scholarly journals Species-specific PCR assay for the detection of Babesia odocoilei

2021 ◽  
pp. 104063872110329
Author(s):  
Hilary J. Burgess ◽  
Betty P. Lockerbie ◽  
Lisanework E. Ayalew ◽  
Antonia Dibernardo ◽  
Kristýna Hrazdilová ◽  
...  
Keyword(s):  
Dna Amplification ◽  
18S Rrna Gene ◽  
Rrna Gene ◽  
Species Sensitivity ◽  
Pcr Assay ◽  
Banding Patterns ◽  
Specific Pcr ◽  
Pcr Product ◽  
Specific Pcr Assay ◽  
Species Specific

We developed a PCR assay for the detection of Babesia odocoilei based on the 18S rRNA gene. Multiple specimens of B. odocoilei were examined, and the assay consistently produced a small specific PCR product of 306 bp. The PCR assay was also challenged with DNA from 13 other Babesia species and 2 Theileria species, originating from 10 different host species; however, nonspecific DNA amplification and multiple banding patterns were observed, and the amplicon banding patterns varied between different isolates of the same species. Sensitivity was determined to be 6.4 pg of DNA, and an estimated 0.0001% parasitism. This assay can be utilized for species-specific differential detection of B. odocoilei.

Species-specific PCR assay for L. infantum/L. donovani discrimination

Acta Tropica ◽  
2006 ◽  
Vol 100 (3) ◽  
pp. 241-245 ◽  
Author(s):  
Mallorie Hide ◽  
Anne-Laure Bañuls
Keyword(s):  
Pcr Assay ◽  
Specific Pcr ◽  
Specific Pcr Assay ◽  
Species Specific

Specific PCR Detection of Arcobacter butzleri, Arcobacter cryaerophilus, Arcobacter skirrowii, and Arcobacter cibarius in Chicken Meat

2009 ◽  
Vol 72 (7) ◽  
pp. 1491-1495 ◽  
Author(s):  
DANIELA PENTIMALLI ◽  
NICOLETTE PEGELS ◽  
TERESA GARCÍA ◽  
ROSARIO MARTÍN ◽  
ISABEL GONZÁLEZ
Keyword(s):  
Pcr Amplification ◽  
Chicken Meat ◽  
Pcr Analysis ◽  
Rrna Gene ◽  
Pcr Assay ◽  
Gyra Gene ◽  
Specific Primers ◽  
Specific Pcr ◽  
Arcobacter Butzleri ◽  
Species Specific

An enrichment PCR assay using species-specific primers was developed for the detection of Arcobacter butzleri, Arcobacter cryaerophilus, Arcobacter skirrowii, and Arcobacter cibarius in chicken meat. Primers for A. cryaerophilus, A. skirrowii, and A. cibarius were designed based on the gyrA gene to amplify nucleic acid fragments of 212, 257, and 145 bp, respectively. The A. butzleri–specific primers were designed flanking a 203-bp DNA fragment in the 16S rRNA gene. The specificity of the four primer pairs was assessed by PCR analysis of DNA from a panel of Arcobacter species, related Campylobacter, Helicobacter species, and other food bacteria. The applicability of the method was then validated by testing 42 fresh retail-purchased chicken samples in the PCR assay. An 18-h selective preenrichment step followed by PCR amplification with the four Arcobacter primer sets revealed the presence of Arcobacter spp. in 85.7% of the retail chicken samples analyzed. A. butzleri was the only species present in 50% of the samples, and 35.7% of the samples were positive for both A. butzleri and A. cryaerophilus. A. skirrowii and A. cibarius were not detected in any of the chicken samples analyzed. The enrichment PCR assay developed is a specific and rapid alternative for the survey of Arcobacter contamination in meat.

PCR Identification of Ruminant Tissue in Raw and Heat-Treated Meat Meals

2006 ◽  
Vol 69 (9) ◽  
pp. 2241-2247 ◽  
Author(s):  
JEONG CHUL HA ◽  
WAN TAE JUNG ◽  
YONG SUK NAM ◽  
TAE WHA MOON
Keyword(s):  
Dna Sequence ◽  
12S Rrna ◽  
Spongiform Encephalopathy ◽  
Rrna Gene ◽  
Specific Pcr ◽  
Pcr Product ◽  
Heat Treated ◽  
Mitochondrial 12S Rrna ◽  
Animal Feedstuffs ◽  
Species Specific

To control the spread of bovine spongiform encephalopathy in cattle through contaminated animal feedstuffs, screening of feed products is essential. We designed five pairs of primers to identify specifically raw and heat-treated tissue from cattle, sheep, goat, deer, and ruminants in general. A forward common primer was designed based on a conserved DNA sequence in the mitochondrial 12S rRNA–tRNAval–16S rRNA gene, and reverse primers were designed to hybridize with a species-specific DNA sequence for each species considered. All primers were developed to create a specific PCR product small enough (less than 200 bp) to be suitable for heat-treated material. To evaluate the effect of heat treatment, a severe sterilization condition (133°C at 300 kPa for 20 min) was chosen. Species-specific amplicons were obtained from all types of heat-treated meat meals. Analysis of laboratory-contaminated vegetable meals revealed that the detection limit of the assay was 0.05% for each species analyzed. This PCR-based analysis can be used as a routine method for detecting banned animal-derived ingredients in raw and heat-treated feedstuffs.

scholarly journals Molecular Fingerprinting of Cryptosporidium Oocysts Isolated during Water Monitoring

2006 ◽  
Vol 72 (8) ◽  
pp. 5428-5435 ◽  
Author(s):  
Rosely A. B. Nichols ◽  
Brian M. Campbell ◽  
Huw V. Smith
Keyword(s):  
Environmental Water ◽  
18S Rrna Gene ◽  
Water Monitoring ◽  
Rrna Gene ◽  
Molecular Fingerprinting ◽  
Oocyst Wall ◽  
Specific Pcr ◽  
Pcr Product ◽  
Allele Specific ◽  
Microscope Slides

ABSTRACT We developed and validated a PCR-based method for identifying Cryptosporidium species and/or genotypes present on oocyst-positive microscope slides. The method involves removing coverslips and oocysts from previously examined slides followed by DNA extraction. We tested four loci, the 18S rRNA gene (N18SDIAG and N18SXIAO), the Cryptosporidium oocyst wall protein (COWP) gene (STN-COWP), and the dihydrofolate reductase (dhfr) gene (by multiplex allele-specific PCR), for amplifying DNA from low densities of Cryptosporidium parvum oocysts experimentally seeded onto microscope slides. The N18SDIAG locus performed consistently better than the other three tested. Purified oocysts from humans infected with C. felis, C. hominis, and C. parvum and commercially purchased C. muris were used to determine the sensitivities of three loci (N18SDIAG, STN-COWP, and N18SXIAO) to detect low oocyst densities. The N18SDIAG primers provided the greatest number of positive results, followed by the N18SXIAO primers and then the STN-COWP primers. Some oocyst-positive slides failed to generate a PCR product at any of the loci tested, but the limit of sensitivity is not entirely based on oocyst number. Sixteen of 33 environmental water monitoring Cryptosporidium slides tested (oocyst numbers ranging from 1 to 130) contained mixed Cryptosporidium species. The species/genotypes most commonly found were C. muris or C. andersoni, C. hominis or C. parvum, and C. meleagridis or Cryptosporidium sp. cervine, ferret, and mouse genotypes. Oocysts on one slide contained Cryptosporidium muskrat genotype II DNA.

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