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Spacer Regions (spacer + regions)
Kinds of Spacer Regions Selected AbstractsDetection and typing of Borrelia burgdorferi sensu lato genospecies in Ixodes persulcatus ticks in West Siberia, RussiaFEMS MICROBIOLOGY LETTERS, Issue 2 2003Anatoly B Beklemishev Abstract The prevalence of Borrelia burgdorferi sensu lato (s.l.) genospecies in West Siberia as well as in many other regions of Russia remains insufficiently investigated. In the present study a total of 151 adult female ticks Ixodes persulcatus Schulze, collected at three localities in eastern regions of West Siberia, where Lyme disease is endemic, were examined for the presence of the spirochete B. burgdorferi s.l. by polymerase chain reaction targeting the 23S,5S rRNA intergenic spacer regions. Spirochetal DNA was detected in on average 15.2±3.0% of the ticks examined. The infection rate of adult ticks with B. burgdorferi s.l. at various localities ranged from 8.6±3.4% to 29.0±7.6%, being greatest in the northernmost site studied and decreasing southwards. The restriction patterns obtained after MseI digestion of the 23S,5S rRNA intergenic spacer amplicons assigned 23 DNA samples to the following genomic groups: 19 to B. garinii (12 to group NT29 and seven to group 20047T), three to B. afzelii, and one to mixed B. afzelii and B. garinii NT29. We have not detected other genospecies, which were found in ticks in Europe, the Russian Far East and Japan. Thus, the ticks examined were associated only with two genospecies of Borrelia burgdorferi s.l. pathogenic to humans (B. garinii and B. afzelii), and B. garinii was the major genospecies infecting adult I. persulcatus in eastern regions of West Siberia. [source] Genetic diversity of the hyperparasite Sphaerellopsis filum on Melampsora willow rustsFOREST PATHOLOGY, Issue 5 2004M. Liesebach Summary The non-specific rust hyperparasite Sphaerellopsis filum occurs naturally on Melampsora rusts of many species of the genus Salix as well as on a large range of other rust genera worldwide. To study the genetic diversity of the hyperparasitic fungus 77 S. filum isolates collected from rusts on willow clones from plantations, clone collections and natural habitats of different sites were investigated using polymerase chain reaction - restriction fragment length polymorphism (PCR-RFLP) analysis of the rDNA internal transcribed spacer regions including 5.8S rDNA and sequence analysis. Additionally, strains from Melampsora poplar rusts (4) and strains of Puccinia abrupta from Parthenium hysterophorus (5) and of P. obscura from Bellis perennis (1) were used for comparisons. Results of genetic analysis demonstrated distinct variation within the S. filum isolates. Two main groups with more than 32% difference between their nucleotide sequences were distinguished, indicating two taxa within S. filum. Within the first main group three profiles (I, II, III) were detected. The differences between these profiles were about 12%. The variation within each profile was very low (less than 2%). The second main group comprised two profiles (IV, V), which differed in 12 to 16% of their nucleotide positions. The isolates of group IV possessed a higher variation (up to 5%) within the group than those of the first main group (I, II, III). Group V was only represented by a single isolate. Neither interrelations between the S. filum profiles and the Melampsora genotypes nor a spatial distribution could be detected. It is remarkable that the six strains of S. filum from Puccinia rusts belong to one subgroup. Résumé Sphaerellopsis filum est un hyperparasite non spécifique des rouilles qui se rencontre naturellement sur les Melampsora affectant le genre Salix ainsi que sur une large gamme d'autres genres de rouille dans le monde entier. Pour caractériser la diversité de ce champignon hyperparasite, 77 isolats de S. filum, récoltés à partir de lésions de rouille sur des clones de plantation, des collections de clones et dans des sites naturels, ont étéétudiés par analyse PCR-RFLP des régions ITS de l'ADNr, incluant le 5.8S rDNA, et par analyse de séquences. Des souches provenant de Melampsora des peupliers (4), de Puccinia abrupta sur Parthenium hysterophorus (5) et de P. obscura sur Bellis perennis (1) ont également été utilisées pour comparaison. L'analyse génétique démontre une variation entre isolats de S. filum. Deux groupes principaux, avec plus de 32% de différence dans leur séquence nucléotidique, se distinguent, indiquant l'existence de deux taxons au sein de S. filum. A l'intérieur du premier groupe, trois profils (I, II, III) sont mis en évidence, avec une différence d'environ 12% entre profils mais très peu de variation (moins de 2%) à l'intérieur d'un profil. Le deuxième groupe comprend deux profils (IV, V) qui différent de 12 à 16% pour leur séquence nucléotidique. Les isolats du groupe IV présentent une variation intra-groupe plus importante (jusqu'à 5%) que ceux des groupes I, II et III. Le groupe V n'est représenté que par un isolat. Aucune relation n'a pu être établié entre les profils de S. filum et les génotypes de Melampsora ou la distribution spatiale. Il est à noter que les six isolats de S. filum provenant de rouilles àPuccinia appartiennent à un seul sous-groupe. Zusammenfassung Der Hyperparasit Sphaerellopsis filum kommt natürlich auf Melampsora -Rostpilzen bei Salicaceae und auf zahlreichen anderen Rostgattungen weltweit vor. Um die genetische Vielfalt dieses hyperparasitischen Pilzes zu untersuchen, wurden 77 S. filum -Proben, die von Weidenrosten in Plantagen, Klonsammlungen und von verschiedenen natürlichen Standorten stammen, isoliert. Die 5.8S-ITS-Abschnitte der rDNA wurden mit Hilfe der PCR-RFLP untersucht und Sequenzen analysiert. Zum Vergleich wurden vier Isolate von Pappelrosten sowie fünf Isolate von Puccinia abrupta von Parthenium hysterophorus und ein Isolat von Puccinia obscura von Bellis perennis herangezogen. Die Ergebnisse der genetischen Untersuchungen zeigten eine deutliche Variation zwischen den S. filum -Isolaten. Zwei Gruppen mit über 32% Unterschied in der Nukleotid-Sequenz ließen sich unterscheiden. Dies deutet auf zwei taxonomische Einheiten von Sphaerellopsis hin. Die erste Gruppe ließ sich in drei Untergruppen (I, II, III) einteilen, deren 5.8S-ITS-Profile im Mittel 12% Unterschied aufwiesen. Die Variation innerhalb dieser drei Profile war sehr gering (,2%). Die zweite Gruppe umfasste zwei Profile (IV, V), die sich an 12 bis 16% Positionen ihrer Nukleotid-Abfolge unterschieden. Die Variation innerhalb von Profil IV war höher (bis 5%) als die der Untergruppen I - III, Profil V war nur durch ein Isolat vertreten. Eine Beziehung des S. filum -Genotyps zum Melampsora -Genotyp der Weide oder der Pappel ließ sich bei dem untersuchten Material nicht nachweisen, ebenso konnte keine geographische Differenzierung gefunden werden. Auffällig ist, dass alle sechs Puccinia -Isolate einer Untergruppe angehörten. [source] Analysis of intraspecies polymorphism in the ribosomal DNA cluster of the cockroach Blattella germanicaINSECT MOLECULAR BIOLOGY, Issue 2 2000D. V. Mukha Abstract HindIII restriction digests of the rDNA repeat unit of the German cockroach, Blattella germanica, reveal significant intraspecies sequence polymorphism. This variability is probably caused by structural differences within the nontranscribed spacer regions (NTS) of the ribosomal repeat unit. HindIII rDNA fragment polymorphisms in three cockroach strains show that individuals from different populations may have different HindIII rDNA patterns, whereas individuals within populations exhibit relatively similar rDNA patterns. We suggest that HindIII restriction fragment polymorphisms within cockroach ribosomal DNA will be a valuable tool for measuring population-level parameters within and between natural cockroach populations. [source] Oligonucleotide microarrays for the detection and identification of viable beer spoilage bacteriaJOURNAL OF APPLIED MICROBIOLOGY, Issue 4 2008D.G. Weber Abstract Aims:, The design and evaluation of an oligonucleotide microarray in order to detect and identify viable bacterial species that play a significant role in beer spoilage. These belong to the species of the genera Lactobacillus, Megasphaera, Pediococcus and Pectinatus. Methods and Results:, Oligonucleotide probes specific to beer spoilage bacteria were designed. In order to detect viable bacteria, the probes were designed to target the intergenic spacer regions (ISR) between 16S and 23S rRNA. Prior to hybridization the ISR were amplified by combining reverse transcriptase and polymerase chain reactions using a designed consenus primer. The developed oligonucleotide microarrays allows the detection of viable beer spoilage bacteria. Conclusions:, This method allows the detection and discrimination of single bacterial species in a sample containing complex microbial community. Furthermore, microarrays using oligonucleotide probes targeting the ISR allow the distinction between viable bacteria with the potential to grow and non growing bacteria. Significance and Impact of the Study:, The results demonstrate the feasibility of oligonucleotide microarrays as a contamination control in food industry for the detection and identification of spoilage micro-organisms within a mixed population. [source] Clustering of Saccharomyces boulardii strains within the species S. cerevisiae using molecular typing techniquesJOURNAL OF APPLIED MICROBIOLOGY, Issue 4 2002G. Mitterdorfer Aims: This study was undertaken to characterize and differentiate therapeutically relevant Saccharomyces yeasts. Among the isolates were so-called Saccharomyces boulardii strains, which are considered as probiotic agents, but whose taxonomic assignment is controversial. Moreover, the discriminative power of the applied molecular typing techniques should be evaluated. Methods and Results: Genotyping was performed using species-specific polymerase chain reaction (PCR), randomly amplified polymorphic DNA-PCR, restriction fragment length polymorphism analysis of rDNA spacer regions and pulsed-field gel electrophoresis. Species-specific PCR assigned all of the product isolates to the species S. cerevisiae. By combining the other techniques, all isolates could be discriminated. Moreover, it could be demonstrated that probiotic S. boulardii strains form a separate cluster located within the species. Conclusions: With the exception of species-specific PCR, all of the applied methodologies were suitable for subspecies typing and indicated a close relationship between the probiotic strains. Significance and Impact of the Study: The methods applied in this study are considered powerful tools for quality control of therapeutically relevant yeasts. It is of crucial importance, especially regarding S. boulardii yeasts, to verify the identity of the correct strain, since the beneficial properties are considered to be strain-specific. [source] GENETIC DIVERSITY AND INTROGRESSION IN TWO CULTIVATED SPECIES (PORPHYRA YEZOENSIS AND PORPHYRA TENERA) AND CLOSELY RELATED WILD SPECIES OF PORPHYRA (BANGIALES, RHODOPHYTA),JOURNAL OF PHYCOLOGY, Issue 2 2009Kyosuke Niwa We investigated the genetic variations of the samples that were tentatively identified as two cultivated Porphyra species (Porphyra yezoensis Ueda and Porphyra tenera Kjellm.) from various natural populations in Japan using molecular analyses of plastid and nuclear DNA. From PCR-RFLP analyses using nuclear internal transcribed spacer (ITS) rDNA and plastid RUBISCO spacer regions and phylogenetic analyses using plastid rbcL and nuclear ITS-1 rDNA sequences, our samples from natural populations of P. yezoensis and P. tenera showed remarkably higher genetic variations than found in strains that are currently used for cultivation. In addition, it is inferred that our samples contain four wild Porphyra species, and that three of the four species, containing Porphyra kinositae, are closely related to cultivated Porphyra species. Furthermore, our PCR-RFLP and molecular phylogenetic analyses using both the nuclear and plastid DNA demonstrated the occurrence of plastid introgression from P. yezoensis to P. tenera and suggested the possibility of plastid introgression from cultivated P. yezoensis to wild P. yezoensis. These results imply the importance of collecting and establishing more strains of cultivated Porphyra species and related wild species from natural populations as genetic resources for further improvement of cultivated Porphyra strains. [source] Occurrence of Toxic Hexadepsipeptides in Preharvest Maize Ear Rot Infected by Fusarium poae in PolandJOURNAL OF PHYTOPATHOLOGY, Issue 1 2007J. Che, kowski Abstract Twenty-seven preharvest maize ears affected by Fusarium poae rot (disease score 36,100%) were selected in 1998 and 1999 in Poland and examined for the occurrence of toxic hexadepsipeptides: beauvericin (BEA), enniatin A, enniatin B and enniatin B1. The identification of F. poae was confirmed by sequence analysis of variable internal transcribed spacer regions and compared with NCBI gene bank DNA sequences. Chemical analyses were performed by HPLC-MS. In 27 ears infected by F. poae were detected: BEA (trace to 46 ,g/g) in 18 samples, enniatin A (trace to 37 ,g/g) in nine samples, enniatin B (trace to 47 ,g/g) in 15 samples and enniatin B1 (trace to 25 ,g/g) in 12 samples. When 20 strains of F. poae isolated from these samples were cultured on rice, all produced BEA (1.9,75 ,g/g), three enniatin A (1.8,2 ,g/g), 12 enniatin B (1.1,5.1 ,g/g) and eight enniatin B1 (1.2,5.2 ,g/g). Occurrence and quantification of enniatin A, enniatin B and enniatin B1 and their co-occurrence with BEA in maize kernels is reported for the first time. [source] Evaluation of the PCR method for identification of Bifidobacterium speciesLETTERS IN APPLIED MICROBIOLOGY, Issue 1 2008S.Y. Youn Abstract Aims:,Bifidobacterium species are known for their beneficial effects on health and their wide use as probiotics. Although various polymerase chain reaction (PCR) methods for the identification of Bifidobacterium species have been published, the reliability of these methods remains open to question. Methods and Results:, In this study, we evaluated 37 previously reported PCR primer sets designed to amplify 16S rDNA, 23S rDNA, intergenic spacer regions, or repetitive DNA sequences of various Bifidobacterium species. Conclusions:, Ten of 37 experimental primer sets showed specificity for B. adolescentis, B. angulatum, B. pseudocatenulatum, B. breve, B. bifidum, B. longum, B. longum biovar infantis and B. dentium. Significance and Impact of the Study:, The results suggest that published Bifidobacterium primer sets should be re-evaluated for both reproducibility and specificity for the identification of Bifidobacterium species using PCR. Improvement of existing PCR methods will be needed to facilitate identification of other Bifidobacterium strains, such as B. animalis, B. catenulatum, B. thermophilum and B. subtile. [source] Survival of inoculated Saccharomyces cerevisiae strain on wine grapes during two vintagesLETTERS IN APPLIED MICROBIOLOGY, Issue 3 2006F. Comitini Abstract Aims:, To investigate the influence of a specific ecological niche, the wine grape, on the survival and development of Saccharomyces cerevisiae. Methods and Results:, A strain with a rare phenotype was sprayed onto the grape surfaces and monitored through two vintages using a specific indicative medium and analysing the internal transcribed spacer regions in the 5·8S rDNA. During the ripening process, there was a progressive colonization of the surface of the undamaged and damaged grapes by epiphytic yeasts, up to the time of harvest. The damaged wine grapes showed a much greater epiphytic yeast population. However, the inoculated S. cerevisiae strain showed a scarce persistence on both undamaged and damaged wine grapes, and the damaged grapes did not appear to improve the grape surface colonization of this strain. Conclusions:, Results indicated that wine grape is not a favourable ecological niche for the development and colonization of S. cerevisiae species. Significance and Impact of the Study:, Results of this work are further evidence that S. cerevisiae is not specifically associated with natural environments such as damaged and undamaged wine grapes. [source] Variation in 16S-23S rRNA intergenic spacer regions among Bacillus subtilis 168 isolatesMOLECULAR MICROBIOLOGY, Issue 1 2001Yevette J. Shaver The genome of the Bacillus subtilis 168-type strain contains 10 ribosomal RNA (rRNA) operons. In the intergenic spacer region (ISR) between the 16S and 23S rRNA genes, five rRNA operons, rrnI-H-G and rrnJ-W, lack a trinucleotide signature region. Precise determination of molecular weight (MW), using electrospray mass spectrometry (MS), of the polymerase chain reaction (PCR) products from a segment of the ISR from the 168-type strain and B. subtilis 168-like strain 23071 demonstrated 114 and 111 basepair (bp) PCR products (due to the presence or absence of the insert in the operons) as predicted from sequence. However, PCR of the ISR segment for five other B. subtilis 168 isolates generated only a 114 bp PCR product, suggesting the presence of the trinucleotide signature region in all rRNA operons for these strains. Additional genetic variability between the seven B. subtilis 168 isolates was demonstrated by restriction fragment length polymorphism (RFLP) of the rRNA operons, with three distinct patterns found upon Southern blot analysis. The 168-type strain and three others (23066, 23067, and 23071) exhibited the same Southern pattern. Thus, operon deletion is not responsible for the absence of a 111 bp product on MS analysis for strains 23066 and 23067. Restriction analysis confirmed the presence of the trinucleotide signature region in the ISR of all rRNA operons for five B. subtilis 168 isolates; sequencing of rrnW/H from a representative strain also upheld this finding. These results help provide a better understanding of variations in sequence, operon number and chromosomal organization, both within a genome and among isolates of B. subtilis subgroup 168. It is also hypothesized that the presence of the trinucleotide insert in certain rRNA operons may play a role in rRNA maturation and protein synthesis. [source] Multiple determinants influence root colonization and induction of induced systemic resistance by Pseudomonas chlororaphis O6MOLECULAR PLANT PATHOLOGY, Issue 6 2006SONG HEE HAN SUMMARY Colonization of the roots of tobacco by Pseudomonas chlororaphis O6 induces systemic resistance to the soft-rot pathogen, Erwinia carotovora ssp. carotovara SCC1. A screen of the transposon mutants of P. chlororaphis O6 showed mutants with about a fivefold reduction in ability to induce systemic resistance to the soft-rot disease. These mutations disrupted genes involved in diverse functions: a methyl-accepting chemotaxis protein, biosynthesis of purines, phospholipase C, transport of branched-chain amino acids and an ABC transporter. Additional mutations were detected in the intergenic spacer regions between genes encoding a GGDEF protein and fumarate dehydratase, and in genes of unknown function. The mutants in the ABC transporters did not display reduced root colonization. However, the other mutants had up to 100-fold reduced colonization levels. Generally the production of metabolites important for interactions in the rhizosphere, phenazines and siderophores, was not altered by the mutations. A reduced induction of systemic resistance by a purine biosynthesis mutant with a disrupted purM gene correlated with poor growth rate, lesser production of phenazines and siderophore and low levels of root colonization. These studies showed that multiple determinants are involved in the induction of systemic resistance, with there being a requirement for strong root colonization. [source] Identification of Candida fabianii as a cause of lethal septicaemiaMYCOSES, Issue 4 2006Giuseppe Valenza Summary Infections caused by rare fungal species of low pathogenic potential become increasingly common in hospital settings. The identification of these species presents a major challenge for the clinical mycology laboratory. We describe a case of fatal septicaemia caused by Candida fabianii. The use of common biochemical approaches led to misidentification of the isolate as Candida utilis. Sequencing of the internal transcribed spacer regions (ITS1 and ITS2) allowed unequivocal species identification. [source] Unique histological characteristics of Scedosporium that could aid in its identificationPATHOLOGY INTERNATIONAL, Issue 2 2010Masatomo Kimura Scedosporium prolificans has been increasingly recognized as an etiological agent of disseminated mycelial infections in profoundly immunocompromised patients. Reported herein is a case of disseminated S. prolificans infection in a patient undergoing anti-neoplastic chemotherapy for acute myeloid leukemia. Antemortem blood culture yielded S. prolificans, which was confirmed on conventional morphological examination and polymerase chain reaction-based DNA sequencing targeting internally transcribed spacer regions. Histopathology of autopsy specimens indicated fungal infection in the heart, lungs, liver, kidneys, spleen, pancreas and gastrointestinal tract, with the development of hemorrhagic and ischemic necrosis. The infecting fungus had developing septate hyphae and was identified as belonging to the genus Scedosporium, on in situ hybridization of tissue. The combination of haphazardly branching hyphae and lemon-shaped conidia appeared to be the most useful distinguishing features to allow differentiation of this fungus from other filamentous fungi in tissue. Three other unique histopathological characteristics of the fungus were noted: (i) parallel hyphae bridged at right angles to produce letter-H patterns; (ii) intravascular conidiation; and (iii) purple conidia in tissue, though these are usually described as brown in most text books. Precise histopathology, in addition to other techniques such as in situ hybridization, can aid in the identification of etiological fungi. [source] Identification, distribution and current taxonomy of Botryosphaeriaceae species associated with grapevine decline in New South Wales and South AustraliaAUSTRALIAN JOURNAL OF GRAPE AND WINE RESEARCH, Issue 1 2010W.M. PITT Abstract Background and Aims:, Botryosphaeriaceae species are recognised as important pathogens of grapevines both in Australia and overseas. The identity, prevalence and distribution of Botryosphaeriaceae species in vineyards throughout the major winegrowing regions of New South Wales (NSW) and South Australia (SA) was determined to provide a foundation for improved disease prevention and management. Methods and Results:, Field surveys from 91 vineyards across NSW and SA resulted in the collection of 2239 diseased wood samples and subsequent isolation of 1258 Botryosphaeriaceae isolates. Morphological identification along with phylogenetic analysis of ribosomal DNA internal transcribed spacer regions (ITS1-5.8S-ITS2) and partial sequences of the translation elongation factor 1-, gene (EF1-,) showed that eight Botryosphaeriaceae species from four phylogenetic lineages occur on grapevines in eastern Australia, including Diplodia seriata, Diplodia mutila, Lasiodiplodia theobromae, Neofusicoccum parvum, Neofusicoccum australe, Botryosphaeria dothidea, Dothiorella viticola and Dothiorella iberica. Conclusions:, The prevalence of individual species varied according to geography and climate. Species of Diplodia and Dothiorella, characterised by thick-walled, pigmented conidia were the most prevalent and were distributed widely throughout both NSW and SA. Species with hyaline conidia, such as Neofusicoccum and Fusicoccum, were isolated less frequently and displayed more limited geographic ranges, whilst only a single isolate of Lasiodiplodia was recovered, this being from the northern most region of NSW. Significance of the Study:, The identification of eight taxa within the Botryosphaeriaceae, and their distributions throughout south-eastern Australia was established and discussed in context with climate, reported optimum growth temperatures, and more recent taxonomic and nomenclatural revisions. We established a sound base for control strategies based on the prevailing species in Australian viticultural regions. [source] Diagnosis of common dermatophyte infections by a novel multiplex real-time polymerase chain reaction detection/identification schemeBRITISH JOURNAL OF DERMATOLOGY, Issue 4 2007M. Arabatzis Summary Background, In the absence of a functional dermatophyte-specific polymerase chain reaction (PCR), current diagnosis of dermatophytoses, which constitute the commonest communicable diseases worldwide, relies on microscopy and culture. This combination of techniques is time-consuming and notoriously low in sensitivity. Objectives, Recent dermatophyte gene sequence records were used to design a real-time PCR assay for detection and identification of dermatophytes in clinical specimens in less than 24 h. Patients and methods, Two assays based on amplification of ribosomal internal transcribed spacer regions and on the use of probes specific to relevant species and species-complexes were designed, optimised and clinically evaluated. One assay was for detecting the Trichophyton mentagrophytes species complex plus T. tonsurans and T. violaceum. The second assayed for the T. rubrum species complex, Microsporum canis and M. audouinii. Results, The analytical sensitivity of both assays was 0·1 pg DNA per reaction, corresponding to 2·5,3·3 genomes per sample. The protocol was clinically evaluated over 6 months by testing 92 skin, nail and hair specimens from 67 patients with suspected dermatophytosis. Real-time PCR detected and correctly identified the causal agent in specimens from which T. rubrum, T. interdigitale, M. audouinii or T. violaceum grew in culture, and also identified a dermatophyte species in an additional seven specimens that were negative in microscopy and culture. Conclusions, This highly sensitive assay also proved to have high positive and negative predictive values (95·7% and 100%), facilitating the accurate, rapid diagnosis conducive to targeted rather than empirical therapy for dermatophytoses. [source] Cutaneous abscess by Trichosporon asahii developing on a steroid injection site in a healthy adultCLINICAL & EXPERIMENTAL DERMATOLOGY, Issue 4 2006S. J. Yun Summary We report a rare case of cutaneous abscess by Trichosporon asahii in an immunocompetent adult. A 31-year-old Korean woman presented to our hospital with a cutaneous abscess. She had received an intralesional steroid injection 4 months earlier on the site of a hypertrophic scar. Direct sequencing of the intergenic spacer regions of the rRNA genes identified T. asahii. The decreased local immunity after the steroid injection might have triggered the infection by T. asahii. A cutaneous abscess formation by T. asahii in an immunocompetent patient is an unusual cutaneous finding that to our knowledge has not been reported previously. The local immune reaction of the skin is important for the prevention of Trichosporon infection. [source] | |||||||||||||||||||||||||