Fingerprint Groups Research Articles

DNA Fingerprinting Analysis of Vegetative Compatibility Groups in Aspergillus flavus from a Peanut Field in Georgia.

The ability of species-specific DNA probe pAF28 to correctly match 75 strains of Aspergillus flavus isolated from a peanut field in Georgia with 1 of 44 distinct vegetative compatibility groupings (VCGs) was assessed. Multiple strains belonging to the same VCG typically produced identical DNA fingerprints, with the exception of VCG 17 and VCG 24, which contained strains that showed 83 and 87% similarity, respectively. A. flavus isolates sharing more than 80% of the fragments are recognized as belonging to the same DNA fingerprint group. Each VCG represented by a single isolate produced unique DNA fingerprints. The results provide further evidence that the pAF28 probe is able to distinguish A. flavus VCGs based on DNA fingerprints and can be used to predict the approximate number of VCGs in a sample population. The DNA probe also hybridized strongly and displayed multiple and distinct bands with other species in Aspergillus section Flavi: A. bombycis, A. caelatus, A. nomius, A. pseudotamarii, and A. tamarii. Although individual strains representing Aspergillus spp. in section Flavi produced DNA fingerprints with multiple bands, the banding patterns could not be used to classify these strains according to species.

Xanthomonas albilineans diversity and identification based on rep-PCR fingerprints.

PCR with BOX and ERIC primers was used to analyze DNA of Xanthomonas albilineans and other bacteria associated with sugarcane. Generated fingerprints permitted clear separation of X. albilineans from other bacteria and revealed variation within the species. Good agreement between fingerprint groups and geographic origin and serovars was observed.

Canadian fingerprint group Mytec acquires BII in US$25 million deal

Canadian fingerprint group Mytec acquires BII in US$25 million deal

Bacterial Leaf Spot Diseases of Leafy Crucifers in Oklahoma Caused by Pathovars of Xanthomonas campestris.

Fields of kale, spinach mustard, and turnip were severely damaged by bacterial leaf spots during 1994 to 1996. Symptoms included circular to angular necrotic lesions with yellow halos and water-soaking on the abaxial leaf surface. Yellow, mucoid strains isolated from leaf spots were identified as Xanthomonas campestris using Biolog. Four strains caused black lesions on stems of cabbage seedlings in an excised cotyledon assay, leaf spots and sunken dark lesions on petioles of spray-inoculated crucifers, and leaf spots on spray-inoculated tomato. These strains were classified as X. campestris pv. armoraciae. Most other strains from leafy crucifers and all strains from a cabbage field caused black rot in the cotyledon assay and in spray-inoculations. Many of these strains also caused leaf spots on collard and kale but not stem and petiole lesions. The strains causing black rot were classified as X. campestris pv. campestris. Cluster analysis of Biolog profiles yielded a small group that contained local strains of both pathovars, and a large group comprised of reference and local strains of each pathovar, and some local, nonpathogenic strains. Five fingerprint groups were identified by rep-polymerase chain reaction using the BOXA1R primer. Local and reference strains of each pathovar occurred in two of the groups. Two pathovars of X. campestris are involved in the leaf spot diseases. Both pathovars were recovered within several fields, and also were recovered along with Pseudomonas syringae pv. maculicola. This is the first report of Xanthomonas leaf spot caused by X. campestris pv. armoraciae in Oklahoma.

Occurrence of Bacteroides fragilis enterotoxin gene-carrying strains in Germany and the United States.

Ninety-three Bacteroides fragilis isolates from different geographic locations were analyzed for the presence of an enterotoxin-encoding gene. It was shown that blood culture isolates were more likely to carry this gene than strains from other sources. All enterotoxin-positive strains belonged to the PCR fingerprint group I.

Evidence of clonal dissemination of multidrug-resistant Streptococcus pneumoniae in Hong Kong.

The relationship between the phenotypic and genotypic characteristics of 105 penicillin-intermediate or -resistant Streptococcus pneumoniae isolates saved during 1994 to 1997 at the Prince of Wales Hospital and Pamela Youde Nethersole Eastern Hospital, Hong Kong, was studied. The pbp genes for penicillin-binding proteins 1a, 2b, and 2x for each isolate were amplified by PCR, and the products were digested with restriction enzymes HinfI and AluI. A combination of the pulsed-field gel electrophoresis (PFGE) profiles, pbp fingerprints, and phenotypic characteristics of capsular types and antibiograms enabled these isolates to be divided into four major groups. Seventy-four percent (78 of 105) of the strains, belonging to serotypes 23F, 19F, and 14, showed indistinguishable pbp fingerprint patterns (group A1, 1-1-1, 1-1-1), with PFGE patterns belonging to group A and its subtypes, suggesting that these strains were closely related. Eighty-three percent (65 of 78) of these isolates were also resistant to tetracycline, erythromycin, chloramphenicol, and trimethoprim. The type 23F isolates were indistinguishable from representative strains of the Spanish 23F clone by these molecular methods, indicating that these strains may be variants of the Spanish 23F clone. Serotype 6B accounted for 19% (20 of 105) of the isolates with reduced penicillin susceptibility and was made up of variants belonging to four different pbp fingerprint groups with the PFGE pattern group B, the predominant group being indistinguishable from that of the Spanish 6B clone. Other PFGE and fingerprint groups were mainly obtained from penicillin-susceptible strains of various serotypes. The results suggest that the rapid emergence of drug-resistant S. pneumoniae in Hong Kong has been due to the rapid dissemination of several successful clones.

Population Structure of the Rice Blast Fungus in Japan Examined by DNA Fingerprinting.

DNA fingerprinting with multilocus probes, MAGGY and MGR586, was conducted to investigate the population structure of the rice blast fungus Magnaporthe grisea in Japan. Two-hundred seventy-eight field isolates collected during 1993-1997 and 22 archival isolates collected in 1976 from various cultivars and locations were examined. Two fingerprint groups were identified at the 70% similarity level in each collection. A one-to-one correspondence was found between the groups in field isolates and the groups in archival isolates. These results suggest that two discrete lineages exist in the rice blast population. They were temporarily designated as JL1 and JL2 (Japanese Lineage). JL1 was a predominant lineage representing 97% and 77% of the field and archival isolates, respectively, and present throughout Japan. In addition, these two lineages corresponded to two of the five lineages previously detected in collections made before 1960, suggesting that the structure of the rice blast population in Japan had drastically changed during 1960-1976. The two lineages showed a very similar virulence spectrum, and no relationship could be recognized between lineages and pathotypes. MAGGY revealed robust groups which were similar to those revealed by MGR586, although the two elements differ in character and structure, suggesting that MAGGY could be used as an alternative probe for monitoring the population dynamics of the rice blast pathogen.

Recovery and characterization of spontaneous, selenate-resistant mutants of Magnaporthe grisea, the rice blast pathogen

There have been numerous biochemical mutants of Magnaporthe grisea previously described. In this study, selenate-resistant (sel) mutants were recovered and characterized. The mutants were recovered as spontaneous, fast-growing sectors arising from initially restricted colonies grown in the presence of 0.1% sodium selenate. Over 94% of the sectors generated were resistant to selenate, and approximately 70% of the selenate-resistant mutants were unable to utilize sulfate sulfur. The sulfate non-utilizing (sul) mutants could be distinguished from wild type by their thin expansive growth with little or no aerial mycelium on a defined minimal medium with sulfate as the sole sulfur source. However, growth of the sul mutant was comparable to the wild type when given L-methionine as a reduced sulfur source. Sul mutants were recovered from all 36 isolates examined representing four distinct MGR586 DNA fingerprint groups commonly found on rice in Arkansas. The frequency of sul mutants recovered from the 36 isolates varied on different media and among the different isolates examined. Generally, sectoring frequency and the number of sul mutants recovered was much higher on rice bran agar amended with selenate (0.1 to 1.4 sectors per colony) than on a defined minimal medium amended with selenate (0.0 to 0.7 sectors per colony). No sectors were observed on potato dextrose agar amended with 0.1% sodium selenate. The mean sectoring frequency among isolates in MGR586 DNA fingerprint group B was significantly higher than among isolates in three other fingerprint groups examined. Complementation tests with multiple sul mutants from a given isolate and sexual crosses among sul mutants of two fertile laboratory isolates indicated that the sul mutation was at a single locus. The sul mutants tested were pathogenic on rice in greenhouse tests and indistinguishable in their virulence phenotype to the corresponding wild type. The sul mutation was stable and may be useful as a marker in epidemiological studies.

Genotypic Diversity of Aspergillus parasiticus in an Illinois Corn Field.

Aspergillus parasiticus was isolated from direct platings of soil from a corn field near Kilbourne, Illinois. Soil contained 0.2 to 4.0 CFU of Aspergillus flavus and/or A. parasiticus per g of soil. Sixty isolates of A. parasiticus, each from a separately collected soil sample, were examined for ability to produce sclerotia and aflatoxins, and were subjected to DNA fingerprinting. PstI digests of total genomic DNA from each isolate were probed using the pAF28 repetitive sequence. Among 60 isolates analyzed, 33 (55%) distinct DNA fingerprint groups were identified (each group sharing less than 80% pAF28 band similarity), including 50 distinct genotypes (83%) with less than 100% pAF28 band similarity. A single A. parasiticus fingerprint group represented 13% of the sample population. The 83% genotypic diversity of the A. parasiticus population was equivalent to the 81% genotypic diversity recorded earlier for a population of 31 A. flavus isolates from the same field soil. Sclerotia were produced by 82% of the 50 A. parasiticus genotypes during dark incubation at 25°C. All isolates of A. parasiticus producedaflatoxin B1B2 and G1G2, whereas only 36% of the 31 A. flavus isolates from these soils produced aflatoxins.

β-Carotene inhibition of aflatoxin biosynthesis among Aspergillus flavus genotypes from Illinois corn

Thirty-nineAspergillus flavus genotypes (DNA fingerprinting) isolated from corn grown in a field near Kilbourne, Illinois were evaluated for their sensitivity to β-carotene (50 μg/ml) inhibition of aflatoxin B1 biosynthesis. Inhibition of aflatoxin was greater than 90% for 28 of the genotypes and >70% for 38 of the 39 genotypes. FiveA. flavus strains (4 fingerprint groups) isolated from molded raw peanuts, NRRL 3239, NRRL 3357, NRRL 6514, NRRL 6515 and NRRL 13135, produced greater quantities of aflatoxin than all 39 genotypes isolated from corn, and were less sensitive to β-carotene inhibition.Aspergillus flavus NRRL 3357 is commonly used as inoculum in variety trials for aflatoxin resistance. Isolate identity and sensitivity to potential inhibitors in corn can be critical in assessing corn resistance to aflatoxin.

A Survey for Strains of Xylella fastidiosa in Citrus Affected by Citrus Variegated Chlorosis and Citrus Blight in Brazil.

Polymerase chain reaction amplification of DNA from various strains of Xylella fastidiosa with tRNA consensus primers produced three different fingerprint groups. The citrus variegated chlorosis (CVC) and mulberry leaf scorch strains were unique and readily separated from each other and all other strains tested. Internal primers were designed based on the sequence of a DNA fragment unique to the CVC strain. An assay was developed with a mixture of these primers and those reported to detect 18 strains of X. fastidiosa. The assay was used to survey citrus in Brazil. The strain identified to be the cause of CVC was found in constant association with trees with CVC symptoms. On occasion, trees with no symptoms were found to have the CVC strain; this was presumably due to presymptomatic infections. No other strains were found in this survey, and X. fastidiosa was not associated with citrus blight.

DNA fingerprinting of Candida rugosa via repetitive sequence-based PCR.

A repetitive sequence-based PCR (rep-PCR) technique was developed to characterize the genotypic relatedness among Candida rugosa isolates. Two repetitive sequences, viz., Care-2 and Com29 from Candida albicans, were used to design primers Ca-21, Ca-22, and Com-21, respectively. When used alone or in combination, these primers generated discriminatory fingerprints by amplifying the adjacent variable regions of the genome. Twenty-three isolates from burn patients, eight from other human sources, and four C. rugosa isolates pathogenic in animals were placed into nine fingerprinting groups. Different primers placed these isolates into identical groups, indicating that rep-PCR is a specific and reproducible technique for molecular characterization of C. rugosa. Moreover, these primers unequivocally discriminated among other important Candida species such as C. albicans, C. glabrata, C. tropicalis, C. krusei, C. parapsilosis, C. kefyr, and C. lusitaniae. These data confirm the conservation of repetitive sequences in Candida species. Because of its ease and sensitivity, rep-PCR offers a relatively rapid and discriminatory method for molecular typing of C. rugosa in outbreaks.

Identification and grouping of Newcastle disease virus strains by restriction site analysis of a region from the F gene.

A 75% region of the F gene (between nucleotides 334 and 1682) of Newcastle disease virus (NDV) RNA was amplified by reverse transcription polymerase chain reaction (RT-PCR). PCR products were cleaved by three restriction endonucleases and the positions of thirty cleavage sites were mapped in more than 200 NDV strains. Restrictions site analysis established six major groups of NDV isolates and unique fingerprints of vaccine strains. Group I comprised lentogenic strains isolated mainly from waterfowl with some from chickens. "Old" (prior to 1960s) North American isolates of varying virulence including lentogenic and mesogenic vaccine strains belonged to group II. Group III included two early isolates from the Far East. Early European strains (Herts 33 and Italien) of the first panzootic (starting in the late 1920s) and their descendants with some modifications were placed into group IV. NDV strains isolated during the second panzootic of chickens (starting in the early 1960s) were classified into two groups. Group V included strains originating in imported psittacines and in epizootics of chickens in the early 1970s. Group V1 comprised strains from the Middle East in the late 1960s and later isolates from Asia and Europe. Pigeon paramyxovirus-1 strains that were responsible for the third panzootic formed a distinct subgroup in group V1. Our grouping of NDV strains has confirmed group differences established by monoclonal antibodies. It is concluded that restriction site analysis of F gene PCR amplicons is a relatively fast, simple and reliable method for the differentiation and identification of NDV strains.

The genetic diversity and epizootiology of infectious hematopoietic necrosis virus

Infectious hematopoietic necrosis virus (IHNV) is a rhabdovirus which causes a serious disease in salmondd fish. The T1 ribonuclease fingerprinttin method was used to compare the RNA genomes of 26 isolates of IHNV recovered from sockeye salmon ( Oncorhynchus nerka), chinook salmon ( O. tshawytscha), and steelhead trout ( O. mykiss) throughout the enzootic portion of western North America. Most of the isolates as a source of genetic variation. In from a single year (1987) to limit time of isolation as a source of genetic variation. In addition, isolates from different years collected at three sites were analyzed to investigate genetic drift or evolution of IHNV within specific locations. All of the isolates examined by T1 fingerprint analysis contained less than a 50% variation in spot location and were represented by a single fingerprint group. The observed variation was estimated to correspond to less than 5% variation in the nucleic acid sequence. However, sufficient variation was detected to separate the isolates into four subgroups which appeared to correlate to different geographic regions. Host species appeared not to be a significant source of variation. The evolutionary and epizootiologic significance of these findings and their relationship to other evidence of genetic variation in IHNV isolates are discussed.

Molecular typing of Cryptococcus neoformans serotype D clinical isolates

Cryptococcus neoformans serotype A is responsible for the majority of cryptococcal infections in AIDS patients. In France, approximately 17% of the patients are infected with serotype D, regardless of their human immunodeficiency virus status. In a retrospective study of 273 patients, we found that serotype D was unevenly distributed in France. We wondered if this was related to the yeast's genetic background. We used karyotyping and DNA fingerprints generated by UT-4p to analyze 40 serotype D clinical isolates. We found an extensive polymorphism, with only two conserved karyotypes from drug-addicted patients living in the same area. Although highly variable, the DNA fingerprints were classified into 10 groups. Four pairs of isolates were identical; three of these pairs were from patients living in the same area, but there was no other correlation with the geographical area. The two isolates with identical karyotypes belonged to the same fingerprint group. Five of the six isolates that made up fingerprint group I were recovered from drug-addicted patients (P < 0.002; chi-square), and all five isolates found in fingerprint group III were from male homosexual patients (P < 0.02). Finally, five of the seven isolates from patients with cryptococcal pneumonia were classified as fingerprint group V (P < 0.04). These results suggest that there are possible relationships between characteristics of the isolates and body localization or even risk factors. Results of the present study warrant other studies on isolates of all serotypes and on isolates from clinical and environmental sources.

Mitochondrial DNA fingerprinting of Acanthamoeba spp. isolated from clinical and environmental sources

Restriction fragment length polymorphism analysis of mitochondrial DNA (mtDNA fingerprinting) was evaluated as an epidemiologic tool for identifying potential reservoirs of Acanthamoeba infection. Fingerprints for 15 clinical isolates recovered by our affiliated laboratories were compared with those for 25 environmental isolates from western Washington State and 10 American Type Culture Collection (ATCC) strains. Seven different fingerprint groups emerged from the analysis of clinical isolates with six selected restriction enzymes (BamHI, BglII, EcoRI, HindIII, KpnI, and SalI). Fourteen (56%) environmental and 4 (40%) ATCC isolates displayed fingerprints similar to those of clinical isolates. In all, five of the seven groups contained one or more environmental and/or ATCC isolates. Comparisons with published mtDNA fingerprints for Acanthamoeba isolates showed that two groups have counterparts in Europe and Japan and in Europe and Australia. The inclusion of environmental isolates demonstrated that the most common clinical isolates do have counterparts readily recoverable from the surrounding environment and that some of these counterparts appear to be geographically widespread.

Molecular epidemiology of penicillin-resistant pneumococci isolated in Nairobi, Kenya.

A total of 26% of the pneumococci isolated from an outpatient clinic in Nairobi, Kenya, during 1991 to 1992 had intermediate levels of penicillin resistance. Gene fingerprinting and DNA sequencing were used to distinguish the penicillin-binding protein (PBP) 1A, 2B, and 2X genes in 23 resistant isolates. Isolates were grouped into those that had identical forms of each of the three PBP genes (fingerprint groups) and those that had identical rRNA gene restriction patterns (ribotypes). Both methods divided the isolates into 11 groups. In a few cases, horizontal gene transfer appeared to have distributed an identical altered PBP gene into different pneumococcal lineages. Eight isolates were indistinguishable by ribotyping or multilocus enzyme electrophoresis and contained identical PBP 1A genes. Although these isolates were therefore members of the same clone, they were divided into two fingerprint groups which contained different PBP 2X and 2B genes. Presumably, members of this clone have acquired different altered PBP 2X and 2B genes on two separate occasions. One of these fingerprint groups contained isolates of serotype 14, whereas the other contained isolates of both serotypes 14 and 7. The identification of isolates in the latter group that are identical by all criteria, except serotype, implies the occurrence of a change in serotype. The predominant serotypes of the penicillin-resistant pneumococci from Nairobi were serotypes 14 and 19. In both cases, isolates of the same serotype which required the same MIC of penicillin were not members of a single clone, indicating that identity of serotype and MIC are not sufficient criteria for defining clones of resistant pneumococci even when the bacteria are isolated from a single clinic.

DNA Fingerprinting to Examine Microgeographic Variation in theMagnaporthe grisea(Pyricularia grisea) Population in Two Rice Fields in Arkansas

DNA fingerprinting was used to analyze microgeographic variation in the Magnaporthe grisea population in Arkansas. One hundred and thirteen isolates were collected from two commercial rice fields (cv. Newbonnet) in 1991. In addition, several reference isolates representing the predominant pathotypes in Arkansas were examined. Total DNA of each isolate was restricted with EcoR1 and probed with a dispersed repeated MGR586 DNA probe. MGR586 DNA fingerprint groups were identified based on DNA restriction fragment length polymorphism similarities. Seven distinct fingerprint groups (designated A through G) were identified among the field and reference isolates. These seven fingerprint groups were very similar to seven of the eight fingerprint groups (lineages) previously reported for M. grisea [...]

Use of tRNA consensus primers to indicate subgroups of Pseudomonas solanacearum by polymerase chain reaction amplification.

Polymerase chain reaction amplification of DNA from 112 Pseudomonas solanacearum strains with the tRNA consensus primers T3A and T5A divided the species into three fingerprint groups. These groups correspond well with previous divisions made by restriction fragment length polymorphism analysis. This polymerase chain reaction test is a facile method for rapidly classifying P. solanacearum strains.

Untersuchungen an Taylorella equigenitalis: Zellwandproteine, Genomfingerprints, Plasmide, Adhäsion und Toxizität

In this study 55 strains of Taylorella equigenitalis isolated from horses of four different studs in Austria, and a comparative strain from the Federal Republic of Germany were investigated by different methods. These investigations were carried out with the help of SDS-PAGE, immunoblotting, the analyses of genomes and by proof of plasmids. Furthermore, pathogenic mechanisms such as adhesion or the formation of toxins were investigated in vitro. On the basis of the results carried out by means of SDS-PAGE and immunoblotting all tested strains of Taylorella equigenitalis were alike, whereas by DNA analyses the strains could be divided into five groups. The comparative strain from the FRG, which clearly differed from the Austrian strains, formed one group all by itself. From three studs, which are related to each other because of an intensive exchange of horses, representatives (n = 53) of three DNA fingerprint groups were isolated. These three fingerprint patterns were very similar to each other, while the hybridisation patterns from the other two Austrian strains were very different. One of these strains, isolated from a diseased mare, could not be distinguished from the other strain isolated from a clinical healthy stallion from the same study by this method. Only 47.3% from the investigated strains showed attachment to HeLa cells, while cell extracts of all of them caused morphological changes of a varying degree of both Y1 and Vero cells. There were no connexions between these adhesion-cytotoxicity-properties and the DNA fingerprint groups as well as the studs, respectively. No plasmids were found in the Taylorella equigenitalis strains used in this study.