scholarly journals Detection of Laribacter hongkongensis using species-specific duplex PCR assays targeting the 16S rRNA gene and the 16S-23S rRNA intergenic spacer region (ISR)

2011 ◽  
Vol 111 (3) ◽  
pp. 625-630 ◽  
Author(s):  
L. Shen ◽  
M. Xiao ◽  
F. Kong ◽  
M. Brown ◽  
J. Sun ◽  
...  
Keyword(s):  
16S Rrna ◽  
Intergenic Spacer ◽  
23S Rrna ◽  
Rrna Gene ◽  
Duplex Pcr ◽  
Intergenic Spacer Region ◽  
Laribacter Hongkongensis ◽  
Pcr Assays ◽  
Species Specific ◽  
The 16S Rrna Gene

scholarly journals Mycoplasma mucosicanis sp. nov., isolated from the mucosa of dogs

2011 ◽  
Vol 61 (4) ◽  
pp. 716-721 ◽  
Author(s):  
Joachim Spergser ◽  
Stefan Langer ◽  
Simone Muck ◽  
Kathrin Macher ◽  
Michael Szostak ◽  
...  
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Gene Sequence ◽  
Intergenic Spacer ◽  
23S Rrna ◽  
Rrna Gene ◽  
Intergenic Spacer Region ◽  
Pcr Rflp ◽  
The 16S Rrna Gene ◽  
Sequence Similarities

Fourteen Mycoplasma strains were isolated from the oral cavity and genital tract of asymptomatic dogs. Isolates had been preliminarily identified by conventional serological testing as Mycoplasma bovigenitalium, but in 16S–23S rRNA intergenic spacer PCR-RFLP assays the isolates exhibited an RFLP pattern distinct from M. bovigenitalium PG11T. Analysis of the 16S rRNA gene placed a representative of the isolates (strain 1642T) in the M. bovigenitalium subcluster of the Mycoplasma bovis cluster of mycoplasmas, with the highest sequence similarities to Mycoplasma californicum ST-6T (96.4 %), M. bovigenitalium PG11T (96.3 %) and Mycoplasma phocirhinis 852T (96.2 %). 16S rRNA gene sequence similarities almost equidistant from three recognized species and results obtained by sequence analysis of the 16S–23S rRNA intergenic spacer region, polar lipid profiles and serological reactions indicated that this organism represents a novel species of the genus Mycoplasma for which the name Mycoplasma mucosicanis sp. nov. is proposed, with strain 1642T ( = ATCC BAA-1895T  = DSM 22457T) as the type strain.

scholarly journals Taxonomy of the canine Mollicutes by 16S rRNA gene and 16S/23S rRNA intergenic spacer region sequence comparison

2004 ◽  
Vol 54 (2) ◽  
pp. 537-542 ◽  
Author(s):  
Victoria J. Chalker ◽  
Joe Brownlie
Keyword(s):  
Phylogenetic Analysis ◽  
16S Rrna ◽  
16S Rrna Gene ◽  
Sequence Comparison ◽  
Intergenic Spacer ◽  
23S Rrna ◽  
Rrna Gene ◽  
16S Rrna Gene Sequences ◽  
Intergenic Spacer Region ◽  
The 16S Rrna Gene

The taxonomy of canine Mollicutes is described, based on phylogenetic analysis of 16S rRNA gene and 16S/23S rRNA intergenic spacer (IGS) region sequences. The nucleotide sequences of the 16S rRNA gene of two untyped mycoplasmas and the IGS region of 11 Mycoplasma species were determined and used for phylogenetic analysis. The two untyped Mycoplasma strains, HRC 689 and VJC 358, were found to be distinct from all known canine mycoplasmas and all published mycoplasma 16S rRNA gene sequences.

scholarly journals The Western Progression of Lyme Disease: Infectious and NonclonalBorrelia burgdorferi Sensu LatoPopulations in Grand Forks County, North Dakota

2014 ◽  
Vol 81 (1) ◽  
pp. 48-58 ◽  
Author(s):  
Brandee L. Stone ◽  
Nathan M. Russart ◽  
Robert A. Gaultney ◽  
Angela M. Floden ◽  
Jefferson A. Vaughan ◽  
...  
Keyword(s):  
Lyme Disease ◽  
16S Rrna ◽  
16S Rrna Gene ◽  
North Dakota ◽  
Intergenic Spacer ◽  
Rrna Gene ◽  
Content Type ◽  
Intergenic Spacer Region ◽  
Grand Forks ◽  
The 16S Rrna Gene

ABSTRACTScant attention has been paid to Lyme disease,Borrelia burgdorferi,Ixodes scapularis, or reservoirs in eastern North Dakota despite the fact that it borders high-risk counties in Minnesota. Recent reports ofB. burgdorferiandI. scapularisin North Dakota, however, prompted a more detailed examination. Spirochetes cultured from the hearts of five rodents trapped in Grand Forks County, ND, were identified asB. burgdorferi sensu latothrough sequence analyses of the 16S rRNA gene, the 16S rRNA gene-ileTintergenic spacer region,flaB,ospA,ospC, andp66. OspC typing revealed the presence of groups A, B, E, F, L, and I. Two rodents were concurrently carrying multiple OspC types. Multilocus sequence typing suggested the eastern North Dakota strains are most closely related to those found in neighboring regions of the upper Midwest and Canada. BALB/c mice were infected withB. burgdorferiisolate M3 (OspC group B) by needle inoculation or tick bite. Tibiotarsal joints and ear pinnae were culture positive, andB. burgdorferiM3 was detected by quantitative PCR (qPCR) in the tibiotarsal joints, hearts, and ear pinnae of infected mice. Uninfected larvalI. scapularisticks were able to acquireB. burgdorferiM3 from infected mice; M3 was maintained inI. scapularisduring the molt from larva to nymph; and further, M3 was transmitted from infectedI. scapularisnymphs to naive mice, as evidenced by cultures and qPCR analyses. These results demonstrate that isolate M3 is capable of disseminated infection by both artificial and natural routes of infection. This study confirms the presence of unique (nonclonal) and infectiousB. burgdorferipopulations in eastern North Dakota.

Differentiation of group VIII Spiroplasma strains with sequences of the 16S–23S rDNA intergenic spacer region

2004 ◽  
Vol 50 (12) ◽  
pp. 1061-1067 ◽  
Author(s):  
Laura B Regassa ◽  
Kimberly M Stewart ◽  
April C Murphy ◽  
Frank E French ◽  
Tao Lin ◽  
...  
Keyword(s):  
Intergenic Spacer ◽  
Analytical Models ◽  
23S Rrna ◽  
Rrna Gene ◽  
Group Viii ◽  
Similarity Coefficients ◽  
Intergenic Spacer Region ◽  
Intergenic Sequences ◽  
The Stability ◽  
The 16S Rrna Gene

Spiroplasma species (Mollicutes: Spiroplasmataceae) are associated with a wide variety of insects, and serology has classified this genus into 34 groups, 3 with subgroups. The 16S rRNA gene has been used for phylogenetic analysis of spiroplasmas, but this approach is uninformative for group VIII because the serologically distinct subgroups generally have similarity coefficients >0.990. Therefore, we investigated the utility of the 16S–23S rRNA spacer region as a means to differentiate closely related subgroups or strains. We generated intergenic sequences and detailed serological profiles for 8 group VIII Spiroplasma strains. Sequence analyses using Maximum Parsimony, Neighbor Joining, and Maximum Likelihood placed the strains into 2 clades. One clade consisted of strains BARC 2649 and GSU5367. The other clade was divided into clusters containing representatives of the 3 designated group VIII subgroups (EA-1, DF-1, and TAAS-1) and 3 previously unclassified strains. The stability of the positions of the strains in various analytical models and the ability to provide robust support for groupings tentatively supported by serology indicates that the 16S–23S intergenic rDNA sequence will prove useful in intragroup analysis of group VIII spiroplasmas.Key words: Mollicutes, Spiroplasma, phylogeny, Tabanidae.

scholarly journals Species-Specific PCR for Identification of Common Contaminant Mollicutes in Cell Culture

2001 ◽  
Vol 67 (7) ◽  
pp. 3195-3200 ◽  
Author(s):  
Fanrong Kong ◽  
Gregory James ◽  
Susanna Gordon ◽  
Anna Zelynski ◽  
Gwendolyn L. Gilbert
Keyword(s):  
16S Rrna ◽  
Cell Cultures ◽  
Bacterial Species ◽  
Intergenic Spacer ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
23S Rrna ◽  
Rrna Gene ◽  
Specific Pcr ◽  
Species Specific

ABSTRACT Mycoplasma arginini, M. fermentans, M. hyorhinis, M. orale, and Acholeplasma laidlawii are the members of the class Mollicutes most commonly found in contaminated cell cultures. Previous studies have shown that the published PCR primer pairs designed to detect mollicutes in cell cultures are not entirely specific. The 16S rRNA gene, the 16S-23S rRNA intergenic spacer region, and the 5′ end of the 23S rRNA gene, as a whole, are promising targets for design of mollicute species-specific primer pairs. We analyzed the 16S rRNA genes, the 16S-23S rRNA intergenic spacer regions, and the 5′ end of the 23S rRNA genes of these mollicutes and developed PCR methods for species identification based on these regions. Using high melting temperatures, we developed a rapid-cycle PCR for detection and identification of contaminant mollicutes. Previously published, putative mollicute-specific primers amplified DNA from 73 contaminated cell lines, but the presence of mollicutes was confirmed by species-specific PCR in only 60. Sequences of the remaining 13 amplicons were identified as those of gram-positive bacterial species. Species-specific PCR primers are needed to confirm the presence of mollicutes in specimens and for identification, if required.

scholarly journals Simultaneous Detection and Identification of Common Cell Culture Contaminant and Pathogenic Mollicutes Strains by Reverse Line Blot Hybridization

2004 ◽  
Vol 70 (3) ◽  
pp. 1483-1486 ◽  
Author(s):  
Hui Wang ◽  
Fanrong Kong ◽  
Peter Jelfs ◽  
Gregory James ◽  
Gwendolyn L. Gilbert
Keyword(s):  
Cell Culture ◽  
16S Rrna ◽  
Cell Line ◽  
Intergenic Spacer ◽  
23S Rrna ◽  
Pcr Assay ◽  
Intergenic Spacer Region ◽  
Specific Pcr ◽  
Specific Pcr Assay ◽  
Species Specific

ABSTRACT We have developed a reverse line blot (RLB) hybridization assay to detect and identify the commonest mollicutes causing cell line contamination (Mycoplasma arginini, Mycoplasma fermentans, Mycoplasma hyorhinis, Mycoplasma orale, and Acholeplasma laidlawii) and human infection (Mycoplasma pneumoniae, Mycoplasma hominis, Mycoplasma genitalium, Ureaplasma parvum, and Ureaplasma urealyticum). We developed a nested PCR assay with “universal” primers targeting the mollicute 16S-23S rRNA intergenic spacer region. Amplified biotin-labeled PCR products were hybridized to membrane-bound species-specific oligonucleotide probes. The assay correctly identified reference strains of 10 mollicute species. Cell cultures submitted for detection of mollicute contamination, clinical specimens, and clinical isolates were initially tested by PCR assay targeting a presumed mollicute-specific sequence of the 16S rRNA gene. Any that were positive were assessed by the RLB assay, with species-specific PCR assay as the reference method. Initially, 100 clinical and 88 of 92 cell culture specimens gave concordant results, including 18 in which two or more mollicute species were detected by both methods. PCR and sequencing of the 16S-23S rRNA intergenic spacer region and subsequent retesting by species-specific PCR assay of the four cell culture specimens for which results were initially discrepant confirmed the original RLB results. Sequencing of amplicons from 12 cell culture specimens that were positive in the 16S rRNA PCR assay but negative by both the RLB and species-specific PCR assays failed to identify any mollicute species. The RLB hybridization assay is sensitive and specific and able to rapidly detect and identify mollicute species from clinical and cell line specimens.

scholarly journals Species Identification and Subtyping ofUreaplasma parvum and Ureaplasma urealyticumUsing PCR-Based Assays

2000 ◽  
Vol 38 (3) ◽  
pp. 1175-1179 ◽  
Author(s):  
Fanrong Kong ◽  
Zhenfang Ma ◽  
Gregory James ◽  
Susanna Gordon ◽  
Gwendolyn L. Gilbert
Keyword(s):  
16S Rrna ◽  
Ureaplasma Urealyticum ◽  
Direct Sequencing ◽  
Intergenic Spacer ◽  
Good Evidence ◽  
23S Rrna ◽  
Rrna Gene ◽  
Specific Primer ◽  
Vaginal Swabs ◽  
Species Specific

There is good evidence that the organism currently known asUreaplasma urealyticum should be divided into two species—U. parvum (previously U. urealyticumbiovar 1) and U. urealyticum (previously U. urealyticum biovar 2). In this study, we designed a series of primers, targeting the 16S rRNA gene and 16S rRNA-23S rRNA intergenic spacer regions, the urease gene subunits, and the 5′ ends of the multiple-banded antigen (MBA) genes, to identify and subtype theseUreaplasma species. All of the species-specific primer pairs could distinguish the two species, but only subtype-specific primer pairs targeting the MBA genes could distinguish subtypes within each species. U. parvum was separated into three subtypes, represented by serovars 1, 3/14, and 6. U. urealyticum was also separated into three subtypes by PCR and/or direct sequencing. Subtype 1 consisted of serovars 2, 5, 8, and 9; subtype 2 contained serovars 4, 10, 12, and 13; and subtype 3 contained serovars 7 and 11. A selection of primer pairs was used to identify and subtype 78 clinical ureaplasma isolates from vaginal swabs of pregnant women and to identify and subtype ureaplasmas directly in 185 vaginal swabs in which they had been previously detected. U. parvum was identified in 228 (87%) of 263 isolates or specimens, and U. urealyticum was identified in 50 (19%) (both were present in 6%). Serovars 3/14 (48%) and 1 (43%) were most common among U. parvum isolates, and subtypes 2 (62%) and 1 (34%) were most common among U. urealyticum isolates. This new PCR-based typing system will facilitate future studies of the relationship between individual Ureaplasma species or subtypes and human disease.

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