Random Amplified Polymorphic DNA Groups Research Articles

Phylogeny Study of 20 Selected Species of Zingiberaceae from Ex situ Collections in Peninsular Malaysia

Zingiberaceae is widely distributed in Malaysia, with 750 species and 31 genera. This family comprises a different number of subfamilies and genera according to different taxonomic classification methods — classical taxonomy: one subfamily and four tribes vs molecular taxonomy: four subfamilies and six tribes. However, the taxonomic classification of Zingiberaceae is still debated, especially the classical taxonomy. It is due to some Zingiberaceae species showing cryptic morphologies that make it difficult to classify them through classical taxonomy, which refers to the unique morphological characteristics of a tribe/species. Therefore, accurate taxonomic classification is required by using a molecular approach. In this study, 20 selected species of Zingiberaceae collected from the Agricultural Conservatory Park, Institute of Bioscience (IBS), Universiti Putra Malaysia (UPM) were taxonomically classified using a molecular method with the help of three random amplified polymorphic DNA (RAPD) and three inter simple sequence repeat (ISSR) markers until the tribe level. The combined RAPD and ISSR unweighted pair group method with arithmetic mean (UPGMA) phylogenetic tree was comparable to Zingiberaceae’s current molecular and classical taxonomy. The 20 selected species were grouped into three tribes (Alpinieae, Zingiberaceae, and Globbeae). This finding has contributed additional biological information to better manage the 20 Zingiberaceae species in the Agricultural Conservatory Park, IBS, UPM. Further studies are needed to explore the genetic diversities and properties of Zingiberaceae species.

Genotyping of Burkholderia pseudomallei Isolated From Patients in South-Western Coastal Region of India

Burkholderia pseudomallei is the causative agent of melioidosis. Various tools have been used to determine the genetic diversity in B. pseudomallei isolates. In this study, Random Amplified Polymorphic DNA (RAPD)-PCR and flagellin gene (fliC) based PCR-Restriction Fragment Length Polymorphism (RFLP) were used to genotype Indian clinical B. pseudomallei isolates. A total of 89 clinical isolates could be grouped in 6 groups (A through F) by RAPD-PCR analysis. Some of the isolates in various groups had identical banding pattern suggesting them to be epidemiologically related. The RAPD groups also correlated with MLST sequence types suggesting the utility of this easy to do typing method. The PCR- RFLP analysis suggested Type III to be the predominant type which is different from other RFLP types reported from Southeast Asia. In conclusion, the results of this study show that RAPD-PCR, a simple genotyping method, may be used for analyzing the B. pseudomallei isolates and also establish epidemiological relevant relatedness among them. The results of fliC PCR-RFLP further suggest the Indian isolates are different from other Southeast Asian isolates.

Genetic relatedness and recombination analysis of Allorhizobium vitis strains associated with grapevine crown gall outbreaks in Europe.

To analyse genetic diversity and epidemiological relationships among 54 strains of Allorhizobium vitis isolated in Europe during an 8-year period and to assess the relative contribution of mutation and recombination in shaping their diversity. By using random amplified polymorphic DNA (RAPD) PCR, strains studied were distributed into 12 genetic groups. Sequence analysis of dnaK, gyrB and recA housekeeping genes was employed to characterize a representative subcollection of 28 strains. A total of 15 different haplotypes were found. Nucleotide sequence analysis suggested the presence of recombination events in A. vitis, particularly affecting dnaK locus. Although prevalence of mutation over recombination was found, impact of recombination was about two times greater than mutation in the evolution of the housekeeping genes analysed. The RAPD analysis indicated high degree of genetic diversity among the strains. However, the most abundant RAPD group was composed of 35 strains, which could lead to the conclusion that they share a common origin and were distributed by the movement of infected grapevine planting material as a most common way of crossing long distances. Furthermore, it seems that recombination is acting as an important driving force in the evolution of A. vitis. As no substantial evidence of recombination was detected within recA gene fragment, this phylogenetic marker could be reliable to characterize phylogenetic relationships among A. vitis strains. We demonstrated clear epidemiological relationship between majority of strains studied, suggesting a need for more stringent phytosanitary measures in international trade. Moreover, this is the first study to report recombination in A. vitis.

Molecular and Phenotypic Characterization of Aerococcus viridans Associated with Subclinical Bovine Mastitis.

Aerococcus viridans is a wide spread bacterium in the environment and clinically this organism is associated with different diseases in animals and humans. However, the geno- and phenotypic characterization of A. viridans associated with bovine mastitis has not yet been reported. The objectives of this study were to investigate the genetic and phenotypic diversity of A. viridans isolates using three different molecular methods including 16S rRNA gene sequencing, pulsed-field gel electrophoresis and random amplified polymorphic DNA (RAPD) along with biochemical tests, including antimicrobial susceptibility test. In total, 60 A. viridans strains were cultured from dairy herds presenting with subclinical mastitis. The results of biochemical tests revealed that most of the isolates (75.0%) were accurately identified by API Rapid 20 Strep system and the majority of A. viridans strains (96.7%) were found to be catalase negative, while two (3.3%) isolates were weakly positive. All isolates were resistant to trimethoprim-sulfamethoxazole, followed by streptomycin (96.7%), tetracycline (65.0%) and clindamycin (56.7%) by minimum inhibition concentration-determining broth microdilution technique. As compared to the sequence of 16S rRNA gene, both PFGE and RAPD showed their capacities to discriminate the intra-species diversity of A. viridans. Furthermore, most of the isolates obtained from the same herd or region belonged to the same major RAPD group, which indicated that RAPD is an appropriate assay for tracking the origins of isolates and epidemiological studies of A. viridans. This is a novel approach to use three molecular techniques and to compare their efficiency regarding the genetic diversity of A. viridans. The data suggest that A. viridans associated with subclinical mastitis has a considerable phenotypic and genotypic diversity.

Alteration of bacterial communities and organic matter in microbial fuel cells (MFCs) supplied with soil and organic fertilizer

The alteration of the organic matter (OM) and the composition of bacterial community in microbial fuel cells (MFCs) supplied with soil (S) and a composted organic fertilizer (A) was examined at the beginning and at the end of 3 weeks of incubation under current-producing as well as no-current-producing conditions. Denaturing gradient gel electrophoresis revealed a significant alteration of the microbial community structure in MFCs generating electricity as compared with no-current-producing MFCs. The genetic diversity of cultivable bacterial communities was assessed by random amplified polymorphic DNA (RAPD) analysis of 106 bacterial isolates obtained by using both generic and elective media. Sequencing of the 16S rRNA genes of the more representative RAPD groups indicated that over 50.4% of the isolates from MFCs fed with S were Proteobacteria, 25.1% Firmicutes, and 24.5% Actinobacteria, whereas in MFCs supplied with A 100% of the dominant species belonged to γ-Proteobacteria. The chemical analysis performed by fractioning the OM and using thermal analysis showed that the amount of total organic carbon contained in the soluble phase of the electrochemically active chambers significantly decreased as compared to the no-current-producing systems, whereas the OM of the solid phase became more humified and aromatic along with electricity generation, suggesting a significant stimulation of a humification process of the OM. These findings demonstrated that electroactive bacteria are commonly present in aerobic organic substrates such as soil or a fertilizer and that MFCs could represent a powerful tool for exploring the mineralization and humification processes of the soil OM.

New method for early detection of two random amplified polymorphic DNA (RAPD) groups of Staphylococcus aureus causing bovine mastitis infection in Paraná State, Brazil

The aim of this work was to develop a fast and accurate molecular approach to allow early detection of two RAPD groups of S. aureus causing bovine mastitis. Seventy five S. aureus isolates from infected animals were characterized by RAPD. Genomic fragments isolated from the unique bands present in either group were cloned and sequenced. Based on the DNA sequences, specific primers were designed to allow for the simultaneous detection of either group by multiplex PCR of S. aureus DNA isolated from clinical and subclinical bovine mastitis. Results showed that these proposed primers set could be used to detect various clinical and subclinical S. aureus isolates as well as the detection of the microorganism in bulk milk. Their use as a specific method for effective and early diagnostic tool for S. aureus infection in dairy herds is suggested.

옥수수 이삭썩음병에 관여하는 Fusarium속균의 동정

In 2007, a total of 77 isolates of Fusarium spp. were obtained from ear rot symptoms of corns collected from 5 locations in Gangwon Province, Korea. The fungal isolates were identified based on their morphological features. Out of the isolates, fifteen isolates were identified as Fusarium verticillioides which formed microconidia in long chains on monophialides. Four isolates were identified as F. subglutinans which formed microconida only on false heads. Six isolates were identified as F. graminearum which produced red pigment in PDA culture. Besides these Fusarium species, F. napiform, F. nygamai, and F. oxysporum were identified from the rest isolates. To assess for genetic diversity of the isolates, a random amplified polymorphic DNA(RAPD) technique was carried out using URP primers. The results from the RAPD analysis showed that the isolates from corn were divided into 6 groups. These RAPD groups of the Fusarium species corresponded to morphological characters of the Fusarium species. The phylogenetic analysis of most isolates by DNA sequencing of EF-1<TEX>$\alpha$</TEX> gene corresponded to morphological characters of the Fusarium species. The results of pathogenicity tests by two inoculation methods revealed that F. verticillioides, F. graminearum and F. subglutinans are strongly pathogenic to corn stalks.

Determination of genetic diversity among Indian isolates of Rhizoctonia bataticola causing dry root rot of chickpea

Genetic diversity among 27 isolates (23 from chickpea and 4 from other host crops) of Rhizoctonia bataticola representing 11 different states of India was determined by random amplified polymorphic DNA (RAPD), internal transcribed spacer restriction fragment length polymorphism (ITSRFLP) and ITS sequencing. The isolates showed variability in virulence test. Unweighted paired group method with arithmetic average cluster analysis was used to group the isolates into distinct clusters. The clusters generated by RAPD grouped all the isolates into six categories at 40% genetic similarity. High level of diversity was observed among the isolates of different as well as same state. Some of the RAPD (OPN 4, OPN 12, and OPN 20) markers clearly distinguished majority of the isolates into the area specific groups. The ITS I, 5.8 rDNA and ITS II regions of 11 isolates representing different RAPD groups were amplified with primers ITS 1 and ITS 4 and digested with seven restriction enzymes. The restriction enzymes DraI, MboI, RsaI, and AluI were found to be suitable for differentiating the isolates into five categories by showing isolate specific ITS-RFLP patterns. The isolates were variable in their nucleotide sequences of the ITS regions. This is the first study on genetic diversity among chickpea isolates of R. bataticola.

Molecular Differentiation of Mycoplasma gallisepticum and Mycoplasma imitans Strains by Pulsed-Field Gel Electrophoresis and Random Amplified Polymorphic DNA

Pulsed-field gel electrophoresis (PFGE) and random amplified polymorphic DNA (RAPD) analysis were used to compare 21 Mycoplasma gallisepticum strains and five M. imitans strains. Each strain of M. gallisepticum typed by PFGE and RAPD methods was genetically quite unique and RAPD and PFGE fingerprinting enabled strain characterization. Relationships between the M. gallisepticum and M. imitans strains were established and dendrograms were drawn from PFGE and RAPD patterns. PFGE group A and RAPD group D were significantly associated with M. imitans strains (P < 0.05). Three M. imitans strains shared the same PFGE and RAPD patterns. The two M. gallisepticum vaccine strains had singular PFGE and RAPD patterns. Thus, PFGE and RAPD can be used to investigate disease outbreaks in vaccinated flocks or for epidemiological tracking. For M. gallisepticum, the RAPD and PFGE discriminatory powers were superior to 0.95 and the in vitro, in ovo and in vivo reproducibility of RAPD and PFGE was 100%. The RAPD drawback was the inconsistent band intensity complicating the interpretation of patterns, while the PFGE limit was its low typeability (86%). Thus, these two molecular typing methods seemed complementary for M. gallisepticum epidemiological studies.

Intraspecific diversity within avocado field isolates of Rosellinia necatrix from south-east Spain

Fifty-five isolates of Rosellinia necatrix, the cause of common avocado white root rot disease, were collected from south-east Spain and characterised according to their virulence behaviour and their molecular patterns to assess broader levels of genetic diversity. Virulence properties were revealed by in vitro inoculation on avocado plants. Differences in reaction types showed variability among these isolates. No sequence differences were observed when the internal transcribed spacer 1 (ITS1) and ITS2 regions and DNA fragments of the β-tubulin, adenosine triphosphatase and translation elongation factor 1 genes were explored in representive isolates from five virulence groups. Random amplified polymorphic DNA (RAPD) amplifications were also performed for each isolate using 19 random primers. Four of these primers revealed polymorphism among isolates and repetitive and discriminative bands were used to build an unweighted pair group with arithmetic mean tree. However, RAPD clustering showed low stability, and no correlation between RAPD and virulence groups was observed, possibly indicating high levels of sexual recombination.

Vegetative compatibility and molecular characterization of Fusarium graminearum isolates from the State of Paraná, Brazil

Fusarium graminearum isolates causing Fusarium head blight in wheat were collected in Brazil and analyzed by random amplified polymorphic DNA (RAPD) markers and vegetative compatibility grouping (VCG). Nitrate non-utilizing mutants (nit) from each isolate were paired to verify heterokaryon formation. Three VCGs were identified among F. graminearum isolates: VCG1 included F-2, F-3 and F-4 isolates; VCG2 included F-1, F-6 and F-9 isolates; VCG3 included F-5, F-7 and F-8 isolates. Based on PCR amplification with eight different primers, the isolates showed great genetic similarity among themselves. Dendrogram analysis demonstrated two RAPD groups: Group A, consisting of isolates F-2 and F-9, and Group B, composed of the remaining isolates. Results suggest the clonal origin of F. graminearum isolates.

Diversity of Phytophthora capsici in Northwest Spain: Analysis of Virulence, Metalaxyl Response, and Molecular Characterization.

Phytophthora crown rot, caused by Phytophthora capsici, is potentially the most destructive disease of pepper in Spain. Phenotypic and genetic diversity of 16 P. capsici isolates collected from 11 farms in northwest Spain was characterized based on virulence, mating type, sensitivity to metalaxyl, and genetic analysis using random amplified polymorphic DNA (RAPD) methods. Low variability was observed among the isolates in their metalaxyl response, with 87.5% being highly sensitive. No isolates of the A2 mating type were detected. More variability was found in the virulence assay, and isolates were classified into two groups according to their pathogenicity on a set of four pepper cultivar differentials. Genetic variation examined with eight RAPD primers generated 92 polymorphic bands and revealed the existence of different patterns among isolates. Cluster analysis using the unweighted pair-group method with arithmetic averages (UPGMA) separated the Spanish isolates into three RAPD groups and established a relationship between the Spanish population and a representative worldwide group of isolates. No correlation was found between groups obtained by RAPD analysis and groups defined by virulence or metalaxyl response.

Genetic Diversity of Didymella bryoniae for RAPD Profiles Substantiated by SCAR Marker in Korea

Twenty isolates of Didymella bryoniae were isolated from infected cucurbit plants in various growing areas of southern Korea in 2001 and 2002. Random Amplified Polymorphic DNA (RAPD) group [RG] I of D. bryoniae was more virulent than RG IV to watermelon. Virulence of the RG I isolate was strong to moderate to cucumber, whereas that of the RG IV varied from strong, moderate to weak. Two hundred seventy-three amplified fragments were produced with 40 primers, and were analyzed by a cluster analysis using UPGMA method with an arithmetic average program of NTSYSPC. At the distance level of 0.7, two major genomic DNA RAPD groups were differentiated among 20 isolates. The RG I included 7 isolates from watermelon and one isolate from melon, whereas the RG IV included 12 isolates from squash, cucumber, watermelon and melon. Amplification of internal transcribed spacer (ITS) region and small subunit rRNA region from the 20 isolates yielded respectively a single fragment. Restriction pattern with 12 restriction enzymes was identical for all isolates tested, suggesting that variation in the ITS and small subunit within the D. bryoniae were low. Amplification of the genomic DNAs of the tested isolates with the sequence characterized amplified regions (SCAR) primer RG IF-RG IR specific for RG I group resulted in a single band of 650bp fragment for 8 isolates out of the 20 isolates. Therefore, these 8 isolates could be assigned into RG I. The same experiments done with RG IIF-RG IIR resulted in no amplified PCR product for the 20 isolates tested. An about 1.4 kb-fragment amplified from the RG IV isolates was specifically hybridized with PCR fragments amplified from genomic DNAs of the RG IV isolates only, suggesting that this PCR product could be used for discriminating the RG IV isolates from the RG I isolates as well other fungal species.

Relationship among Lepista species determined by CAPS and RAPD

To determine the relationship of Australian members of the genus Lepista with those from other parts of the world, genetic variation of isolates representing 27 accessions was assayed by cleaved amplified polymorphic sequences (CAPS) and random amplified polymorphic DNA (RAPD). CAPS and RAPD identified eight and seven groups, respectively. CAPS Groups 1 and 2 and RAPD Group 1 consisted of French and Australian accessions classified as morphospecies L. nuda. CAPS Group 3 and RAPD Groups 2 and 2A consisted of mostly Australian isolates identified as L. sordida var. sordida or L. sordida var. umbonata. Isolates earlier identified as morphospecies L. sp. were also placed in CAPS Group 3 and RAPD Group 2A indicating that these isolates are L. sordida var. sordida. In addition, three smaller groups were distinguished. A French isolate of L. sordida var. sordida was placed in distinctly separate CAPS and RAPD groups to Australian L. sordida var. sordida Groups 4,4 respectively. A French isolate of L. sordida var. aianthina was placed in CAPS and RAPD Groups 3,3. An accession of L. saeva was placed in CAPS Group 6 and RAPD Group 5, separate from other isolates. RAPD Groups 6 and 7 consist, respectively, of Greek and American accessions of L. nuda that were only distantly related to the Australian and French accessions of this morphospecies: CAPS also separated these isolates from each other and from all other isolates. The data suggest that the classification of morphospecies and varieties within Lepista cannot be determined on the basis of morphology alone. The Greek and American accessions of L. nuda are separated from the French and Australian accessions and may not be L. nuda. Similarly, the Australian accessions currently classified as L. sordida var. sordida together with the accessions of L. sordida var. umbonata are distinct from the French accessions of L. sordida var. sordida suggesting that Australian isolates may represent a new species or variety. Further studies using a combination of morphological and genetic characterization are required to provide a more accurate understanding of Lepista species.

Characterization of Iranian Isolates of Fusarium oxysporum on the Basis of RAPD Analysis, Virulence and Vegetative Compatibility

AbstractFusarium yellow disease of chickpea (Cicer arietinum), caused by the vascular wilt pathogen Fusarium oxysporum, is a major production constraint in chickpea‐growing regions of Iran. Fifteen isolates of F. oxysporum were classified on the basis of vegetative compatibility (the ability to form heterokaryons). Vegetative compatibility was assessed by pairing nit mutants. The isolates were assigned to three vegetative compatibility groups (VCGs): VCG1, VCG2, and VCG3 with 5, 3, and 7 members, respectively. These isolates were also characterized by virulence assay. On the basis of disease severity on chickpea seedlings, they were divided into three groups: highly virulent (HV), moderately virulent (MV) and weakly virulent (WV). Twelve arbitrary decamer primers were tested on the fungal isolates by polymerase chain reaction. Random amplified polymorphic DNA (RAPD) analysis indicated considerable genetic variation among these isolates. Comparison of the results of VCG tests and the virulence assay indicated that all seven HV isolates belong to a single vegetative compatibility group (VCG3), and all three WV isolates are placed in VCG1. There was no relationship between RAPD groups and the other two characteristics of Iranian F. oxysporum isolates.

Characterization of Fusarium oxysporum Isolates Obtained from Cucumber in China by Pathogenicity, VCG, and RAPD.

Thirty-four isolates of Fusarium oxysporum, obtained in China from cucumber plants showing either Fusarium wilt (F. oxysporum f. sp. cucumerinum) or root and stem rot (F. oxysporum f. sp. radicis-cucumerinum) symptoms, were characterized by pathogenicity, vegetative compatibility, and random amplified polymorphic DNA (RAPD). Of these, 23 isolates were identified by pathogenicity as F. oxysporum f. sp. cucumerinum, and one as F. oxysporum f. sp. radicis-cucumerinum, while 10 isolates were avirulent on cucumber, melon, sponge gourd, and pumpkin. The Chinese isolates of F. oxysporum f. sp. cucumerinum were assigned to RAPD groups III and XXI and to vegetative compatibility group (VCG) 0183, four new VCGs, 0184 to 0187, and a single-member VCG included in the artificial VCG 018-. The Chinese isolate of F. oxysporum f. sp. radicis-cucumerinum was assigned to RAPD group I and bridging VCG 0260/0261. The occurrence of F. oxysporum f. sp. radicis-cucumerinum on cucumber is reported for the first time in China.

Population Structure of Magnaporthe grisea from North‐western Himalayas and its Implications for Blast Resistance Breeding of Rice

AbstractNeutral and pathogenicity markers were used to analyse the population structure of Magnaporthe grisea rice isolates from the north‐western Himalayan region of India. Random amplified polymorphic DNA (RAPD)‐based DNA fingerprinting of 48 rice isolates of M. grisea with five primers (OPA‐04, OPA‐10, OPA‐13, OPJ‐06 and OPJ‐19) showed a total of 65 RAPD bands, of which 54 were polymorphic. Cluster analysis of 48 rice isolates of M. grisea on the basis of these 65 RAPD bands revealed the presence of high genotypic diversity and continuous DNA fingerprint variation in the pathogen population. No correlation was observed between RAPD patterns and virulence characteristics of the pathogen. The observed population structure contrasted with presumed clonal reproductive behaviour of the pathogen and indicated the possibility of ongoing genetic recombination in the pathogen population. Analysis of the virulence organization of five RAPD groups (RG1–RG5) using 20 rice genotypes comprising at least 15 resistance genes revealed that no combination of resistance genes would confer resistance against all RAPD fingerprint groups present in the M. grisea rice population. The possible implications of the observed population structure of M. grisea for blast resistance breeding have been discussed.

Morphological and genetic characteristics of Flavobacterium columnare isolates: correlations with virulence in fish

Variability in pathogenicity of Flavobacterium columnare makes disease treatment difficult because there is currently no way to easily recognize those strains that warrant aggressive treatments. In order to identify suitable virulence markers, 17 isolates of F. columnare were cultured from six different fish species. The DNA from all isolates was analysed using random amplified polymorphic DNA (RAPD). Bootstrap analysis of the RAPD data produced a tree with three major groups supported by bootstrap scores of 80-100%. Virulence of the isolates was determined by bath exposure of channel catfish, Ictaluruspunctatus (Rafinesque), and golden shiners, Notemigonus crysoleucas (Mitchill), to broth cultures of F. columnare. In channel catfish, 13 of 17 isolates produced 100% mortality within 48 h post-exposure. All isolates of cyprinid fish origin clustered in a single RAPD group. At least two of the four isolates that were not virulent in channel catfish were of cyprinid fish origin. There was a wide variation in cell morphology between isolates with lengths of cells or cell chains ranging from 3 to 11 microm, even under identical culture conditions. Most of the shorter or single cell isolates fell into a single RAPD group. No clear association was identified between virulence and any other characteristic, including RAPD group.

Genetic Diversity of Ascochyta rabiei in Canada.

Assessment of variability of Ascochyta rabiei (teleomorph: Didymella rabiei) was based on virulence tests of 40 isolates and on random amplified polymorphic DNA (RAPD) analysis of 39 isolates from Canada. In addition, isolates of A. rabiei from other countries were assessed in the virulence (18 isolates) and RAPD (20 isolates) analyses. Seven isolates of A. lentis (teleo-morph: Didymella lentis) and two of A. pinodes (teleomorph: Mycosphaerella pinodes) also were included in the RAPD analysis. Significant line-isolate interactions in the virulence tests indicated that certain isolates were virulent only on certain lines. Canadian isolates were grouped into 14 pathotypes using eight chickpea differentials. These groupings also encompassed 17 of the 18 isolates from other countries. RAPD analysis of all 68 isolates using 8 primers produced 112 fragments, of which 96% were polymorphic. Similarities among A. rabiei isolates from Canada ranged from 20 to 100%. In the RAPD dendrogram, all five A. rabiei isolates from Australia, three of six from Syria, three of five from the United States, and one of two from India clustered within the major groups of Canadian isolates. There was a weak association between RAPD and pathotype groups. A. rabiei was 45% similar to A. lentis and only 14% similar to A. pinodes. The levels of DNA variability and virulence among isolates show that the population of A. rabiei in Canada is highly diverse.

Genetic variation among asexual progeny of Phytophthora infestans detected with RAPD and AFLP markers

Genotypic variation among 32 single‐zoospore isolates (SZI) of Phytophthora infestans, derived asexually from two hyphal‐tip parental isolates (PI‐105 and PI‐1) of the US‐8 genotype, was assessed with 80 random amplified polymorphic DNA (RAPD) primers and 18 amplified fragment length polymorphic DNA (AFLP) primer pairs. In previous investigations, the SZIs from parental isolate PI‐105 showed high levels of virulence variability and were differentiated into 14 races, whereas the SZIs from PI‐1 showed identical virulence to the parent. The purpose of this investigation was to determine if phenotypic variation observed among SZIs of P. infestans could be detected at the DNA level in these isolates. Polymorphism was detected with 51 RAPD primers and with all 18 AFLP primer pairs in PI‐105 SZIs. In SZIs from PI‐1, polymorphism was also detected with 25 RAPD primers and 17 AFLP primer pairs. Cluster analysis using the unweighted pair‐group method with arithmetic averages (UPGMA) separated the SZIs from parent PI‐105 into six virulence groups, 11 RAPD groups and three AFLP groups. Cluster analysis of PI‐1 SZIs, which all belong to the same virulence group, differentiated them into four RAPD groups and six AFLP groups. No close correlation among RAPD, AFLP and virulence groups could be established within the two progenies of SZIs. Results of this study suggest that there is a considerable level of inherent genetic variability among SZIs derived asexually from the same parental isolate. The possible mechanisms and implications of this genetic variation are discussed.