scholarly journals Detection of Brucella melitensis in bovine milk and milk products from apparently healthy animals in Egypt by real-time PCR

2014 ◽  
Vol 8 (10) ◽  
pp. 1339-1343 ◽  
Author(s):  
Gamal Wareth ◽  
Falk Melzer ◽  
Mandy C Elschner ◽  
Heinrich Neubauer ◽  
Uwe Roesler
Keyword(s):  
Public Health ◽  
Real Time ◽  
Real Time Pcr ◽  
Dairy Products ◽  
Enzyme Linked Immunosorbent Assay ◽  
Human Consumption ◽  
Rt Pcr ◽  
Milk Products ◽  
Milk Samples ◽  
Apparently Healthy

Introduction: Brucellosis in Egypt is an endemic disease among animals and humans. In endemic developing countries, dairy products produced from untreated milk are a potential threat to public health. The aim of this study was to detect brucellae in milk and milk products produced from apparently healthy animals to estimate the prevalence of contamination. Methodology: Two hundred and fifteen unpasteurized milk samples were collected from apparently healthy cattle (n = 72) and buffaloes (n = 128) reared on small farms, and from milk shops (n = 15) producing dairy products for human consumption. All milk samples were examined by indirect enzyme-linked immunosorbent assay (iELISA) and real-time PCR (RT-PCR) to detect Brucella antibodies and Brucella-specific DNA, respectively. Results: Using iELISA, anti-Brucella antibodies were detected in 34 samples (16%), while RT-PCR amplified Brucella-specific DNA from 17 milk samples (7.9%). Species-specific IS711 RT-PCR identified 16 of the RT-PCR-positive samples as containing B. melitensis DNA; 1 RT-PCR-positive sample was identified as containing B. abortus DNA. Conclusions: The detection of Brucella DNA in milk or milk products sold for human consumption, especially the highly pathogenic species B. melitensis, is of obvious concern. The shedding of Brucella spp. in milk poses an increasing threat to consumers in Egypt. Consumption of dairy products produced from non-pasteurized milk by individual farmers operating under poor hygienic conditions represents an unacceptable risk to public health.

scholarly journals Prevalence of Bovine viral diarrhea virus in cattle herds from Basrah and Nassirya Provinces by direct and indirect Elisa and Real time qPCR

2012 ◽  
Vol 11 (2) ◽  
pp. 1
Author(s):  
B. A. Jarullah, J. Aed Gati, and A. Saleh
Keyword(s):  
Real Time ◽  
Real Time Pcr ◽  
Indirect Elisa ◽  
Positive Sample ◽  
Enzyme Linked Immunosorbent Assay ◽  
Rt Pcr ◽  
Diarrhea Virus ◽  
Virus Rna ◽  
Two Samples ◽  
Cattle Herds

The current study was conducted to investigate the prevalence of BVD virus in Basrah and Nassirya city by using ELISA and RT-PCR. Two hundreds and eighty two samples of non vaccinated cattle sera samples collected from two regions of Iraq (188 samples from Nassirya city and 92 samples from Basrah city). Samples tested by Enzyme Linked Immunosorbent Assay (ELISA) antigen capture. Positive results were 20 samples ( 8 sample in Thi-Qar and 12 positive samples from Basrah). All samples submitted to indirect ELISA(IDEXX HerdCheck ELISA )for detect BVDV antibodies .Genotyping of all 20 positive samples to antigen detection were tested by Real time PCR, using Cador BVDV ½ kit, after extraction of virus RNA by QIAamp mini kit. The results revealed that there were 20 positive sample according to direct ELISA(Ag detection), while 66 sample were positive to indirect ELISA, as well as, the result of RT-PCR showed that there were two sample positive to BVDV type-1 (one sample form each city).Key words: BVDV, Genotype, ELISA, Iraq, Real time PCR.

scholarly journals Harmonization of Bordetella pertussis Real-Time PCR Diagnostics in the United States in 2012

2014 ◽  
Vol 53 (1) ◽  
pp. 118-123 ◽  
Author(s):  
Margaret M. Williams ◽  
Thomas H. Taylor ◽  
David M. Warshauer ◽  
Monte D. Martin ◽  
Ann M. Valley ◽  
...  
Keyword(s):  
Public Health ◽  
Real Time ◽  
Bordetella Pertussis ◽  
Real Time Pcr ◽  
The United States ◽  
Extraction Methods ◽  
Clinical Samples ◽  
Acid Extraction ◽  
Rt Pcr ◽  
Content Type

Real-time PCR (rt-PCR) is an important diagnostic tool for the identification ofBordetella pertussis,Bordetella holmesii, andBordetella parapertussis. Most U.S. public health laboratories (USPHLs) target IS481, present in 218 to 238 copies in theB. pertussisgenome and 32 to 65 copies inB. holmesii. The CDC developed a multitarget PCR assay to differentiateB. pertussis,B. holmesii, andB. parapertussisand provided protocols and training to 19 USPHLs. The 2012 performance exercise (PE) assessed the capability of USPHLs to detect these threeBordetellaspecies in clinical samples. Laboratories were recruited by the Wisconsin State Proficiency Testing program through the Association of Public Health Laboratories, in partnership with the CDC. Spring and fall PE panels contained 12 samples each of viableBordetellaand non-Bordetellaspecies in saline. Fifty and 53 USPHLs participated in the spring and fall PEs, respectively, using a variety of nucleic acid extraction methods, PCR platforms, and assays. Ninety-six percent and 94% of laboratories targeted IS481in spring and fall, respectively, in either singleplex or multiplex assays. In spring and fall, respectively, 72% and 79% of USPHLs differentiatedB. pertussisandB. holmesiiand 68% and 72% identifiedB. parapertussis. IS481cycle threshold (CT) values forB. pertussissamples had coefficients of variation (CV) ranging from 10% to 28%. Of the USPHLs that differentiatedB. pertussisandB. holmesii, sensitivity was 96% and specificity was 95% for the combined panels. The 2012 PE demonstrated increased harmonization of rt-PCRBordetelladiagnostic protocols in USPHLs compared to that of the previous survey.

scholarly journals Investigation of hepatitis A and E viruses in mussels collected from the Bosphorus, in Istanbul, Turkey – short communication

2018 ◽  
Vol 36 (No. 3) ◽  
pp. 215-220
Author(s):  
Huseyin Yilmaz ◽  
Utku Cizmecigil ◽  
Eda Tarakci ◽  
Ozge Aydin ◽  
Aysun Yilmaz ◽  
...  
Keyword(s):  
Public Health ◽  
Real Time ◽  
Real Time Pcr ◽  
Gel Electrophoresis ◽  
Aquatic Environment ◽  
Hepatitis A ◽  
Short Communication ◽  
Rt Pcr ◽  
Pooled Samples ◽  
Water Borne

Hepatitis A and E viruses (HAV and HEV) are recognised as food- and water-borne infections and shellfish are a well-documented source of these viruses. The presence of HAV and HEV in mussels has not previously been investigated in Bosphorus, Istanbul, Turkey, and therefore the aim of this study was to investigate the frequency of HAV and HEV in mussels collected from this location. A total of 92 pooled samples representing 736 mussels originating along the Bosphorus coast were collected from fish distributors. RNA was extracted using the RNeasy Plant Mini Kit and a TaqMan real-time RT–PCR was performed using primers specific to HAV and HEV. Amongst the 92 pooled samples tested, 3 (3.3%) were found to be positive for HAV by the TaqMan real-time RT-PCR. No HEV was detected in any of the mussels tested. A 174 bp product was observed on gel electrophoresis with HAV positive samples detected by SYBR Green real-time PCR after reverse transcription. This product was sequenced and confirmed to contain HAV sequences by alignment using the data in GenBank. This is the first report describing the presence of HAV in mussels harvested in the Bosphorus, Istanbul, Turkey, indicating the potential contamination of aquatic environment and risk to public health.

Zika virus infection in asymptomatic persons in Myanmar, 2018

2020 ◽  
Vol 114 (6) ◽  
pp. 440-447
Author(s):  
Mya Myat Ngwe Tun ◽  
Saw Wut Hmone ◽  
Aung Min Soe ◽  
Elizabeth Luvai ◽  
Khine Mya Nwe ◽  
...  
Keyword(s):  
Real Time ◽  
Zika Virus ◽  
Pacific Islands ◽  
Neutralization Test ◽  
Immunoglobulin M ◽  
Enzyme Linked Immunosorbent Assay ◽  
Rt Pcr ◽  
Zikv Infection ◽  
Healthy Persons ◽  
Apparently Healthy

Abstract Background Zika virus (ZIKV) is a mosquito-borne flavivirus. Outbreaks of ZIKV infection have occurred in Africa, Southeast Asia, the Pacific Islands, the Americas and the Caribbean. Although most ZIKV infections are asymptomatic, cases of neurological manifestations have been described. The aim of the present study was to identify the prevalence of ZIKV infection among the asymptomatic persons in Myanmar in 2018. Methods A total of 284 serum samples from apparently healthy persons were collected from Yangon, Myanmar in 2018. They were analysed for ZIKV infection by immunoglobulin M (IgM) capture enzyme-linked immunosorbent assay (ELISA), IgG indirect ELISA, 50% focus reduction neutralization test, real-time reverse transcription polymerase chain reaction (RT-PCR) and conventional RT-PCR. Results Of the 284 apparently healthy persons, 31.3% were positive for the presence of IgM against ZIKV and 94.3% were positive for anti-flavivirus IgG. Among the ZIKV IgM-positive samples, we confirmed ZIKV infection in 15.8% of asymptomatic persons by neutralization test and real-time RT-PCR. Conclusions We conclude that ZIKV infection was increasing among asymptomatic persons in the same area in Myanmar during 2018 compared with 2017. It is highly recommended to strengthen the surveillance system for ZIKV to prevent possible outbreaks.

scholarly journals Monitoring Current-Season Spread of Potato virus Y in Potato Fields Using ELISA and Real-Time RT-PCR

Plant Disease ◽  
2013 ◽  
Vol 97 (5) ◽  
pp. 641-644 ◽  
Author(s):  
Manphool S. Fageria ◽  
Mathuresh Singh ◽  
Upeksha Nanayakkara ◽  
Yvan Pelletier ◽  
Xianzhou Nie ◽  
...  
Keyword(s):  
Real Time ◽  
Potato Virus ◽  
Potato Virus Y ◽  
Enzyme Linked Immunosorbent Assay ◽  
Rt Pcr ◽  
Pcr Assay ◽  
Current Season ◽  
Seed Market ◽  
Polymerase Chain ◽  
Time Of Harvest

The current-season spread of Potato virus Y (PVY) was investigated in New Brunswick, Canada, in 11 potato fields planted with six different cultivars in 2009 and 2010. In all, 100 plants selected from each field were monitored for current-season PVY infections using enzyme-linked immunosorbent assay (ELISA) and real-time reverse-transcription polymerase chain reaction (RT-PCR) assay. Average PVY incidence in fields increased from 0.6% in 2009 and 2% in 2010 in the leaves to 20.3% in 2009 and 21.9% in 2010 in the tubers at the time of harvest. In individual fields, PVY incidence in tubers reached as high as 37% in 2009 and 39% in 2010 at the time of harvest. Real-time RT-PCR assay detected more samples with PVY from leaves than did ELISA. A higher number of positive samples was also detected with real-time RT-PCR from growing tubers compared with the leaves collected from the same plant at the same sampling time. PVY incidence determined from the growing tubers showed a significant positive correlation with the PVY incidence of tubers after harvest. Preharvest testing provides another option to growers to either top-kill the crop immediately to secure the seed market when the PVY incidence is low or leave the tubers to develop further for table or processing purposes when incidence of PVY is high.

scholarly journals PP-040 Real-time PCR assay for identification of Brucella abortus in bulk milk samples in Iran

2010 ◽  
Vol 14 ◽  
pp. S36-S37 ◽  
Author(s):  
S. Moshkelani ◽  
M. Javaheri Koupaei ◽  
S. Rabiei ◽  
A. Doosti
Keyword(s):  
Real Time ◽  
Brucella Abortus ◽  
Real Time Pcr ◽  
Pcr Assay ◽  
Milk Samples ◽  
Bulk Milk

Identification of L. casei and L. rhamnosus in the dairy products using Real-Time PCR assay

2015 ◽  
Vol 4 (5) ◽  
pp. 222-225
Author(s):  
K. G. Li ◽  
G. P. Pogossian ◽  
A. K. Moldagulova ◽  
E. E. Bekenova ◽  
A. Abdikadirova ◽  
...  
Keyword(s):  
Real Time ◽  
Real Time Pcr ◽  
Dairy Products ◽  
Production Control ◽  
High Efficiency ◽  
High Specificity ◽  
Industrial Test ◽  
Fermented Dairy Products ◽  
Species Specific ◽  
Fermented Dairy

  Lactobacilli are essential and important biological objects used in food pro-duction and medicine. One of the sufficient problems is fast, reliable and highly specific identification of lactobacilli in the scientific research and cur-rent production control. We represent two species-specific real-time PCR in the present study to discriminate L. rhamnosus and L. casei basing on the unique peptidoglycan-hydrolase genes p40 and p75 respectively. PCR pri-mers and probes were designed to provide high specificity discrimination via high temperature of PCR annealing stage. High efficiency of the reactions is provided by the size of amplified DNA fragments minimization. Reliable re-producibility of the target sequences amplification and fluorescence detec-tion provide a basis for the future creation of industrial test-systems for op-erational control in the production of fermented dairy products.

scholarly journals Toxoplasma gondii and pre-treatment protocols for polymerase chain reaction analysis of milk samples: a field trial in sheep from Southern Italy

2017 ◽  
Vol 6 (1) ◽  
Author(s):  
Alice Vismarra ◽  
Elena Barilli ◽  
Maura Miceli ◽  
Carlo Mangia ◽  
Cristina Bacci ◽  
...  
Keyword(s):  
Toxoplasma Gondii ◽  
Real Time ◽  
Dna Extraction ◽  
Real Time Pcr ◽  
Southern Italy ◽  
Raw Milk ◽  
Type I ◽  
Treatment Protocols ◽  
Milk Samples ◽  
Pre Treatment

Toxoplasmosis is a zoonotic disease caused by the protozoan <em>Toxoplasma gondii</em>. Ingestion of raw milk has been suggested as a risk for transmission to humans. Here the authors evaluated pre-treatment protocols for DNA extraction on <em>T. gondii</em> tachyzoite-spiked sheep milk with the aim of identifying the method that resulted in the most rapid and reliable PCR positivity. This protocol was then used to analyze milk samples form sheep from three different farms in southern Italy, including Real Time PCR for DNA quantification and PCR-RFLP for genotyping. The pre-treatment protocol using EDTA and Tris-HCl to remove casein gave the best results in the least amount of time compared to the others on spiked milk samples. One sample of 21 collected from sheep farms was positive on one-step PCR, Real Time PCR and resulted in a Type I genotype at one locus (SAG3). Milk usually contains a low number of tachyzoites and this could be a limiting factor for molecular identification. Our preliminary data has evaluated a rapid, cost-effective and sensitive protocol to treat milk before DNA extraction. The results of the present study also confirm the possibility of <em>T. gondii</em> transmission through consumption of raw milk and its unpasteurized derivatives.

scholarly journals Isolation and molecular characterization of Mycobacterium tuberculosis complex isolated from raw milk in some dairy farms in Egypt

2016 ◽  
Vol 5 (2) ◽  
pp. 105
Author(s):  
Heba Hussien ◽  
Eman Mahrous
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Real Time ◽  
Real Time Pcr ◽  
Culture Method ◽  
Raw Milk ◽  
Accurate Method ◽  
Tuberculosis Complex ◽  
Milk Samples ◽  
Source Of Infection

<p>This study was conducted to detect <em>Mycobacterium tuberculosis</em> complex in milk in three Egyptian Governorates; El-Sharkia, El-Menoufia and El-Behera Governorates. 300 milk samples were collected from tuberculin positive cases, 18 (6.0%) were shedding <em>Mycobacterium tuberculosis</em> complex in their milk which detected by real time PCR. On another hand, 170 milk samples were collected from tuberculin negative cases, 5 (2.9%) were shedding <em>Mycobacterium tuberculosis</em> complex in their milk which detected by real time PCR. All milk samples were examined by three techniques including Microscopic examination, culture and real time PCR. Real time PCR is more rapid and accurate method than microscopic and culture method. The isolated colonies from culture were examined by Multiplex PCR to demonstrate the source of infection either human or animal source.</p>

Complement C3 Genotyping of Slow and Fast Variants by Real Time PCR-High Resolution Melting

2012 ◽  
Vol 10 (3) ◽  
pp. 329-334 ◽  
Author(s):  
D.M. Valero-Hervás ◽  
P. Morales ◽  
M.J. Castro ◽  
P. Varela ◽  
M. Castillo-Rama ◽  
...  
Keyword(s):  
High Resolution ◽  
Real Time ◽  
Real Time Pcr ◽  
High Resolution Melting ◽  
Low Cost ◽  
Complement C3 ◽  
Amplification Refractory Mutation System ◽  
Rt Pcr ◽  
Dna Amount ◽  
Mutation System

“Slow” and “Fast” C3 complement variants (C3S and C3F) result from a g.304C>G polymorphism that changes arginine to glycine at position 102. C3 variants are associated with complement-mediated diseases and outcome in transplantation. In this work C3 genotyping is achieved by a Real Time PCR - High Resolution Melting (RT-PCR-HRM) optimized method. In an analysis of 49 subjects, 10.2% were C3FF, 36.7% were C3SF and 53.1% were C3SS. Allelic frequencies (70% for C3S and 30% for C3F) were in Hardy-Weinberg equilibrium and similar to those published previously. When comparing RT-PCR-HRM with the currently used Tetraprimer-Amplification Refractory Mutation System PCR (T-ARMS-PCR), coincidence was 93.8%. The procedure shown here includes a single primer pair and low DNA amount per reaction. Detection of C3 variants by RT-PCR-HRM is accurate, easy, fast and low cost, and it may be the method of choice for C3 genotyping.

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