+ Site Statistics
+ Search Articles
+ Subscribe to Site Feeds
EurekaMag Most Shared ContentMost Shared
EurekaMag PDF Full Text ContentPDF Full Text
+ PDF Full Text
Request PDF Full TextRequest PDF Full Text
+ Follow Us
Follow on FacebookFollow on Facebook
Follow on TwitterFollow on Twitter
Follow on LinkedInFollow on LinkedIn

+ Translate

Quantitative 16S rDNA-targeted polymerase chain reaction and oligonucleotide hybridization for the detection of Paenibacillus azotofixans in soil and the wheat rhizosphere

Quantitative 16S rDNA-targeted polymerase chain reaction and oligonucleotide hybridization for the detection of Paenibacillus azotofixans in soil and the wheat rhizosphere

FEMS Microbiology Ecology 19(3): 153-164

A molecular method for the detection of Paenibacillus azotofixans in soil and the wheat rhizosphere was developed. The system consisted of polymerase chain reaction (PCR) amplification of part of the variable V1 to V4 regions of the 16S ribosomal RNA gene, followed by hybridization with a specific oligonucleotide probe homologous to part of the intervening region. In vitro specificity tests showed that the detection system worked specifically for P. azotofixans strains, and did not detect other Paenibacillus species or species of other bacterial genera. Vegetative cells of a rifampicin resistant P. azotofixans derivative were trackable in Flevo silt loam (FSL) soil in 24 h experiments using both selective plating and most probable number (MPN)-PCR combined with probing, and plate counts parallelled MPN-PCR estimations of numbers of specific targets. MPN-PCR allowed for the detection of down to 102 introduced cells per g of dry soil. Introduced P. azotofixans spores did not form colonies on selective plates, but were detectable via PCR. The P. azotofixans populations introduced into the silt loam soil suffered a slow decline of the detectable plate count over a period of 14 days. MPN-PCR revealed a similar decline of the number of specific DNA targets. Greater numbers of targets were found in wheat rhizosphere from Flevo silt loam soil, and these numbers persisted throughout the experiment. Soil drying resulted in enhanced persistence of the target sequences, whereas in a constantly moist soil the numbers of target sequences declined. Rewetting of dried soil resulted in declining target sequence numbers. The MPN-PCR detection method is adequate to assess the impact of stress conditions affecting P. azotofixans in FSL and probably other soils, since it abolishes the need for culturing or specific markers and is direct and unambiguous due to its high specificity.

(PDF emailed within 0-6 h: $19.90)

Accession: 009285625

Download citation: RISBibTeXText

DOI: 10.1111/j.1574-6941.1996.tb00208.x

Related references

rDNA targeted oligonucleotide primers for the identification of pathogenic yeasts in a polymerase chain reaction. Journal of Industrial Microbiology 14(6): 475-477, 1995

16S rRNA-targeted polymerase chain reaction and oligonucleotide hybridization to screen for Azoarcus spp., grass-associated diazotrophs. Applied and Environmental Microbiology 59(11): 3816-3824, 1993

Detection of Plasmodium sporozoites in mosquitoes by polymerase chain reaction and oligonucleotide rDNA probe, without dissection of the salivary glands. Medical and Veterinary Entomology 9(4): 433-437, 1995

Detection of Morganella morganii, a prolific histamine former, by the polymerase chain reaction assay with 16S rDNA-targeted primers. Journal of Food Protection 66(8): 1385-1392, 2003

Phenotypic and genetic diversity of Paenibacillus azotofixans strains isolated from the rhizoplane or rhizosphere soil of different grasses. Journal of Applied Microbiology 84(2): 216-226, 1998

Detection and identification method of three mycobacterium species and genus specific detection by polymerase chain reaction and DNA hybridization with alkaline phosphatase labeled oligonucleotide probe. Kansenshogaku Zasshi. Journal of the Japanese Association for Infectious Diseases 70(2): 141-150, 1996

Detection of RAS mutations in archival testicular germ cell tumors by polymerase chain reaction and oligonucleotide hybridization. Genes Chromosomes & Cancer. 5(2): 109-118, 1992

A 23S rDNA-targeted polymerase chain reaction-based system for detection of Staphylococcus aureus in meat starter cultures and dairy products. Journal of Food Protection 62(10): 1150-1156, 1999

Activity and DNA contamination of commercial polymerase chain reaction reagents for the universal 16S rDNA real-time polymerase chain reaction detection of bacterial pathogens in blood. Diagnostic Microbiology and Infectious Disease 66(1): 41-49, 2008

Comparison of paenibacillus azotofixans strains isolated from rhizoplane, rhizosphere, and non-root-associated soil from maize planted in two different brazilian soils. Applied and Environmental Microbiology 64(10): 3860-3868, 1998

Detection of Ralstonia solanacearum in tomato rhizosphere soil by polymerase chain reaction. Phytopathology 88(9 SUPPL ): S59, Sept, 1998

Rapid detection of Campylobacter fetus by polymerase chain reaction combined with non-radioactive hybridization using an oligonucleotide covalently bound to microwells. Molecular and Cellular Probes 14(4): 233-240, 2000

Comparison of the effectiveness of bacterial culture, 16S rDNA directed polymerase chain reaction, and checkerboard DNA-dNA hybridization for detection of Fusobacterium nucleatum in endodontic infections. Journal of Endodontics 28(2): 86-89, 2002

A consensus polymerase chain reaction-oligonucleotide hybridization approach for the detection of chromosomal translocations in pediatric bone and soft tissue sarcomas. American Journal of Clinical Pathology 104(6): 627-633, 1995