method, fluorescent antibodytest (FAT), only delivers reliable results when performed on fresh brain material and thereforedecomposed samples are rarely submitted for diagnostic testing. Severely decomposed brainmaterial was tested for the presence of rabies virus genomic material using a quantitativereal-time reverse transcription polymerase chain reaction (q-real-time RT-PCR) assaywhen conventional molecular methods were unsuccessful. This may be a useful tool in theinvestigation of cases where the opportunity to sample the suspected animals post mortem wasforfeited and which would not be possible with conventional testing methodologies becauseof the decomposition of the material.

RT-PCR microlocalization of mRNAs for calbindin D28k and vitamin D receptor in the murine nephron

1996 ◽  
Vol 270 (4) ◽  
pp. F677-F681 ◽  
Author(s):  
L. Liu ◽  
A. Khastgir ◽  
J. M. McCauley ◽  
S. T. Dunn ◽  
J. H. Morrissey ◽  
...  
Keyword(s):  
Vitamin D ◽  
Vitamin D Receptor ◽  
Calcium Binding ◽  
Spatial Relationship ◽  
Pcr Primers ◽  
Rt Pcr ◽  
Calbindin D28k ◽  
Actin Mrna ◽  
Polymerase Chain ◽  
Convoluted Tubules

The spatial relationship between vitamin D receptor (VDR) and calbindin D28k [calcium binding protein D28k (CaBP-D28k)] gene expression within the murine kidney was studied by localizing their mRNAs in discrete nephron structures using reverse transcription-polymerase chain reaction (RT-PCR). Primers for beta-actin mRNA were used as a control for the presence of tissue during RT-PCR for CaBP-D28k mRNA. mRNA for CaBP-D28k was found only in distal convoluted tubules (DCTs), connecting tubules (CNTs), and cortical collecting ducts (CCDs). In contrast, VDR mRNA was detected in glomeruli, S2 proximal convoluted tubules, cortical thick ascending limbs of Henle's loop, DCTs, CNTs, and initial CCDs. The presence of both VDR and CaBP-D28k mRNA in DCTs, CNTs, and CCDs is consistent with the hypothesis that cacitriol acts via the VDR to stimulate CaBP-D28k synthesis. Conversely, the presence of VDR mRNA in other parts of the nephron suggests that calcitriol has genomically mediated actions within the kidney in addition to stimulation of CaBP-D28k synthesis.

scholarly journals Rapid Detection and Quantification of Rhynchosporium secalis in Barley Using a Polymerase Chain Reaction

2011 ◽  
Vol 42 (No. 3) ◽  
pp. 111-114
Author(s):  
J. Gubiš ◽  
M. Hudcovicová ◽  
M. Gubišová
Keyword(s):  
Gel Electrophoresis ◽  
Pcr Primers ◽  
Rhynchosporium Secalis ◽  
Agarose Gel Electrophoresis ◽  
Artificial Infection ◽  
Rt Pcr ◽  
Chain Reaction ◽  
Green Approach ◽  
Total Dna ◽  
Polymerase Chain

PCR primers for diagnosis of Rhynchosporium secalis in seed samples of barley were developed. For the quantification of the pathogen in seed samples a real-time PCR with SYBR Green approach was used. Amounts from 1.8 to 419.1 pg of R. secalis DNA per 100 ng of total DNA were detected in 18 samples of barley seeds contaminated by R. secalis in field conditions. The correctness of this quantitative analysis was checked using an artificial infection of seeds with 1, 2, 5 and 20% level of infection by R. secalis. The level of contamination of artificially infected samples decreased with a lowering amount of added seed powder contaminated by the pathogen, the correlation coefficient for this analysis was 0.98. While the primer pair used in these analyses shows cross-reactions with other pathogens (P. teres, Drechslera tritici-repentis, F. culmorum and F. poe), it is recommended to check the products of RT-PCR by agarose-gel electrophoresis, in which these pathogens are easily distinguishable from R. secalis by different lengths of the amplified fragments.  

scholarly journals Retrospective observational RT-PCR analyses on 688 babies born to 843 SARS-CoV-2 positive mothers, placental analyses and diagnostic analyses limitations suggest vertical transmission is possible.

2021 ◽  
Vol 13 (1) ◽  
pp. 53-66 ◽  
Author(s):  
G. Bahadur ◽  
M. Bhat ◽  
S. Acharya ◽  
D. Janga ◽  
B. Campbell ◽  
...  
Keyword(s):  
Vertical Transmission ◽  
Research Question ◽  
Diagnostic Testing ◽  
Placental Pathology ◽  
Rt Pcr ◽  
Chain Reaction ◽  
Serial Testing ◽  
Medical Termination ◽  
Polymerase Chain ◽  
Positive Results

Research question: Is there vertical transmission (from mother to baby antenatally or intrapartum) after SARS-CoV-2 (COVID-19) infected pregnancy? Study design: A systematic search related to SARS-CoV-2 (COVID-19), pregnancy, neonatal complications, viral and vertical transmission. The duration was from December 2019 to May 2020. Results: A total of 84 studies with 862 COVID positive women were included. Two studies had ongoing pregnancies while 82 studies included 705 babies, 1 miscarriage and 1 medical termination of pregnancy (MTOP). Most publications (50/84, 59.5%), reported small numbers (<5) of positive babies. From 75 studies, 18 babies were COVID-19 positive. The first reverse transcription polymerase chain reaction (RT-PCR) diagnostic test was done in 449 babies and 2 losses, 2nd RT-PCR was done in 82 babies, IgM tests were done in 28 babies, and IgG tests were done in 28 babies. On the first RT-PCR, 47 studies reported time of testing while 28 studies did not. Positive results in the first RT-PCR were seen in 14 babies. Earliest tested at birth and the average time of the result was 22 hours. Three babies with negative first RT-PCR became positive on the second RT-PCR at day 6, day 7 and at 24 hours which continued to be positive at 1 week. Four studies with a total of 4 placental swabs were positive demonstrating SARS-CoV-2 localised in the placenta. In 2 studies, 10 tests for amniotic fluid were positive for SARS-CoV-2. These 2 babies were found to be positive on RT-PCR on serial testing. Conclusion: Diagnostic testing combined with incubation period and placental pathology indicate a strong likelihood that intrapartum vertical transmission of SARS-CoV-2 (COVID-19) from mother to baby is possible.

scholarly journals Use of Degenerate Primers for Partial Sequencing and RT-PCR-Based Assays of Grapevine Leafroll-Associated Viruses 4 and 5

1998 ◽  
Vol 88 (11) ◽  
pp. 1238-1243 ◽  
Author(s):  
Geoffrey Routh ◽  
Yun-Ping Zhang ◽  
Pasquale Saldarelli ◽  
Adib Rowhani
Keyword(s):  
Heat Shock Protein 70 ◽  
Plasmid Vector ◽  
Pcr Primers ◽  
Pcr Detection ◽  
Rt Pcr ◽  
Chain Reaction ◽  
Degenerate Primers ◽  
Double Stranded Rna ◽  
Pcr Products ◽  
Polymerase Chain

Double-stranded RNA (dsRNA) was purified from grapevines infected with grapevine leafroll-associated viruses 4 (GLRaV-4) and 5 (GLRaV-5), two putative closteroviruses. Reverse-transcriptase polymerase chain reaction (RT-PCR) was performed on this dsRNA using degenerate oligonucleotides designed to amplify an approximately 550- to 650-nucleotide fragment from the heat shock protein 70 homolog (HSP70) of the known closteroviruses. RT-PCR products of the appropriate molecular weight were gel-isolated and cloned into the plasmid vector pGEM-T. Clones of RT-PCR products generated by using these primers on dsRNA isolated from a plant infected with GLRaV-4 were sequenced. This sequence was used to develop an immunocapture RT-PCR (IC-RT-PCR) detection protocol capable of detecting GLRaV-4. Similar clones were made from dsRNA isolated from a plant infected with GLRaV-5. These clones were also sequenced. The two sequences were compared, and RT-PCR primers were developed that were able to amplify cDNA from both. These experiments demonstrate that degenerate primers that amplify closterovirus HSP70 sequences can be used to successfully generate sequences useful for IC-RT-PCR detection of these viruses. These data also suggest that it is feasible to use HSP70 sequences to design PCR primers capable of more general PCR detection of multiple GLRaV serotypes. Lastly, the presence of closterovirus-like HSP70 sequences in these putative closteroviruses implies that they are indeed members of this taxonomic group.

scholarly journals Characteristics of Anti-SARS-CoV-2 Antibodies in Recovered COVID-19 Subjects

Viruses ◽  
2021 ◽  
Vol 13 (4) ◽  
pp. 697
Author(s):  
Angela Huynh ◽  
Donald M. Arnold ◽  
James W. Smith ◽  
Jane C. Moore ◽  
Ali Zhang ◽  
...  
Keyword(s):  
Population Level ◽  
Enzyme Linked Immunosorbent Assay ◽  
The Novel ◽  
Rt Pcr ◽  
S Protein ◽  
Viral Neutralization ◽  
Humoral Immune ◽  
Global Pandemic ◽  
Polymerase Chain ◽  
Ig G

Coronavirus Disease 2019 (COVID-19) is a global pandemic caused by the novel severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). While detection of SARS-CoV-2 by polymerase chain reaction with reverse transcription (RT-PCR) is currently used to diagnose acute COVID-19 infection, serological assays are needed to study the humoral immune response to SARS-CoV-2. Anti-SARS-CoV-2 immunoglobulin (Ig)G/A/M antibodies against spike (S) protein and its receptor-binding domain (RBD) were characterized in recovered subjects who were RT-PCR-positive (n = 153) and RT-PCR-negative (n = 55) using an enzyme-linked immunosorbent assay (ELISA). These antibodies were also further assessed for their ability to neutralize live SARS-CoV-2 virus. Anti-SARS-CoV-2 antibodies were detected in 90.9% of resolved subjects up to 180 days post-symptom onset. Anti-S protein and anti-RBD IgG titers correlated (r = 0.5157 and r = 0.6010, respectively) with viral neutralization. Of the RT-PCR-positive subjects, 22 (14.3%) did not have anti-SARS-CoV-2 antibodies; and of those, 17 had RT-PCR cycle threshold (Ct) values > 27. These high Ct values raise the possibility that these indeterminate results are from individuals who were not infected or had mild infection that failed to elicit an antibody response. This study highlights the importance of serological surveys to determine population-level immunity based on infection numbers as determined by RT-PCR.

scholarly journals A Peptide-Based Magnetic Chemiluminescence Enzyme Immunoassay for Serological Diagnosis of Coronavirus Disease 2019

2020 ◽  
Vol 222 (2) ◽  
pp. 189-193 ◽  
Author(s):  
Xue-fei Cai ◽  
Juan Chen ◽  
Jie- li Hu ◽  
Quan-xin Long ◽  
Hai-jun Deng ◽  
...  
Keyword(s):  
Real Time ◽  
Severe Pneumonia ◽  
Rt Pcr ◽  
Chain Reaction ◽  
Cutoff Value ◽  
Assay Performance ◽  
Positive Rate ◽  
Polymerase Chain ◽  
Ig G ◽  
Chemiluminescence Enzyme Immunoassay

Abstract Background Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), a novel β-coronavirus, causes severe pneumonia and has spread throughout the globe rapidly. The disease associated with SARS-CoV-2 infection is named coronavirus disease 2019 (COVID-19). To date, real-time reverse-transcription polymerase chain reaction (RT-PCR) is the only test able to confirm this infection. However, the accuracy of RT-PCR depends on several factors; variations in these factors might significantly lower the sensitivity of detection. Methods In this study, we developed a peptide-based luminescent immunoassay that detected immunoglobulin (Ig)G and IgM. The assay cutoff value was determined by evaluating the sera from healthy and infected patients for pathogens other than SARS-CoV-2. Results To evaluate assay performance, we detected IgG and IgM in the sera from confirmed patients. The positive rate of IgG and IgM was 71.4% and 57.2%, respectively. Conclusions Therefore, combining our immunoassay with real-time RT-PCR might enhance the diagnostic accuracy of COVID-19.

scholarly journals Bell's Palsy Associated with SARS-CoV-2 Infection in a 2-Year-Old Child

2021 ◽  
Author(s):  
Serina Bsales ◽  
Birk Olson ◽  
Sunanda Gaur ◽  
Dalya Chefitz ◽  
Mary Carayannopoulos ◽  
...  
Keyword(s):  
Antibody Test ◽  
Bell’S Palsy ◽  
Rt Pcr ◽  
Bell's Palsy ◽  
Pediatric Case ◽  
Positive Antibody ◽  
Child Patient ◽  
Triggering Factor ◽  
Polymerase Chain ◽  
Ig G

AbstractBell's palsy (BP) is an acute, unilateral facial nerve palsy (FNP) that is a diagnosis of exclusion, sometimes associated with infectious causes. In this article, we described a previously healthy 2-year-old child patient who presented with left-sided facial droop, positive severe acute respiratory syndrome-coronavirus-2 (SARS-CoV-2) real-time reverse transcription polymerase chain reaction (RT-PCR), positive SARS-CoV-2 immunoglobulin (Ig)-G antibody, and negative cerebrospinal fluid (CSF) SARS-CoV-2 (PCR and serology). This is the second reported pediatric case of BP in the setting of SARS-CoV-2, and the first in a child without comorbidities. Due to the positive antibody test, we presented the idea that SARS-CoV-2 could be a triggering factor of the FNP, possibly occurring in the later stages of disease.

scholarly journals Analysis and Forecasting of Global RT-PCR Primers for SARS-CoV-2

2020 ◽  
Author(s):  
Gowri Nayar ◽  
Ed Seabolt ◽  
Mark Kunitomi ◽  
Akshay Agarwal ◽  
Kristen L. Beck ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Reverse Transcription ◽  
Animal Cell ◽  
Geographic Location ◽  
High Specificity ◽  
Pcr Primers ◽  
Rt Pcr ◽  
Chain Reaction ◽  
Polymerase Chain ◽  
Over Time

Rapid tests for active SARS-CoV-2 infections rely on reverse transcription polymerase chain reaction (RT-PCR). RT-PCR uses reverse transcription of RNA into complementary DNA (cDNA) and amplification of specific DNA (primer and probe) targets using polymerase chain reaction (PCR). The technology makes rapid and specific identification of the virus possible based on sequence homology of nucleic acid sequence and is much faster than tissue culture or animal cell models. However, the technique can lose sensitivity over time as the virus evolves and the target sequences diverge from the selective primer sequences. Different primer sequences have been adopted in different geographic regions. As we rely on these existing RT-PCR primers to track and manage the spread of the Coronavirus, it is imperative to understand how SARS-CoV-2 mutations, over time and geographically, diverge from existing primers used today. In this study, we analyze the performance of the SARS-CoV-2 primers in use today by measuring the number of mismatches between primer sequence and genome targets over time and spatially. We find that there is a growing number of mismatches, an increase by 2% per month, as well as a high specificity of virus based on geographic location.

Sign in / Sign up

Export Citation Format

Share Document