Identification and DNA fingerprinting of Legionella strains by randomly amplified polymorphic DNA analysis

Abstract

The randomly amplified polymorphic DNA (RAPD) technique was used in the development of a fingerprinting (typing) and identification protocol for Legionella strains. Twenty decamer random oligonucleotide primers were screened for their discriminatory abilities. Two candidate primers were selected. By using a combination of these primers, RAPD analysis allowed for the differentiation between all different species, between the serogroups, and further differentiation between subtypes of the same serogroup. The usefulness of RAPD analysis was also evaluated with outbreak-related clinical and environmental isolates previously typed by the restriction fragment length polymorphism technique. RAPD analysis proved to be as accurate as other genotypic methods, reproducible, and highly discriminatory and is a valuable new alternative to traditional fingerprinting and identification of Legionella species and strains.

Diversity Suppression‐Subtractive Hybridization Array for Profiling Genomic DNA Polymorphisms

“…In conclusion, the method presented here is demonstrated to be a new, highly effective and reliable approach for profiling DNA polymorphisms, detecting unique polymorphic fragments and generating dendrograms. In comparison with other approaches used to probe DNA polymorphisms, such as RAPD (Bansal and McDonell 1997), SNP (Blazej et al 2003), multi-locus DNA fingerprinting (Coyer et al 1995) and AFLP (Botstein et al 1980), this method has several marked advantages. The most important feature of the method is that it combines SSH and DNA polymorphism detection to construct a platform for DNA polymorphism analysis that is independent of DNA sequence information.…”

“…Polymorphisms of genomic DNA between individuals or species have received increasing attention because they are useful for tracing genetic traits and studying biological diversity. Many molecular techniques have been developed for analyzing DNA polymorphism, such as ribotyping (Salloum et al 2002;Regnault et al 1997), random amplified polymorphic DNA (RAPD; Bansal and McDonell, 1997;LoPresti et al 1998), restriction fragment length polymorphism (RFLP; Botstein et al 1980), simple-sequence repeats (SSR; Weber and May 1989) and amplified fragment length polymorphism (AFLP; Vos et al 1995). These methods have been widely used in DNA polymorphism research, and have made great contributions to our understanding of genomic polymorphisms.…”

Presence of Legionellaceae in warm water supplies and typing of strains by polymerase chain reaction

“…Testing a set of 10 serogroup 1 reference strains primer pair Lpm-1\Lpm-2 were able to distinguish between 9 strains and primer ERIC2 was able to distinguish between 8 strains (data not shown). Bansal and McDonell [23] used PCR-based DNA fingerprinting technique with a combination of two random primers (double RAPD) to study its discriminatory ability with 67 well-defined legionella reference strains (representing 39 species) and 120 outbreak-related environmental and clinical isolates. For reference strains they obtained a unique strain-specific array of fragments for each species, serovar, and subtype.…”

Evaluation of arbitrarily primed polymerase chain reaction analysis for typing Legionella pneumophila

“…Even if the discriminatory power of AP-PCR could be marginally increased by using additional primers, the use of two different primers does not always expand it, as shown by Grattard et a1 [14]. In a recent study [23], a combination of primers was carefully selected from a larger set based on testing their discriminatory ability on a few isolates of Legionella. Table 1 shows strains (n=62) isolated during the period 1981-94 from environmental sources and nosocomial cases of legionellosis acquired in four hospitals, one nursing home (Courcelles) and a chemical plant (Seneffe), 11 community-acquired cases, and an air-conditioning system, and reference strain N C T C 11404.…”