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Edwards, S.G., O’Callaghan, J., Dobson, D.W. 2002. PCR-based detection and quantification of mycotoxigenic fungi. Mycological Research 106 :1005–1025. Dobson D.W. PCR
-specific PCR-based assays for the detection of Fusarium species and a comparison with the whole seed agar plate method and trichothecene analysis. International Journal of Food Microbiology 103 :271–284. Gaba
be tested by PCR (based on 10% prevalence and 95% confidence, representing one piglet/litter) four times at 30-day intervals. PRRSV is extremely diverse in Europe ( Balka et al., 2018 ) and in Hungary ( Szabó et al., 2020 ); therefore, it
, Jung M , Meller S , Kristiansen G Improved PCR performance using template DNA from formalin-fixed and paraffin-embedded tissues by overcoming PCR inhibition . PLoS One 8 , 1 – 10 ( 2013 ) 3
, Thomson RB Jr. , Peterson LR , Kaul KL : Real-time PCR for detection and identification of Plasmodium spp . J Clin Microbiol 43 , 2435 – 2440 ( 2005 ) 8. Berry
A Mycoplasma bovis species-specific PCR assay has been developed with improvement of a previously described method (Ghadersohi et al., 1997). This test and its semi-nested version (Hayman and Hirst, 2003) did not function at all in our hands. A new reverse primer (Mbr2) was designed using previously published sequence data. For testing specificity, DNA was extracted from the most frequently occurring mycoplasma species and bacteria of bovine origin. The new PCR detected only Mycoplasma bovis. Moreover, no cross-reaction was observed with the genetically closest relative species, M. agalactiae. The target organism could be detected in a dose as low as 150 CFU ml-1 in broth cultures using ethidium-bromide-stained agarose gels.
the spread of MRSA and SA [ 6 , 7 ]. The German MRSA screening guidelines [ 8 ] recommend culture-based methods as the basis of MRSA screening. Due to the high negative predictive value (NPV) and rapid turnaround time, PCR-based methods are
biopsies. If only cysts are seen, discrimination of E. histolytica from non-pathogenic species can be performed either by PCR or antigen testing [ 2 ]. While diagnostic accuracy of modern real-time PCR for the detection of E. histolytica in stool
The goal of this study was to improve the diagnostic applicability of genus- and serovar- (S. Enteritidis and S. Typhimurium) specific PCR systems in screening faecal and caecal samples of poultry, poultry feed and poultrymeat for Salmonella, by keeping the opportunity to obtain Salmonella cultures from positive samples. Peptone broth pre-enrichment cultures of the samples were tested by PCR. In faecal and caecal samples from broiler chicks a strong inhibitory action was frequently observed. This could be reduced markedly by the addition of bovine serum albumin (BSA) acting as amplification facilitator. The results of testing pre-enrichment cultures from artificially contaminated faecal, poultry feed and poultrymeat samples (using S. Enteritidis, S. Typhimurium and S. Hadar as contaminants) suggest that the sensitivity of the above systems is 101-102 CFU g-1 sample. The testing of 95 caecal samples from slaughtered chicks resulted in 49% culture-positive and 76% PCR-positive samples. The suitability of a generic real-time PCR for testing faecal samples of poultry was also studied. Its detection limit for these samples was found to be lower than that of the diagnostic PCR system. Both methods reduced the time required for Salmonella detection to 24-30 h, and the advantage of the real-time PCR was its increased sensitivity. We have established a diagnostic and a real-time PCR system for rapid and reliable genus- and serovar- (S. Enteritidis and S. Typhimurium) specific detection of Salmonella for monitoring purposes in the poultry food chain. Sensitivity is equal to, or higher than, that of the standard bacterial culture method, and the method still provides the Salmonella culture if needed.
Recently, a PCR-derived method for serotyping Streptococcus pneumoniae has been devised to substitute the conventional antiserum phenotypic method. The method initially used a multiplex PCR reaction, dividing the isolates into 6 different groups based on the detected PCR gel pattern. In order to optimise and refine this crucial step, the Taguchi technique was employed, which can evaluate the individual effect of six parameters (in this case: primers, MgCl 2 , nucleotide mix, polymerase and buffer), with only 18 experiments; varying the parameter levels in an orthogonal matrix which suppresses the interactions between them. With this method, clear and sharp bands were observed in 5 experiments out of the 18, while the PCR did not work reliably in the remaining cases. In addition, the PCR-based technique could be rendered more economic by the 10-fold lowering of the quantities of two primers. The modified reaction yielded identical results to those obtained with the original method.Furthermore, we have designed serotype-specific primers for 11 new serotypes. The most important ones are those that can distinguish the very closely related, but equally important serotypes 6A and 6B.
