scholarly journals Revisiting Brucellosis in Small Ruminants of Western Border Areas in Pakistan

Pathogens ◽  
2020 ◽  
Vol 9 (11) ◽  
pp. 929 ◽  
Author(s):  
Tariq Jamil ◽  
Khushal Khan Kasi ◽  
Falk Melzer ◽  
Muhammad Saqib ◽  
Qudrat Ullah ◽  
...  
Keyword(s):  
Real Time ◽  
Real Time Pcr ◽  
Small Ruminants ◽  
Kappa Statistics ◽  
Flock Size ◽  
Serological Tests ◽  
Perfect Agreement ◽  
Parallel Testing ◽  
Brucella Infection ◽  
Western Border

Brucellosis, globally known bacterial zoonosis, is endemic to Pakistan. B. abortus in bovines, B. melitensis in small ruminants and B. canis in dogs mainly cause this disease. A total of 1821 sera (1196 from sheep and 625 from goats) from animal herds near the Pakistan–Afghanistan border were collected. In parallel testing of sera for anti-Brucella antibodies (B. abortus and B. melitensis) was carried out by RBPT and indirect ELISA. The presence of Brucella DNA in sera was tested by real-time PCR. The overall percentage of seropositive samples was 0.99 (18/1821) by both tests. All positive samples originated from Baluchistan territory which translated into 1.76% (18/1021). None of the positive sera had signals for Brucella DNA and none of sera from goats carried detectable antibodies. Both tests showed an almost perfect agreement with Kappa statistics. The flock size was found to be associated with the presence of anti-Brucella antibodies. The samples of Khyber Pakhtunkhwa (KPK) tested negative in both serological tests and hence were not processed for real-time PCR. The present study shows the presence of anti-Brucella antibodies in sheep in the Baluchistan region of Pakistan. Diagnostic services need to be improved and test and slaughter policies might be implemented for eradication of Brucella infection in these areas. Awareness about the infection is needed at the farmer’s level. Isolation and molecular biology of the isolates could help with understanding the prevailing etiology in a better way.

scholarly journals Diagnostic Efficiency of Different Serological Tests and Real time PCR for Detecting Brucella Infection in Camels' Sera

2017 ◽  
Vol 24 (1) ◽  
pp. 132-146
Author(s):  
Mahmoud E.R. Hamdy ◽  
Mahmoud H. Abdel Haleem ◽  
Mohamed K. Al-kholi ◽  
Soliman S. Hazem
Keyword(s):  
Real Time ◽  
Real Time Pcr ◽  
Diagnostic Efficiency ◽  
Serological Tests ◽  
Brucella Infection

scholarly journals The Light and Shadow of Rapid Serological Tests for SARS-CoV-2 Infection: Results from a Study in a Large Emergency Department

2020 ◽  
Vol 17 (18) ◽  
pp. 6493
Author(s):  
Daniela Loconsole ◽  
Francesca Centrone ◽  
Caterina Morcavallo ◽  
Silvia Campanella ◽  
Anna Sallustio ◽  
...  
Keyword(s):  
Emergency Department ◽  
Real Time ◽  
Real Time Pcr ◽  
Respiratory Symptoms ◽  
Large Scale ◽  
Serological Test ◽  
Serological Tests ◽  
Serological Assay ◽  
Infected People ◽  
Screening Programs

A critical point in the management of the SARS-CoV-2 pandemic is the need to promptly identify the greatest number of infected people and to implement strict public health measures. In this study, the performance of a rapid serological test in a clinical setting was evaluated. Samples from 819 consecutive patients (with or without respiratory symptoms) admitted to a large Emergency Department were tested between 23 March and 21 April 2020. Patient samples were tested in a real-time PCR assay and a serological assay. In total, 148/819 patients (18.1%) tested positive for SARS-CoV-2 by real-time PCR. The serological test revealed that 70/819 patients (8.5%) had anti-SARS-CoV-2 IgM and/or IgG. The prevalence of anti-SARS-CoV-2 antibodies was significantly higher in patients with respiratory symptoms lasting for >7 days than in those with respiratory symptoms lasting for 0–7 days (p < 0.001). The serological assay had an overall sensitivity of 35.1% and an overall specificity of 97.3%. A high negative predictive value (96.7%) was reported for patients without respiratory symptoms. The results confirm that rapid serological assays alone are not sufficient for diagnosis of SARS-CoV-2 infection but can be incorporated into large-scale screening programs during periods in which the virus circulation is low.

scholarly journals Serologic and Molecular Detection of Mycoplasma gallisepticum in Layer Flocks in South Marmara Region of Turkey

2021 ◽  
pp. 29-37
Author(s):  
Elçin Günaydın ◽  
Özlem Kardoğan ◽  
Gülşen Goncagül ◽  
Yavuz Çokal
Keyword(s):  
Real Time ◽  
Real Time Pcr ◽  
Molecular Detection ◽  
Preventive Measures ◽  
Mycoplasma Gallisepticum ◽  
Marmara Region ◽  
Time Loss ◽  
Serological Tests ◽  
Primary Screening ◽  
Pooled Samples

Background: Due to the economic impacts of Mycoplasma gallisepticum (MG) infection in poultry, it is essential to have a fast, reliable and accurate diagnostic test to diagnose the infection. Aims: It was aimed to examine the presence of MG in the South Marmara Region of Turkey where extensive commercial layer flocks exist by RPA, ELISA and real-time PCR. Materials and Methods: In the study, 981 sera and 160 tracheal swab samples (20 swabs per each flock) obtained from eight layer flocks were examined for the presence of MG-antibody by RPA, ELISA, and the presence of MG by real-time PCR, respectively. Results: MG-seropositive flock rate was determined to be 100% by RPA. Twenty-three of the RPA positive sera in each flock LA, LB, LC, LD, LF, LG, and 17 RPA positive sera in flock LE (due to 17 positive RPA sera obtained) were examined for the presence of MG antibody by ELISA, and MG-seropositive flock rate was determined to be 87.5%. As a result of the examination of a total of 32 tracheal swab samples (20 swabs perflock/5 swabs=4 pooled samples, 8 flocksX4 pooled samples= 32 samples) for the presence of MG, real-time PCR positive flock rate was found to be 75%. Conclusion: To decide the flock whether it is infected or not and the initiate effective preventive measures against MG infection as soon as possible; serology should be applied simultaneously with bacteriology and/or PCR to prevent time loss due to shortcomings of serological tests used as primary screening test such as cross reactions, sensitivity and specificity problems.

scholarly journals A Novel Real-Time PCR-Based Screening Test with Pooled Fecal Samples for Bovine Johne’s Disease

2020 ◽  
Vol 58 (12) ◽  
Author(s):  
Satoko Kawaji ◽  
Reiko Nagata ◽  
Yasutaka Minegishi ◽  
Yumi Saruyama ◽  
Akiko Mita ◽  
...  
Keyword(s):  
Real Time ◽  
Real Time Pcr ◽  
Johne's Disease ◽  
Johne’S Disease ◽  
Enzyme Linked Immunosorbent Assay ◽  
Pcr Assay ◽  
Serological Tests ◽  
Fecal Samples ◽  
Content Type ◽  
Cattle Herds

ABSTRACT Johne’s disease (JD) is an economically important infectious disease in livestock farming caused by Mycobacterium avium subsp. paratuberculosis. As an alternative to serological tests, which are used mainly for the screening of whole herds, we developed a novel ResoLight-based real-time PCR (RL-PCR) assay with pooled fecal samples for the detection of fecal shedders in cattle herds. The RL-PCR assay included an internal amplification control (IC) which was amplified using the same primer pair as the target molecule M. avium subsp. paratuberculosis IS900 and differentiated based on melting temperatures. Individual fecal suspensions were pooled and concentrated by centrifugation to avoid a loss of sensitivity by the dilution effect. Combined with a DNA extraction kit (Johne-PureSpin; FASMAC), no inhibition of PCR amplification was observed with up to 15 fecal samples in a pool. The detection limit of RL-PCR at a pool size of 10 was 10 M. avium subsp. paratuberculosis organisms per gram of feces, which was comparable to that of individual testing. A total of 2,654 animals in 12 infected herds were screened by individual antibody-enzyme-linked immunosorbent assay (ELISA) and the RL-PCR assay using pooled feces. Fifty animals were diagnosed with JD through the screening by RL-PCR, compared with only 5 by ELISA (which were also positive in RL-PCR). In 7 JD-free herds, the results of 4 out of 327 pools (1.2%) were invalid due to the lack of IC amplification, and then animals were confirmed negative individually. Our results suggest that implementation of herd screening by pooled RL-PCR would advance the monitoring and control of JD in cattle herds.

Real time PCR assay for differentiation of Brucella abortus and Brucella melitensis

2017 ◽  
Author(s):  
Vinay Kumar ◽  
Sushila Maan ◽  
Aman Kumar ◽  
Kanisht Batra ◽  
Deepika Deepika ◽  
...  
Keyword(s):  
Real Time ◽  
Brucella Abortus ◽  
Real Time Pcr ◽  
Brucella Melitensis ◽  
Economic Losses ◽  
Conventional Pcr ◽  
Serological Tests ◽  
Detectable Difference ◽  
Pcr Assays ◽  
Respective Species

Brucellosis is one of the zoonotic diseases of major concern and can cause huge economic losses to livestock industry. Serological tests and bacterial isolation are considered as the gold standard assay for diagnosis of Brucella spp. but they are time-consuming, hazardous and lack specificity. To control and eradicate a disease, a confirmatory diagnostic method which is sensitive, quick and specific is the foremost requirement. Therefore in this study, we evaluated the performances of two newly designed TaqMan real-time PCR assays targeting the BruAB_0168 gene and BMEII0466 gene for Brucella abortus and Brucella melitensis (respectively). Both the assays were found to be highly specific in differentiation of respective species. Both the assays can detect as low as 0.02 fg of DNA and there was no detectable difference found in sensitivity of these two tests. R2 value and efficiency of these tests ranged from 0.992 - 0.998 and 100- 106%, respectively showing that these assays are highly efficient. Compared to conventional PCR assays these qPCR assays were 100 times higher sensitive. In conclusion, the present study showed that the developed real-time PCR assays are more sensitive, specific, have high reproducibility and repeatability and are faster than serological and conventional PCR methods for differentiation of Brucella abortus and Brucella melitensis.

scholarly journals Real-time PCR for detecting circulating dengue virus in the Guangdong Province of China in 2006

2008 ◽  
Vol 57 (12) ◽  
pp. 1547-1552 ◽  
Author(s):  
Zhijun Bai ◽  
Licheng Liu ◽  
Zeng Tu ◽  
Lisi Yao ◽  
Jianwei Liu ◽  
...  
Keyword(s):  
Dengue Virus ◽  
Real Time ◽  
Real Time Pcr ◽  
Cross Reactivity ◽  
Clinical Samples ◽  
Sequencing Analysis ◽  
Serum Samples ◽  
Serological Tests ◽  
Wide Range ◽  
Pcr Assays

Dengue virus (DENV) causes a wide range of diseases in humans, from the acute febrile illness dengue fever (DF) to life-threatening dengue haemorrhagic fever/dengue shock syndrome. We developed four real-time quantitative PCR assays for each serotype of DENV based on computational analysis. These assays had high sensitivity and specificity without cross-reactivity for the four serotypes. To evaluate the performance of these assays in detecting and typing the virus in clinical samples, we analysed 64 serum samples from Guangdong during 2006. The results showed that 71 % of those samples were positive by the DEN-1 assay. The DENV assay results, in agreement with the serological tests and sequencing analysis, showed that the pathogen resulting in the DF explosion in Guangdong in 2006 belonged to DEN-1. Compared to the serological assays, the real-time PCR assays that we developed were much more sensitive in the 1–3 days after onset of the symptoms.

scholarly journals Real-time PCR Biochip for On-Site Detection of Coxiella Burnetii in Ticks

2021 ◽  
Author(s):  
A-Tai Truong ◽  
Bo-Ram Yun ◽  
Jiyeon Lim ◽  
Subin Min ◽  
Mi-Sun Yoo ◽  
...  
Keyword(s):  
Real Time ◽  
Coxiella Burnetii ◽  
Real Time Pcr ◽  
Q Fever ◽  
Pcr Amplification ◽  
Pcr Detection ◽  
Economic Damage ◽  
Haemaphysalis Longicornis ◽  
Perfect Agreement ◽  
Close Relationship

Abstract Background: Q fever, a zoonosis caused by Coxiella burnetii, has adverse effects on public health. Ticks are the natural reservoirs of C. burnetii and they contribute to the transmission of the pathogen. A tool for rapid, sensitive, and accurate detection of C. burnetii from ticks is important for the prevention of Q fever. Methods: Ultra-rapid real-time PCR (UR-qPCR) as a chip-based real-time PCR system was developed for the detection of C. burnetii from ticks. The UR-qPCR system was established and evaluated for the rapidity, sensitivity, and specificity of C. burnetii detection. Results: C. burnetii was detected using UR-qPCR from 5,644 larval, nymphal, and adult ticks from 408 pools collected from livestock and epidemiologically linked environments in two provinces, Gangwon and Jeju, in Korea. Ticks from three species were identified; Haemaphysalis longicornis accounted for the highest number, present in 333 of 408 pools (81.62%), followed by Haemaphysalis flava in 62 pools (15.19%) and Ixodes nipponensis in 13 pools (3.19%). The rapidity and sensitivity of PCR detection was demonstrated with the sufficient amplification and detection of approximately 56 copies of C. burnetii DNA with only 20 min of PCR amplification. The kappa value for the diagnostic agreement between UR-qPCR and stationary qPCR was in perfect agreement (p = 1). PCR detection and sequencing indicated that C. burnetii was present in 5 of the 408 pools (1.23%), in which four pools contained H. longicornis and one pool contained H. flava. The infection rates of C. burnetii in the tick pools collected from Gangwon and Jeju Provinces were 1.70% and 0.58%, respectively. Phylogenetic analysis indicated a close relationship between the detected C. burnetii and those originated from goats, humans, and ticks in different countries, such as USA, France, Germany, and Serbia. Conclusions: The results of this study could be important for the prevention and control of Q fever in the two provinces. The UR-qPCR with its features of mobility, sensitivity, and rapidity is helpful for constructing early alert systems in the field for C. burnetii in ticks and for alleviating the transmission and economic damage due to Q fever.

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