Nucleotide diversity of Japanese isolates of infectious hematopoietic necrosis virus (IHNV) based on the glycoprotein gene

Infectious hematopoietic necrosis virus (IHNV), a member of the genus Novirhabdovirus, causes a highly lethal disease of salmonid fish. In the present study, G gene nucleotide sequences of 9 Japanese IHNV isolates obtained from 1971 to 1996 were analyzed to evaluate the genetic diversity and compared with IHNV isolates from North America and Europe. A radial phylogenetic tree revealed 5 major clusters including 3 genogroups (U, M and L) for North American isolates and 1 genogroup for European isolates. Five Japanese isolates from 1971 to 1982 appeared in the cluster for genogroup U, while the remaining Japanese isolates from 1980 to 1996 formed a new genogroup, JRt (Japanese rainbow trout). Maximum nucleotide diversity among the Japanese isolates was 4.5%, which was greater than that within the North American isolates (3.6%), and the degree of nucleotide diversity within Japanese isolates was increased by inclusion of the genogroup JRt isolates. It was concluded that Japanese isolates shared a common source with the genogroup U of the North American isolates and that there were large divergences between Japanese isolates before and after the 1980s.     

Use of RAPD-PCR fingerprinting to detect genetic diversity of soil Streptomyces isolates

Random amplified polymorphic DNA (RAPD) was used for identification and assessment of genetic diversity between isolates of Streptomyces from soil. Genomic DNA from 18 Streptomyces isolates and 2 reference strains were amplified using four different 10-mer primers. Different DNA fingerprinting patterns were obtained for all the isolates. Electrophoretic and cluster analysis of the amplification products revealed incidence of polymorphism among the isolates and none of them was identical to the reference strains although there were some common amplification bands. Two highly divergent groups were determined among the isolates. The results indicate that RAPD is an efficient method for discriminating and studying genetic diversity of Streptomyces isolates.     

Multilocus sequence typing of Campylobacter jejuni isolates from New South Wales, Australia

AIMS: Multilocus sequence typing (MLST) was used to examine the diversity and population structure of Campylobacter jejuni isolates associated with sporadic cases of gastroenteritis in Australia, and to compare these isolates with those from elsewhere. METHODS AND RESULTS: A total of 153 Camp. jejuni isolates were genotyped. Forty sequence types (STs) were found, 19 of which were previously undescribed and 21 identified in other countries. The 19 newly described STs accounted for 43% of isolates, 16 of which were assigned to known clonal complexes. Eighty-eight percent of isolates were assigned to a total of 15 clonal complexes. Of these, four clonal complexes accounted for 60% of isolates. Three STs accounted for nearly 40% of all isolates and appeared to be endemic, while 21 STs were represented by more than one isolate. Seven infections were acquired during international travel, and the associated isolates all had different STs, three of which were exclusive to the travel-acquired cases. Comparison of serotypes among isolates from clonal complexes revealed further diversity. Eight serotypes were identified among isolates from more than one clonal complex, while isolates from six clonal complexes displayed serotypes not previously associated with those clonal complexes. CONCLUSIONS: Multilocus sequence typing is a useful tool for the discrimination of subtypes and examination of the population structure of Camp. jejuni associated with sporadic infections. SIGNIFICANCE AND IMPACT OF THE STUDY: This study highlights the genotypic diversity of Camp. jejuni in Australia, demonstrating that STs causing disease have both a global and a local distribution evident from the typing of domestically and internationally acquired Camp. jejuni isolates.     

Genetic diversity analysis of a world-wide collection of <Emphasis Type="Italic">Ascochyta rabiei</Emphasis> isolates using sequence tagged microsatellite markers

A previously characterized compound microsatellite locus ARMS1, containing penta- and decameric repeat units, has been reported to reveal genetic diversity in Ascochyta rabiei (Pass.) Labr. isolates. Therefore, 37 isolates of Ascochyta rabiei collected from different states of India and 38 isolates from fifteen other countries in the world were examined for their diversity at this locus. Twenty-six alleles on the basis of size (228--451 base pairs) were detected in the world isolates examined, while 15 alleles (287--418 base pairs) were observed in isolates from the Indian subcontinent. To the best of our knowledge, this study is the first to demonstrate diversity in representative Ascochyta rabiei isolates from different parts of the world at the ARMS1 locus.     

Molecular assessment of diversity among endophytic diazotrophs isolated from subtropical Indian sugarcane

Twenty-two endophytic bacterial isolates from the roots of sugarcane were compared morphologically, biochemically and genetically. Gram staining, colony pigment, texture and other cultural characteristics were taken for morphological characterization. Oxidation-fermentation tests for D-glucose and D-sucrose, production of acid and hydrogen from different carbon source, oxidase activity, antibiotic and drug resistance patterns were chosen as the biochemical and physiological criteria. Twelve random decamer primers were used to analyze and compare these isolates through RAPD among themselves as well as with known standard diazotrophic strains. The isolates were compared through dendrograms constructed on the basis of similarity patterns obtained from biochemical and RAPD analysis. The estimated diversity through RAPD analysis was more evident than the diversity on the basis of morphological and biochemical characters. Within Acetobacter group, the isolates showed substantial genetic diversity for future exploitation as PGPRs and diazotrophic associative endophytes.     

Genetic diversity among clonal isolates of Karenia brevis as measured with microsatellite markers

Karenia brevis is the major harmful bloom-forming dinoflagellate in the Gulf of Mexico yet little is known about the intraspecific genetic diversity of this species. Here we describe nine new microsatellite markers and, combined with nine previously described microsatellites, use them to genotype 40 cultured isolates of K. brevis. Genetic diversity identified from cultured isolates was compared with the genetic diversity identified from two field samples to assess how well the current cultures represent the field population. Thirty-nine unique haplotypes were identified from 40 cultured isolates of K. brevis using 18 microsatellite markers. Genetic diversity was similar between cultured isolates and the two field samples. The success of 18 microsatellite markers to distinguish individual isolates supports the use of microsatellites as a genetic tool for diagnostic identification of cultured isolates of K. brevis.     

Genetic Diversity of Rice False Smut Fungus, Ustilaginoidea virens and its Pronounced Differentiation of Populations in North China

Rice false smut caused by Ustilaginoidea virens is an important constraint affecting rice yield and quality in Asia. In China, rice false smut is especially severe in the japonica rice-growing areas in the North China. Nothing is known neither about the diversity of the pathogen in this region nor about the characteristic of its local population. In this study, 110 U. virens isolates sampled from Liaoning and Beijing of North China were analysed using amplified fragment length polymorphism (AFLP) markers to primarily understand the genetic diversity of this pathogen and its population characteristics. At the genetic distance of 0.32, all isolates were divided into two groups. Group A consists of two subgroups differentiated at the genetic distance of 0.55, subgroup 1 included all isolates from Liaoning with an average genetic similarity over 0.82 and subgroup 2 included 27 isolates from Beijing with similarity of 0.74. Group B consists of 28 isolates from Beijing having a diversity of 0.054. The isolates from the Liaoning province, where rice false smut has generated different take-all epidemic for 20 years, showed a genetic diversity of 0.305, which was approximately equally distributed within and among populations. Whereas genetic diversity was 0.458 among isolates from Beijing, an extremely high level of genetic differentiation among 55 isolates was observed in this disease hotspot. Our results suggested that the populations among different locations where sexual stage of the pathogen was rare to be found within ecological region were similar, and the variation of this pathogen has mainly arisen via asexual mechanisms. The migration through human activities in breeding perhaps provides a means of transporting the pathogen from one region to another.     

Genetic and Phenotypic Diversity among Botrytis cinerea Isolates in Iran

Forty-four Botrytis cinerea isolates from different hosts and geographical regions were studied for colony morphology, mycelial growth rate at different temperatures, pathogenicity and molecular diversity. Botrytis cinerea isolates had temperature optima of 20–25°C and isolates showed variation in growth rate at different temperatures. Two morphological types were identified among tested isolates: mycelial and sclerotial. The pathogenicity of isolates was tested on grapevine leaves, and it was revealed that nine of 44 isolates were non-pathogenic and among them seven were of mycelial type. There was no correlation between mycelium growth rate and pathogenicity. Genetic diversity was investigated using nine arbitrary decaprimers. No relationship was found between molecular clusters and geographical region or sampling time; whereas isolates from the same plant host tended to cluster with each other. Seven of nine non-pathogenic isolates were separated from pathogenic ones.     

Diversity of Toxic Shock Syndrome Toxin 1-Positive Staphylococcus aureus Isolates

Staphylococcus aureus isolates from women with nasal, anal, or vaginal colonization were evaluated for population diversity by pulsed-field gel electrophoresis. Cluster analysis of restriction patterns revealed diversity indices of 0.89 and 0.99 for toxic shock syndrome toxin 1-positive and -negative isolates, respectively. Toxin-producing strains were isolated more frequently from the nares than from other sites.     

Genetic diversity in Monilinia laxa populations in stone fruit species in Hungary

The objectives of this study were firstly, to determine the genetic diversity of Monilinia laxa isolates from Hungary, using the PCR-based inter-simple sequence repeat (ISSR) and randomly amplified polymorphic DNA (RAPD) technique; secondly, to prepare genetic diversity groups based on the dendrograms; and finally, to select some relevant isolates to study their fungicide sensitivity. 55 and 77 random amplified polymorphic ISSR and RAPD markers, of which 23 and 18 were polymorphic and 32 and 59 monomorphic, respectively, were used to assess the genetic diversity and to study the structure of M. laxa populations in Hungary. 27 isolates out of 57 ones were confirmed as M. laxa from several orchards (subpopulations) in three geographical regions, in various inoculum sources and in various hosts, were used. 10 fungicides and 12 isolates selected from genetic diversity groups based on the ISSR dendrograms were used to determine the fungicide sensitivity of the selected isolates. The analysis of population structure revealed that genetic diversity within locations, inoculum sources and host (H _S ) accounted for 99 % of the total genetic diversity (H _T ), while genetic diversity among locations, inoculum sources and host represented only 1 %. The relative magnitude of gene differentiation between subpopulations (G _ST ) and the estimate of the number of migrants per generation (Nm) averaged 0.005–0.009 and 53.9–99.2, respectively, for both ISSR and RAPD data set. The results obtained in dendrograms were in accordance with the gene diversity analysis. Grouping of isolates in the dendrograms was irrespective of whether they came from the same or different geographical locations. There was no relationship between clustering among isolates from inoculum sources and hosts. In the fungicide sensitivity tests, five isolates out of 12 were partly insensitive to boscalid+piraclostrobin, cyprodinil, fenhexamid or prochloraz. Obtained results in genetic diversity of M. laxa populations are discussed together with implications for the management of brown rot.     

Diversity of nitrogen-fixing cyanobacteria under various ecosystems of Thailand: I. Morphology,physiology and genetic diversity

The diversity among 853 isolates of nitrogen-fixing cyanobacteria obtained from soil samples collected from different ecosystems including mountainous, forest and cultivated areas in the central, northern and northeastern regions of Thailand was examined. Most isolates showed slow growth rate and had filamentous, heterocystous cells. The percentage of heterocysts in the filaments of different isolates varied from 8.3 to 9.6. Only a few strains showed high nitrogen-fixing potential, while most of the strains exhibited low capacity for nitrogen fixation. Anabaena and Nostoc were the dominant genera among these isolates. One hundred and two isolates were randomly selected from this diverse collection to determine the extent of genetic diversity on the basis of DNA fingerprinting using the PCR method. Based on the PCR products obtained by using a combination of three primers, all strains could be distinguished from one another. When a subset of 45 isolates of Nostoc and a subset of 44 isolates of Anabaena were further analysed by PCR, a wide range of diversity was observed within each of these genera.     

Characterization of Fusarium oxysporum Isolates from Common Bean and Sugar Beet Using Pathogenicity Assays and Random-amplified Polymorphic DNA Markers

Fusarium wilt is an economically important fungal disease of common bean and sugar beet in the Central High Plains (CHP) region of the USA, with yield losses approaching 30% under appropriate environmental conditions. The objective of this study was to characterize genetic diversity and pathogenicity of isolates of Fusarium oxysporum obtained from common bean and sugar beet plants in the CHP that exhibited Fusarium wilt symptoms. A total of 166 isolates of F. oxysporum isolated from diseased common bean plants were screened for pathogenicity on the universal susceptible common bean cultivar ‘UI 114’. Only four of 166 isolates were pathogenic and were designated F. oxysporum f.sp. phaseoli (Fop). A set of 34 isolates, including pathogenic Fop, F. oxysporum f.sp. betae (Fob) isolates pathogenic on sugar beet, and non‐pathogenic (Fo) isolates, were selected for random‐amplified polymorphic DNA (RAPD) analysis. A total of 12 RAPD primers, which generated 105 polymorphic bands, were used to construct an unweighted paired group method with arithmetic averages dendrogram based on Jaccard's coefficient of similarity. All CHP Fop isolates had identical RAPD banding patterns, suggesting low genetic diversity for Fop in this region. CHP Fob isolates showed a greater degree of diversity, but in general clustered together in a grouping distinct from Fop isolates. As RAPD markers revealed such a high level of genetic diversity across all isolates examined, we conclude that RAPD markers had only limited usefulness in correlating pathogenicity among the isolates and races in this study.     

The structure and interrelationship of fungal populations in native and cultivated soils

Mitochondrial DNA haplotypes have been characterized for 120 isolates of the asexual fungus Fusarium oxysporum. Sixty of these isolates were obtained from soil in a native grassland in the San Joaquin Valley of California, including 20 isolates from each of six different sampling locations. The same sampling strategy was used to obtain 60 additional isolates from an agricultural field of the same soil type directly adjacent to the native soil. Twenty-three different mitochondrial DNA haplotypes were identified among the 120 isolates, including 11 haplotypes represented by two or more isolates and 12 that were unique. The five most common mitochondrial DNA haplotypes accounted for 93 (78%) of the 120 isolates. Isolates representing each of these five mitochondrial DNA haplotypes were found both in the cultivated and in the native soil. Seventy-two per cent of the isolates found in the cultivated soil were associated with the same mitochondrial DNA haplotype as one or more isolates in the native soil. The remaining isolates in the cultivated soil were associated with comparatively rare mitochondrial DNA haplotypes, most of which showed a close relationship to one of the haplotypes found in the native soil. Hierarchial gene diversity analysis indicated that a significant proportion of the mitochondrial DNA haplotype diversity was attributable to differences between sampling sites in the native soil but not in the cultivated soil. This may reflect significant spatial structuring of genetic diversity in populations of F. oxysporum in a native soil. The proportion of mtDNA haplotype diversity attributable to differences between populations in the native and cultivated soils was not significant. This suggests that our entire collection, encompassing strains from both native and cultivated soils, is representative of a single population of F. oxysporum.     

Entamoeba histolytica: genetic diversity of clinical isolates from Bangladesh as demonstrated by polymorphisms in the serine-rich gene.

The varied organ tropisms and clinical presentations of infection by Entamoeba histolytica have stimulated interest in the role of parasite genetic diversity in virulence. We investigated genetic diversity among 54 E. histolytica isolates from Bangladesh by analyzing polymorphism in the serine-rich gene by nested PCR on DNA extracted from stool and liver aspirate pus. We detected both size and restriction site polymorphisms among the isolates within this endemic area. A combination of the nested PCR results and the AluI digestion of the PCR products examined yielded 25 distinct DNA banding patterns among the 42 stool isolates and an additional 9 distinct patterns among the 12 liver abscess isolates. Approximately half of the isolates had unique polymorphisms. Interestingly, the majority of E. histolytica from the liver had polymorphisms which were not present in intestinal isolates from the same geographic area. These data are consistent with the existence of genetic differences between E. histolytica which cause intestinal and those which cause hepatic disease. We conclude that there is genetic diversity within E. histolytica isolates from an endemic population as reflected in serine-rich E. histolytica protein gene polymorphism. The correlation of genetic differences with the pathogenic potential of E. histolytica strains and the implications of genetic diversity for the immunoprophylaxis of amebiasis require further study.     

Hammondia heydorni: evidence of genetic diversity among isolates from dogs

Canine isolates of Hammondia heydorni from Argentina, Brazil, and the United States were analysed for genetic diversity. A total of 14 isolates were tested for their ability to produce amplification using three PCR assays, one targeting the common toxoplasmatiid ITS-1 region and 2 amplifying novel, H. heydorni-specific loci, HhAP7 and HhAP10. While the ITS-1 fragments could be amplified from all isolates, only six isolates were capable of amplifying the fragments from the novel loci. The PCR products were further investigated for genetic diversity using restriction fragment length polymorphism (RFLP) and single strand conformation polymorphism (SSCP) techniques. Polymorphism in the digestion pattern was evident only at the HhAP10 locus, differentiating two of the Argentinean isolates from the remainder. Mobility shifts on SSCP gels revealed that the two Argentinean isolates were not only different from the other four isolates, but also differed from each other, both at the HhAP7 and HhAP10 loci. The ITS-1 fragments of all isolates were identical by RFLP. However, two distinct mobility patterns resulted when the products were electrophoresed on SSCP gels. Based on the sequence data from the ITS-1 and the two random loci, the isolates could be broadly classified into two distinct groups, within which minor polymorphisms were evident. In contrast, very little heterogeneity occurred in the sequences of corresponding ITS-1 regions of Neospora caninum and Toxoplasma gondii isolates. Thus, it is concluded that there is a considerable degree of microheterogeneity among isolates of H. heydorni. This diversity should be taken into consideration while attempting to elucidate the systematics, diagnostics, and biology of H. heydorni in relation to N. caninum.     

中国产禾本科植物内生真菌的遗传多样性及rDNA-ITS系统发育分析

采用RAPD技术对分离自我国的13株Epichlo?spp.和9株Neotyphodiumspp.的禾本科植物内生真菌进行了遗传多样性分析,同时对其中的4株进行了rDNA-ITS序列分析及系统发育研究。RAPD分析结果显示:原产自我国的菌株与原产自欧洲的菌株N.uncinatum之间亲缘关系较远;我国的21个菌株之间也存在一定的遗传多样性。rDNA-ITS序列分析表明:原产自我国的菌株聚为一枝,表明我国的Neotyphodium属真菌很有可能直接由我国的Epichlo?属真菌演化而来的新的类群;我国的Epichlo?属真菌有可能和国外推测的一个未确定的Neotyphodium属真菌的杂交进化起源(LAC)有关。     

法国蜜环菌Armillaria gallica菌株遗传多样性的ISSR分析

在中国东北地区共采集到53个法国蜜环菌Armillaria gallica菌株,用ISSR（Inter-Simple Sequence Repeat）标记技术对这些菌株进行遗传多样性分析。用6个ISSR引物扩增所得条带表明,ISSR标记在蜜环菌中存在较高的多态性;亲缘关系树状图表明,有3个菌株遗传分化明显;其余50个分别来自3个不同地理居群的菌株聚成一类,亲缘关系较近,没有表现出地理隔离。     

Genetic and plasmid diversity within natural populations of Pseudomonas syringae with various exposures to copper and streptomycin bactericides.

We examined the genetic and plasmid diversity within natural populations of Pseudomonas syringae isolated from three ornamental pear nurseries in eastern Oklahoma. The bactericide spray regimen differed at each nursery; copper and streptomycin, only copper, and no bactericides were applied at nurseries I, II, and III respectively. Resistance to copper (Cur) and resistance to streptomycin (Smr) were determined for 1,938 isolates of P. syringae; isolates from nurseries I and II were generally Cur Sms; whereas most isolates from nursery III were Cus Sms. The plasmid profiles of 362 isolates were determined, and six, one, seven, and four plasmid profiles were obtained for Cur, Smr, Cur Smr, and Cus Sms isolates, respectively. All Smr plasmids contained sequences homologous to the strA and strB Smr genes from broad-host-range plasmid RSF1010 and were associated with Smr transposon Tn5393. Plasmids were placed into two groups on the basis of hybridization to the oriV and par sequences from pOSU900, a cryptic plasmid in P. syringae pv. syringae. A total of 100 randomly chosen P. syringae isolates from nurseries I and III were analyzed for genetic diversity by using the arbitrarily primed PCR (AP-PCR) technique. An analysis of chromosomal genotypes by AP-PCR revealed a high degree of genetic diversity among the isolates, and the results of this analysis indicated that the isolates could be clustered into two distinct groups. The plasmid profiles were specific to isolates belonging to particular AP-PCR groups. Within each AP-PCR group, identical plasmid profiles were produced by isolates that had different chromosomal genotypes, implying that plasmid transfer has played an important role in the dissemination of Cur and Smr within the populations studied.     

Genetic diversity of Acanthamoeba isolated from ocean sediments

Genetic diversity of 18 Acanthamoeba isolates from ocean sediments was evaluated by comparing mitochondrial (mt) DNA RFLP, 18S rDNA sequences and by examining their cytopathic effects on human corneal epithelial cells versus reference strains. All isolates belonged to morphologic group II. Total of 16 restriction phenotypes of mtDNA from 18 isolates demonstrated the genetic diversity of Acanthamoeba in ocean sediments. Phylogenetic analysis using 18s rDNA sequences revealed that the 18 isolates were distinct from morphological groups I and III. Fifteen isolates showed close relatedness with 17 clinical isolates and A. castellanii Castellani and formed a lineage equivalent to T4 genotype of Byers group. Two reference strains from ocean sediment, A. hatchetti BH-2 and A. griffini S-7 clustered unequivocally with these 15 isolates. Diversity among isolates was also evident from their cytopathic effects on human corneal cells. This is the first time describing Acanthamoeba diversity in ocean sediments in Korea.     

Use of ribotyping and random amplified polymorphic DNA to differentiate isolates of Burkholderia andropogonis

AIMS: The aim of this study was to determine the genetic diversity among isolates of Burkholderia andropogonis from various host plant species and geographic locations. METHODS AND RESULTS: Both random amplified polymorphic DNA (RAPD) and ribotyping analyses were used to assess the diversity of B. andropogonis isolates and compare these results with pathogenicity assays carried out on a number of common hosts of the organism. CONCLUSIONS: Both RAPD and ribotyping analyses revealed a high level of genetic diversity between isolates of B. andropogonis. Both methods demonstrated a similar clustering of isolates. However, there was no strict correlation between the genetic diversity revealed and the original host, geographic location or pathogenicity of the isolates. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first report on the genetic diversity of isolates of B. andropogonis. The great degree of diversity revealed in this study contrasts with the lack of phenotypic diversity within this species.