scholarly journals Molecular Cloning and Sequencing of thearoA Gene from Actinobacillus pleuropneumoniaeand Its Use in a PCR Assay for Rapid Identification

1999 ◽  
Vol 37 (5) ◽  
pp. 1575-1578 ◽  
Author(s):  
Carmen Hernanz Moral ◽  
Alberto Cascón Soriano ◽  
María Sánchez Salazar ◽  
Javier Yugueros Marcos ◽  
Susana Suárez Ramos ◽  
...  
Keyword(s):  
Rapid Identification ◽  
Chromosomal Dna ◽  
Pcr Assay ◽  
Specific Pcr ◽  
Specificity And Sensitivity ◽  
Pcr Products ◽  
Dna Fragment ◽  
Specific Pcr Assay ◽  
Dna Specificity ◽  
K 12

The gene (aroA) of Actinobacillus pleuropneumoniae, serotype 2, encoding 5-enolpyruvylshikimate-3-phosphate synthase was cloned by complementation of the aroA mutation in Escherichia coli K-12 strain AB2829, and the nucleotide sequence was determined. A pair of primers from the 5′ and 3′ termini were selected to be the basis for development of a specific PCR assay. A DNA fragment of 1,025 bp was amplified from lysed A. pleuropneumoniaeserotypes 1 to 12 of biovar 1 or from isolated DNA. No PCR products were detected when chromosomal DNAs from other genera were used as target DNAs; however, a 1,025-bp DNA fragment was amplified whenActinobacillus equuli chromosomal DNA was used as a target, which could be easily differentiated by its NAD independence. The PCR assay developed was very sensitive, with lower detection limits of 12 CFU with A. pleuropneumoniae cells and 0.8 pg with extracted DNA. Specificity and sensitivity make this PCR assay a useful method for the rapid identification and diagnosis of A. pleuropneumoniae infections.

Rapid identification of cervus antlers by species-specific PCR assay

2019 ◽  
Vol 34 (9) ◽  
pp. 1315-1319 ◽  
Author(s):  
Yaya Yang ◽  
Yang Zheng ◽  
Beibei Lu ◽  
Zhaoqun Jiao ◽  
Liqun Chen ◽  
...  
Keyword(s):  
Rapid Identification ◽  
Pcr Assay ◽  
Specific Pcr ◽  
Specific Pcr Assay ◽  
Species Specific

scholarly journals Growth of Paecilomyces variotii in B0 (diesel), B100 (biodiesel) and B7 (blend), degradation and molecular detection

2015 ◽  
Vol 75 (3) ◽  
pp. 541-547 ◽  
Author(s):  
J Gassen ◽  
FM Bento ◽  
APG Frazzon ◽  
MF Ferrão ◽  
IV Marroni ◽  
...  
Keyword(s):  
Dry Weight ◽  
Infrared Analysis ◽  
Paecilomyces Variotii ◽  
Pcr Assay ◽  
Pseudallescheria Boydii ◽  
Specific Pcr ◽  
Fuel Storage ◽  
Oil Water ◽  
Specific Pcr Assay ◽  
Dna Specificity

AbstractThe introduction of biodiesel to diesel may allow the fuel to be more susceptible to microorganism growth, especially during incorrect storage. To evaluate the effect of adding biodiesel in pure diesel on the growth of Paecilomyces variotii, microcosms containing pure diesel (B0), blend diesel/biodiesel (B7) and pure biodiesel (B100) were used. In microcosm with minimal mineral medium and B0, B7 or B100, after 60 days, the biomass (dry weight) formed at interface oil-water in B7 and B100 was significantly higher when compared to that of B0. Infrared analysis showed reduction of the carbonile fraction in B7 and B100 suggesting formation of intermediate compounds in B7. To monitor possible contamination of fuel storage tank by P. variotii samples were collected and analysed by specific-PCR assay for detection of P. variotii spores in the aqueous phase. This method was able to detect a minimum of 103 spores ml–1, corresponding to 0.0144 ng µl–1 of DNA. Specificity was tested against Aspergillus fumigatus and Pseudallescheria boydii.

scholarly journals Molecular Analysis of the rfb O Antigen Gene Cluster of Salmonella enterica Serogroup O:6,14 and Development of a Serogroup-Specific PCR Assay

2003 ◽  
Vol 69 (10) ◽  
pp. 6099-6105 ◽  
Author(s):  
Collette Fitzgerald ◽  
Rachel Sherwood ◽  
Linda L. Gheesling ◽  
Frances W. Brenner ◽  
Patricia I. Fields
Keyword(s):  
Gene Cluster ◽  
Sequence Similarity ◽  
Rapid Identification ◽  
Homologous Sequence ◽  
Pcr Assay ◽  
Typing Method ◽  
Specific Test ◽  
O Antigen ◽  
Specific Pcr ◽  
Specific Pcr Assay

ABSTRACT The Kauffmann-White scheme for serotyping Salmonella recognizes 46 somatic (O) antigen groups, which together with detection of the flagellar (H) antigens form the basis for serotype identification. Although serotyping has become an invaluable typing method for epidemiological investigations of Salmonella, it does have some practical limitations. We have been characterizing the genes required for O and H antigen biosynthesis with the goal of developing a DNA-based system for the determination of serotype in Salmonella. The majority of the enzymes involved in O antigen biosynthesis are encoded by the rfb gene cluster. We report the sequencing of the rfb region from S. enterica serotype Sundsvall (serogroup O:6,14). The S. enterica serotype Sundsvall rfb region is 8.4 kb in length and comprises six open reading frames. When compared with other previously characterized rfb regions, the serogroup O:6,14 sequence is most related to serogroup C1. On the basis of DNA sequence similarity, we identified two genes from the mannose biosynthetic pathway, two mannosyl transferase genes, the O unit flippase gene and, possibly, the O antigen polymerase. The whole cluster is derived from a low-G+C-content organism. Comparative sequencing of an additional serogroup O:6,14 isolate (S. enterica serotype Carrau) revealed a highly homologous sequence, suggesting that O antigen factors O:24 and O:25 (additional O factors associated with serogroup O:6,14) are encoded outside the rfb gene cluster. We developed a serogroup O:6,14-specific PCR assay based on a region of the putative wzx (O antigen flippase) gene. This provides the basis for a sensitive and specific test for the rapid identification of Salmonella serogroup O:6,14.

Identification of Klebsiella oxytoca using a specific PCR assay targeting the polygalacturonase pehX gene

2003 ◽  
Vol 154 (8) ◽  
pp. 587-592 ◽  
Author(s):  
Gennadiy Kovtunovych ◽  
Tetyana Lytvynenko ◽  
Valentyna Negrutska ◽  
Olena Lar ◽  
Sylvain Brisse ◽  
...  
Keyword(s):  
Klebsiella Oxytoca ◽  
Pcr Assay ◽  
Specific Pcr ◽  
Specific Pcr Assay

scholarly journals Genus Level Identification of Mycobacteria from Clinical Specimens by Using an Easy-To-Handle Mycobacterium-Specific PCR Assay

1998 ◽  
Vol 36 (3) ◽  
pp. 614-617 ◽  
Author(s):  
Fritz Stauffer ◽  
Heinrich Haber ◽  
Armin Rieger ◽  
Robert Mutschlechner ◽  
Petra Hasenberger ◽  
...  
Keyword(s):  
Positive Result ◽  
Internal Control ◽  
Specific Probe ◽  
Culture Result ◽  
Pcr Assay ◽  
Clinical Specimens ◽  
Genus Level ◽  
Specific Pcr ◽  
Specific Pcr Assay ◽  
Culture Positive

An easy-to-handle Mycobacterium-specific PCR assay for detection of the presence of a wide range of mycobacterial species in clinical samples was evaluated. The performance of the genus probe was compared with the performance of probes specific forMycobacterium tuberculosis and Mycobacterium avium and with that of standard culture. In addition, the utility of an internal control in monitoring amplification inhibitors was studied. Of 545 respiratory and 325 nonrespiratory specimens (a total of 870 specimens), 58 (6.7%) showed the presence of amplification inhibitors, as determined by a negative result for the internal control. Of these 58 specimens, 31 (53%) were stool specimens; other material, even citrate blood after lysis of erythrocytes, did not pose a problem with regard to inhibition of PCR amplification. Eighty-one of the remaining 812 specimens had a positive Mycobacterium culture result. Of these culture-positive specimens, 58 (71.6%) showed a positive result with the Mycobacterium genus-specific probe. Seventy-two samples had a positive result with theMycobacterium-specific probe but a negative culture result. Of these 72 samples, 26 samples were regarded as true positive, either because the M. tuberculosis- or M. avium-specific probe was also positive at the same time or because other specimens from the same patient taken at the same time were culture positive. The sensitivity of theMycobacterium-specific probe was 78.5% and the specificity was 93.5%. This study showed that pretesting of clinical specimens for mycobacteria to the genus level with aMycobacterium-specific probe offers the routine clinical laboratory the possibility of detecting tuberculous and nontuberculous mycobacteria with one test. Furthermore, specimens testing positive with the genus-specific probe can be immediately identified with species-specific probes.

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