scholarly journals Lymph-Node-Sampler-Based Nucleic Acid Detection Helps to Diagnose and Control African Swine Fever in Pig Production

2021 ◽  
Author(s):  
Xiaowen Li ◽  
Yang Li ◽  
Mingyu Fan ◽  
Shiran Fan ◽  
Wenchao Gao ◽  
...  
Keyword(s):  
Lymph Node ◽  
Nucleic Acid ◽  
African Swine Fever ◽  
Economic Losses ◽  
Nucleic Acid Detection ◽  
Serum Samples ◽  
Pig Production ◽  
Lymph Node Sampling ◽  
Time Period ◽  
And Control

Abstract Background: African swine fever (ASF) is a highly contagious hemorrhagic and transboundary animal disease. It has rapidly spread to many regions of the world and is responsible for serious production and economic losses. However, clinical diagnosis is impractical for the similar classic symptoms of ASF and several other normal diseases. How to make a definite diagnosis is a top priority. Results: In the present study, lymph node samples were collected by the patented “lymph node sampler”, which makes the sampling safer, more efficient and minimally invasive. The ASF virus (ASFV) contents of lymph node sample as well as serum, oral fluid, nasal and rectal swab samples from pig production line were detected by real-time PCR. The big data results demonstrated that the lymph node samples contained more stable and strongly positive ASFV nucleic acid than the porcine exudate and serum samples. Taking the lymph node sample is one of the assistant ways in diagnosing pigs suspected of having ASF. We supervised twenty farms that realized the accurate diagnosis and eliminating of the risk in the shortest time period by using lymph node sampler. Conclusions: Our findings suggest that the lymph node sample is an ideal tissue for diagnosing ASFV infection. The lymph node sampler is a convenient tool for lymph node sampling for practitioners.

scholarly journals Development and evaluation of duplex TaqMan real-time PCR assay for detection and differentiation of wide-type and MGF505-2R gene-deleted African swine fever viruses

2020 ◽  
Author(s):  
zhenhua Guo ◽  
Kunpeng Li ◽  
Songlin Qiao ◽  
Xinxin Chen ◽  
Ruiguang Deng ◽  
...  
Keyword(s):  
Real Time ◽  
Real Time Pcr ◽  
Large Scale ◽  
Limit Of Detection ◽  
African Swine Fever ◽  
Economic Losses ◽  
Clinical Samples ◽  
Diagnosis Method ◽  
Pcr Method ◽  
And Control

Abstract Background: African swine fever (ASF) is the most important disease to the pigs and cause serious economic losses to the countries with large-scale swine production. Vaccines are recognized as the most useful tool to prevent and control ASF virus (ASFV) infection. Currently, the MGF505 and MGF360 gene-deleted ASFVs or combined with CD2v deletion were confirmed to be the most promising vaccine candidates. Thus, it is essential to develop a diagnosis method to discriminate wide-type strain from the vaccines used.Results: In this study, we established a duplex TaqMan real-time PCR based on the B646L gene and MGF505-2R gene. The sequence alignment showed that the targeted regions of primers and probes are highly conserved in the genotype II ASFVs. The duplex real-time assay can specifically detect B646L and MGF505-2R gene single or simultaneously without cross-reaction with other porcine viruses tested. The limit of detection was 5.8 copies and 3.0 copies for the standard plasmids containing B646L and MGF505-2R genes, respectively. Clinical samples were tested in parallel by duplex real-time PCR and a commercial ASFV detection kit. The detection results of these two assays against B646L gene were well consistent.Conclusion: We successfully developed and evaluated a duplex TaqMan real-time PCR method which can effectively distinguish the wide type and MGF505 gene-deleted ASFVs. It would be a useful tool for the clinical diagnosis and control of ASF.

scholarly journals Cas12a-Based On-Site and Rapid Nucleic Acid Detection of African Swine Fever

2019 ◽  
Vol 10 ◽  
Author(s):  
Jing Bai ◽  
Haosi Lin ◽  
Haojian Li ◽  
Yang Zhou ◽  
Junshan Liu ◽  
...  
Keyword(s):  
Nucleic Acid ◽  
African Swine Fever ◽  
Nucleic Acid Detection

scholarly journals Development and evaluation of duplex TaqMan real-time PCR assay for detection and differentiation of wide-type and MGF505-2R gene-deleted African swine fever viruses

2020 ◽  
Vol 16 (1) ◽  
Author(s):  
Zhenhua Guo ◽  
Kunpeng Li ◽  
Songlin Qiao ◽  
Xin-xin Chen ◽  
Ruiguang Deng ◽  
...  
Keyword(s):  
Real Time ◽  
Real Time Pcr ◽  
Large Scale ◽  
Limit Of Detection ◽  
African Swine Fever ◽  
Economic Losses ◽  
Clinical Samples ◽  
Diagnosis Method ◽  
Pcr Method ◽  
And Control

Abstract Background African swine fever (ASF) is the most important disease to the pigs and cause serious economic losses to the countries with large-scale swine production. Vaccines are recognized as the most useful tool to prevent and control ASF virus (ASFV) infection. Currently, the MGF505 and MGF360 gene-deleted ASFVs or combined with CD2v deletion were confirmed to be the most promising vaccine candidates. Thus, it is essential to develop a diagnosis method to discriminate wide-type strain from the vaccines used. Results In this study, we established a duplex TaqMan real-time PCR based on the B646L gene and MGF505-2R gene. The sequence alignment showed that the targeted regions of primers and probes are highly conserved in the genotype II ASFVs. The duplex real-time assay can specifically detect B646L and MGF505-2R gene single or simultaneously without cross-reaction with other porcine viruses tested. The limit of detection was 5.8 copies and 3.0 copies for the standard plasmids containing B646L and MGF505-2R genes, respectively. Clinical samples were tested in parallel by duplex real-time PCR and a commercial ASFV detection kit. The detection results of these two assays against B646L gene were well consistent. Conclusion We successfully developed and evaluated a duplex TaqMan real-time PCR method which can effectively distinguish the wide type and MGF505 gene-deleted ASFVs. It would be a useful tool for the clinical diagnosis and control of ASF.

scholarly journals Development and Evaluation of Duplex TaqMan Real-Time PCR Assay for Detection and Differentiation of Wide-Type and MGF505 Gene-Deleted African Swine Fever Viruses

2020 ◽  
Author(s):  
Zhenhua Guo ◽  
Kunpeng Li ◽  
Songlin Qiao ◽  
Xinxin Chen ◽  
Ruiguang Deng ◽  
...  
Keyword(s):  
Real Time ◽  
Real Time Pcr ◽  
Large Scale ◽  
Limit Of Detection ◽  
African Swine Fever ◽  
Economic Losses ◽  
Clinical Samples ◽  
Diagnosis Method ◽  
Pcr Method ◽  
And Control

Abstract Background: African swine fever (ASF) is the most important disease to the pigs and cause serious economic losses to the countries with large-scale swine production. Vaccines are recognized as the most useful tool to prevent and control ASF virus (ASFV) infection. Currently, the MGF505 and MGF360 gene-deleted ASFVs or combined with CD2v deletion were confirmed to be the most promising vaccine candidates. Thus, it is essential to develop a diagnosis method to discriminate wide-type strain from the vaccines used. Results: In this study, we established a duplex TaqMan real-time PCR based on the B646L gene and MGF505-2R gene. The sequence alignment showed that the targeted regions of primers and probes are highly conserved in the genotype II ASFVs. The duplex real-time assay can specifically detect B646L and MGF505-2R gene single or simultaneously without cross-reaction with other porcine viruses tested. The limit of detection was 5.8 copies and 3.0 copies for the standard plasmids containing B646L and MGF505-2R genes, respectively. Clinical samples were tested in parallel by duplex real-time PCR and a commercial ASFV detection kit. The detection results of these two assays against B646L gene were well consistent. Conclusion: We successfully developed and evaluated a duplex TaqMan real-time PCR method which can effectively distinguish the wide type and MGF505 gene-deleted ASFVs. It would be a useful tool for the clinical diagnosis and control of ASF.

scholarly journals Sensitivity evaluation of 2019 novel coronavirus (SARS-CoV-2) RT-PCR detection kits and strategy to reduce false negative

2020 ◽  
Author(s):  
Yunying Zhou ◽  
Fengyan Pei ◽  
Li Wang ◽  
Huailong Zhao ◽  
Huanjie Li ◽  
...  
Keyword(s):  
Viral Load ◽  
Nucleic Acid ◽  
Prevention And Control ◽  
False Negative ◽  
Infection Prevention And Control ◽  
Nucleic Acid Detection ◽  
Rt Pcr ◽  
Mixed Sample ◽  
And Control ◽  
Selection Of

Abstract Background: In absence of effective vaccines, infection prevention and control of SARS-CoV-2 through diagnostic testing and quarantine is critical. Early detection and differential diagnosis of respiratory infections increases the chances for successful control of COVID-19 disease. The nucleic acid RT-PCR test is regarded as the current standard for molecular diagnosis with high sensitivity. However, the highest specificity confirmation target ORF1ab gene is considered to be less sensitive than other targets in clinical application. In addition, a large amount of recent evidence indicates that the initial missed diagnosis of asymptomatic patients with SARS-CoV-2 and discharged patients with “re-examination positive” may be due to low viral load, and the ability of rapid mutation of SARS-CoV-2 also increases the rate of false negative results. Moreover, the current used mixed sample nucleic acid detection is helpful to seek out the early community transmission of SARS-CoV-2, but the detection kit needs ultra-high detection sensitivity. Methods: From January to March 2020, 10 confirmed specimens of 2019-nCoV were recruited from three 2019-nCOV nucleic acid testing center. Five different amplification kits with three different primers and probes sources were selected. RT-PCR and continuous amplification of nasopharyngeal and oropharyngeal swabs and environmental specimens were performed to validate the sensitivity of the commercially available Nucleic acid test kits. Results: The results showed that ORF1ab gene can still be reported as positive at 1:10 dilution and the N gene even at 1:40 dilution with kit-1 through the verification of multiple positive samples, While other kits have less sensitive. The results in the suspicious range of weakly positive nasopharyngeal and oropharyngeal swabs and environmental specimens could be reported as positive after another re-amplification.Conclusions: Through evaluate the sensitivity of different nucleic acid detection kits, this study provide direct evidence for the selection of kits for mixed sample detection or make recommendations for the selection of validation kit, which is of great significance for the prevention and control of the current epidemic and the discharge criteria of low viral load patients.

scholarly journals Assessment of the Diagnostic Ability of Four Detection Methods Using Three Sample Types of COVID-19 Patients

2021 ◽  
Vol 11 ◽  
Author(s):  
Fei Yu ◽  
Guoliang Xie ◽  
Shufa Zheng ◽  
Dongsheng Han ◽  
Jiaqi Bao ◽  
...  
Keyword(s):  
Nucleic Acid ◽  
Reverse Transcription ◽  
Detection Methods ◽  
Nucleic Acid Detection ◽  
Sample Type ◽  
Rt Pcr ◽  
Chain Reactions ◽  
Polymerase Chain Reactions ◽  
Oropharyngeal Swab ◽  
And Control

BackgroundViral nucleic acid detection is considered the gold standard for the diagnosis of coronavirus disease 2019 (COVID-19), which is caused by SARS-CoV-2 infection. However, unsuitable sample types and laboratory detection kits/methods lead to misdiagnosis, which delays the prevention and control of the pandemic.MethodsWe compared four nucleic acid detection methods [two kinds of reverse transcription polymerase chain reactions (RT-PCR A: ORF1ab and N testing; RT-PCRB: only ORF1ab testing), reverse transcription recombinase aided amplification (RT-RAA) and droplet digital RT-PCR (dd-RT-PCR)] using 404 samples of 72 hospitalized COVID-19 patients, including oropharyngeal swab (OPS), nasopharyngeal swabs (NPS) and saliva after deep cough, to evaluate the best sample type and method for SARS-CoV-2 detection.ResultsAmong the four methods, dd-RT-PCR exhibited the highest positivity rate (93.0%), followed by RT-PCR B (91.2%) and RT-RAA (91.2%), while the positivity rate of RT-PCR A was only 71.9%. The viral load in OPS [24.90 copies/test (IQR 15.58-129.85)] was significantly lower than that in saliva [292.30 copies/test (IQR 20.20-8628.55)] and NPS [274.40 copies/test (IQR 33.10-2836.45)]. In addition, if OPS samples were tested alone by RT-PCR A, only 21.4% of the COVID-19 patients would be considered positive. The accuracy of all methods reached nearly 100% when saliva and NPS samples from the same patient were tested simultaneously.ConclusionsSARS-CoV-2 nucleic acid detection methods should be fully evaluated before use. High-positivity rate methods such as RT-RAA and dd-RT-PCR should be considered when possible. Furthermore, saliva after deep cough and NPS can greatly improve the accuracy of the diagnosis, and testing OPS alone is not recommended.

scholarly journals Development of a Potential Penside Colorimetric LAMP Assay Using Neutral Red for Detection of African Swine Fever Virus

2021 ◽  
Vol 12 ◽  
Author(s):  
Yang Wang ◽  
Junfei Dai ◽  
Yongsheng Liu ◽  
Jifei Yang ◽  
Qian Hou ◽  
...  
Keyword(s):  
African Swine Fever Virus ◽  
Animal Health ◽  
Colorimetric Detection ◽  
Monitoring Program ◽  
African Swine Fever ◽  
Lamp Assay ◽  
Neutral Red ◽  
Economic Losses ◽  
One Step ◽  
And Control

African swine fever (ASF) has caused huge economic losses to the swine industry worldwide. Since there is no commercial ASF vaccine available, an early diagnosis is extremely important to prevent and control the disease. In this study, ASF virus (ASFV) capsid protein-encoding gene (p72) was selected and used to design primers for establishing a one-step visual loop-mediated isothermal amplification (LAMP) assay with neutral red, a pH-sensitive dye, as the color shift indicator. Neutral red exhibited a sharp contrast of color change from faint orange (negative) to pink (positive) during LAMP for detection of ASFV. The designed primer set targeting highly conserved region of the p72 gene was highly specific to ASFV and showed no cross-reactivity with other swine viruses. The detection limit for the one-step visual LAMP developed was 10 copies/reaction based on the recombinant plasmid containing the p72 gene of ASFV. More importantly, the developed one-step visual LAMP showed high consistency with the results of the real-time polymerase chain reaction (qPCR) method recommended by World Organization for Animal Health (OIE). Furthermore, the results demonstrate that the colorimetric detection with this LAMP assay could be directly applied for the whole blood and serum samples without requiring genome extraction. Based on our results, the developed one-step visual LAMP assay is a promising penside diagnostic tool for development of early and cost-effective ASF monitoring program that would greatly contribute to the prevention and control of ASF.

scholarly journals Development of a chemiluminescence immunoassay to accurately detect African swine fever virus antibodies in serum

2021 ◽  
Author(s):  
Yong Yang ◽  
Changjie Lv ◽  
Junqing Fan ◽  
Ya Zhao ◽  
Lili Jiang ◽  
...  
Keyword(s):  
African Swine Fever Virus ◽  
Animal Health ◽  
Serum Antibody ◽  
African Swine Fever ◽  
Animal Husbandry ◽  
Economic Losses ◽  
Effective Vaccine ◽  
Serum Samples ◽  
Working Parameters ◽  
World Organization

AbstractThe outbreak of African swine fever (ASF) has caused significant economic losses to animal husbandry worldwide. Currently, there is no effective vaccine or treatment available to control the disease, and therefore, efficient disease control is dependent on early detection and diagnosis of ASF virus (ASFV). In this study, a chemiluminescent immunoassay (CLIA) was developed using the ASFV protein p54 as a serum diagnostic antigen and an anti-p54 monoclonal antibody. After optimizing the working parameters of the CLIA, the sensitivity of the established CLIA was 1:128, ASFV-specific serum antibody was identified, and there was no cross-reaction with other swine virus antibodies. After testing 49 clinical serum samples, the consistency rate between the CLIA and the World Organization for Animal Health (OIE) recommended commercial kit was 100%. Thus, this CLIA had a high degree of specificity, sensitivity, and reliability, and could be used as a rapid detection method for epidemiological investigations of ASFV infection.

scholarly journals Economic model for calculation of direct and indirect economical losses from African swine fever occurrence

2019 ◽  
Vol 22 (2) ◽  
pp. 227-236 ◽  
Author(s):  
P. Ivanova ◽  
E. Ivanova
Keyword(s):  
Economic Model ◽  
African Swine Fever ◽  
Control Measures ◽  
Economic Losses ◽  
Passive Surveillance ◽  
The Status ◽  
And Control ◽  
Disease Free ◽  
Indirect Economic Losses ◽  
Economical Losses

This study aimed to develop a model for calculation of direct and indirect economic losses in the event of African swine fever (ASF) occurrence. Calculation of the costs included the category of the affected holdings together with the specific biosecurity measures maintained therein and control measures to be imposed in the event of disease. Various scenarios for active and passive surveillance of the disease were developed, in order to prove the absence of virus circulation and to restore the status of the areas as ASF disease free-country.

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