| 1. |
- Bunikis, Ignas, et al.
(författare)
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Multiplex PCR as a tool for validating plasmid content of Borrelia burgdorferi.
- 2011
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Ingår i: Journal of Microbiological Methods. - : Elsevier BV. - 0167-7012 .- 1872-8359. ; 86:2, s. 243-7
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Tidskriftsartikel (refereegranskat)abstract
- Borrelia burgdorferi has an unusual genomic structure containing 21 plasmids. These plasmids carry genes that are essential for infectivity and survival of the spirochetes in vivo. Several plasmids are lost during cultivation in vitro, which might lead to a heterogeneous population after multiple passages and loss of infectivity in laboratory animals. Herein, we present a simple and inexpensive multiplex PCR method that detects the complete plasmid profile of B. burgdorferi B31 in just two PCR tubes.
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| 2. |
- Schueler, Wolfgang, et al.
(författare)
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Complete Genome Sequence of Borrelia afzelii K78 and Comparative Genome Analysis
- 2015
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Ingår i: PLOS ONE. - : Public Library of Science (PLoS). - 1932-6203. ; 10:3
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Tidskriftsartikel (refereegranskat)abstract
- The main Borrelia species causing Lyme borreliosis in Europe and Asia are Borrelia afzelii, B. garinii, B. burgdorferi and B. bavariensis. This is in contrast to the United States, where infections are exclusively caused by B. burgdorferi. Until to date the genome sequences of four B. afzelii strains, of which only two include the numerous plasmids, are available. In order to further assess the genetic diversity of B. afzelii, the most common species in Europe, responsible for the large variety of clinical manifestations of Lyme borreliosis, we have determined the full genome sequence of the B. afzelii strain K78, a clinical isolate from Austria. The K78 genome contains a linear chromosome (905,949 bp) and 13 plasmids (8 linear and 5 circular) together presenting 1,309 open reading frames of which 496 are located on plasmids. With the exception of lp28-8, all linear replicons in their full length including their telomeres have been sequenced. The comparison with the genomes of the four other B. afzelii strains, ACA-1, PKo, HLJ01 and Tom3107, as well as the one of B. burgdorferi strain B31, confirmed a high degree of conservation within the linear chromosome of B. afzelii, whereas plasmid encoded genes showed a much larger diversity. Since some plasmids present in B. burgdorferi are missing in the B. afzelii genomes, the corresponding virulence factors of B. burgdorferi are found in B. afzelii on other unrelated plasmids. In addition, we have identified a species specific region in the circular plasmid, cp26, which could be used for species determination. Different non-coding RNAs have been located on the B. afzelii K78 genome, which have not previously been annotated in any of the published Borrelia genomes.
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| 3. |
- Ameur, Adam, et al.
(författare)
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CanvasDB : a local database infrastructure for analysis of targeted- and whole genome re-sequencing projects
- 2014
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Ingår i: Database. - : Oxford University Press (OUP). - 1758-0463. ; , s. bau098-
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Tidskriftsartikel (refereegranskat)abstract
- CanvasDB is an infrastructure for management and analysis of genetic variants from massively parallel sequencing (MPS) projects. The system stores SNP and indel calls in a local database, designed to handle very large datasets, to allow for rapid analysis using simple commands in R. Functional annotations are included in the system, making it suitable for direct identification of disease-causing mutations in human exome-(WES) or whole-genome sequencing (WGS) projects. The system has a built-in filtering function implemented to simultaneously take into account variant calls from all individual samples. This enables advanced comparative analysis of variant distribution between groups of samples, including detection of candidate causative mutations within family structures and genome-wide association by sequencing. In most cases, these analyses are executed within just a matter of seconds, even when there are several hundreds of samples and millions of variants in the database. We demonstrate the scalability of canvasDB by importing the individual variant calls from all 1092 individuals present in the 1000 Genomes Project into the system, over 4.4 billion SNPs and indels in total. Our results show that canvasDB makes it possible to perform advanced analyses of large-scale WGS projects on a local server.
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| 4. |
- Ameur, Adam, et al.
(författare)
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Comprehensive profiling of the vaginal microbiome in HIV positive women using massive parallel semiconductor sequencing
- 2014
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Ingår i: Scientific Reports. - : Springer Science and Business Media LLC. - 2045-2322. ; 4, s. 4398-
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Tidskriftsartikel (refereegranskat)abstract
- Infections by HIV increase the risk of acquiring secondary viral and bacterial infections and methods are needed to determine the spectrum of co-infections for proper treatment. We used rolling circle amplification (RCA) and Ion Proton sequencing to investigate the vaginal microbiome of 20 HIV positive women from South Africa. A total of 46 different human papillomavirus (HPV) types were found, many of which are not detected by existing genotyping assays. Moreover, the complete genomes of two novel HPV types were determined. Abundance of HPV infections was highly correlated with real-time PCR estimates, indicating that the RCA-Proton method can be used for quantification of individual pathogens. We also identified a large number of other viral, bacterial and parasitic co-infections and the spectrum of these co-infections varied widely between individuals. Our method provides rapid detection of a broad range of pathogens and the ability to reconstruct complete genomes of novel infectious agents.
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| 5. |
- Ameur, Adam, et al.
(författare)
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De Novo Assembly of Two Swedish Genomes Reveals Missing Segments from the Human GRCh38 Reference and Improves Variant Calling of Population-Scale Sequencing Data
- 2018
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Ingår i: Genes. - : MDPI AG. - 2073-4425 .- 2073-4425. ; 9:10
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Tidskriftsartikel (refereegranskat)abstract
- The current human reference sequence (GRCh38) is a foundation for large-scale sequencing projects. However, recent studies have suggested that GRCh38 may be incomplete and give a suboptimal representation of specific population groups. Here, we performed a de novo assembly of two Swedish genomes that revealed over 10 Mb of sequences absent from the human GRCh38 reference in each individual. Around 6 Mb of these novel sequences (NS) are shared with a Chinese personal genome. The NS are highly repetitive, have an elevated GC-content, and are primarily located in centromeric or telomeric regions. Up to 1 Mb of NS can be assigned to chromosome Y, and large segments are also missing from GRCh38 at chromosomes 14, 17, and 21. Inclusion of NS into the GRCh38 reference radically improves the alignment and variant calling from short-read whole-genome sequencing data at several genomic loci. A re-analysis of a Swedish population-scale sequencing project yields > 75,000 putative novel single nucleotide variants (SNVs) and removes > 10,000 false positive SNV calls per individual, some of which are located in protein coding regions. Our results highlight that the GRCh38 reference is not yet complete and demonstrate that personal genome assemblies from local populations can improve the analysis of short-read whole-genome sequencing data.
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| 6. |
- Andrade, Pedro, et al.
(författare)
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Regulatory changes in pterin and carotenoid genes underlie balanced color polymorphisms in the wall lizard
- 2019
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Ingår i: Proceedings of the National Academy of Sciences of the United States of America. - : Proceedings of the National Academy of Sciences. - 0027-8424 .- 1091-6490. ; 116:12, s. 5633-5642
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Tidskriftsartikel (refereegranskat)abstract
- Reptiles use pterin and carotenoid pigments to produce yellow, orange, and red colors. These conspicuous colors serve a diversity of signaling functions, but their molecular basis remains unresolved. Here, we show that the genomes of sympatric color morphs of the European common wall lizard (Podarcis muralis), which differ in orange and yellow pigmentation and in their ecology and behavior, are virtually undifferentiated. Genetic differences are restricted to two small regulatory regions near genes associated with pterin [sepiapterin reductase (SPR)] and carotenoid [beta-carotene oxygenase 2 (BCO2)] metabolism, demonstrating that a core gene in the housekeeping pathway of pterin biosynthesis has been coopted for bright coloration in reptiles and indicating that these loci exert pleiotropic effects on other aspects of physiology. Pigmentation differences are explained by extremely divergent alleles, and haplotype analysis revealed abundant transspecific allele sharing with other lacertids exhibiting color polymorphisms. The evolution of these conspicuous color ornaments is the result of ancient genetic variation and cross-species hybridization.
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| 7. |
- Bárcena-Uribarri, Iván, et al.
(författare)
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P66 porins are present in both Lyme disease and relapsing fever spirochetes : a comparison of the biophysical properties of P66 porins from six Borrelia species
- 2010
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Ingår i: Biochimica et Biophysica Acta - Biomembranes. - : Elsevier. - 0005-2736 .- 1879-2642. ; 1798:6, s. 1197-1203
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Tidskriftsartikel (refereegranskat)abstract
- The genus Borrelia is the cause of the two human diseases: Lyme disease (LD) and relapsing fever (RF). BothLD and RF Borrelia species are obligate parasites and are dependent on nutrients provided by their hosts. Thefirst step of nutrient uptake across the outer membrane of these Gram-negative bacteria is accomplished bywater-filled channels, so-called porins. The knowledge of the porin composition in the outer membranes ofthe different pathogenic Borrelia species is limited. Only one porin has been described in relapsing feverspirochetes to date, whereas four porins are known to be present in Lyme disease agents. From these, theBorrelia burgdorferi outer membrane channel P66 is known to act as an adhesin and was well studied as aporin. To investigate if P66 porins are expressed and similarly capable of pore formation in other Borreliacausing Lyme disease or relapsing fever three LD species (B. burgdorferi, B. afzelii, B. garinii) and three RFspecies (B. duttonii, B. recurrentis and B. hermsii) were investigated for outer membrane proteins homologousto P66. A search in current published RF genomes, comprising the ones of B. duttonii, B. recurrentis and B.hermsii, indicated that they all contained P66 homologues. The P66 homologues of the six Borrelia specieswere purified to homogeneity and their pore-forming abilities as well as the biophysical properties of thepores were analyzed using the black lipid bilayer assay.
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| 8. |
- Bárcena-Uribarri, Iván, et al.
(författare)
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Use of nonelectrolytes reveals the channel size and oligomeric constitution of the Borrelia burgdorferi P66 porin
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Annan publikation (övrigt vetenskapligt/konstnärligt)abstract
- The outer membrane protein P66 of the Lyme disease spirochete Borrelia burgdorferi is capable of pore formation with an atypical high single-channel conductance of 11 nS in 1 M KCl. We studied in a non-theoretical manner the diameter of the P66 channel by analyzing its single-channel conductance in black lipid bilayers in the presence of different nonelectrolytes with known hydrodynamic radii. Furthermore, we calculated the filling of the channel with these nonelectrolytes and the results revealed that nonelectrolytes with hydrodynamic radii of 0.34 nm or smaller pass through the pore, whereas neutral molecules with greater radii only partially filled the channel or were not able to enter it at all. Thus, the diameter of the P66 entrance was determined to be ≤ 1.9 nm with a constriction site diameter of about 0.7 nm. Furthermore, the P66-induced membrane conductance could be blocked by 80-90% after addition of the nonelectrolytes PEG 400, PEG 600 and maltohexaose in the low millimolar range. Interestingly, the analysis of the power density spectra of P66 after blockage with nonelectrolytes revealed no chemical interaction responsible for channel block. The blockage of one P66 single-channel conductance unit of 11 nS occurred by seven subconducting states, thus indicating a heptameric organization of the P66 oligomer. This organization of P66 as a heptamer was confirmed by Blue Native PAGE and immunoblot analysis, which demonstrated that P66 forms a complex with a mass of approximately 460 kDa.
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| 9. |
- Berggrund, Malin, et al.
(författare)
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Temporal changes in the vaginal microbiota in self-samples and its association with persistent HPV16 infection and CIN2
- 2020
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Ingår i: Virology Journal. - : Springer Science and Business Media LLC. - 1743-422X. ; 17
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Tidskriftsartikel (refereegranskat)abstract
- BackgroundThe vaginal microbiota has been reported to be associated with HPV infection and cervical cancer. This study was performed to compare the vaginal microbiota at two timepoints in women performing self-sampling and had a persistent or transient HPV16 infection. The women were tested for 12 high-risk HPV (hrHPV) types but only women with single type (HPV16) were included to reduce confounding variables.MethodsIn total 96 women were included in this study. Of these, 26 were single positive for HPV16 in the baseline test and HPV negative in the follow-up test and 38 were single positive for HPV16 in both tests and diagnosed with CIN2+ in histology. In addition, 32 women that were negative for all 12 HPV tested were included. The samples of vaginal fluid were analyzed with the Ion 16S™ Metagenomics Kit and Ion 16S™ metagenomics module within the Ion Reporter™ software.ResultsK-means clustering resulted in two Lactobacillus-dominated groups, one with Lactobacillus sp. and the other specifically with Lactobacillus iners. The two remaining clusters were dominated by a mixed non-Lactobacillus microbiota. HPV negative women had lower prevalence (28%) of the non-Lactobacill dominant cluster in the baseline test, as compared to women with HPV16 infection (42%) (p value = 0.0173). Transition between clusters were more frequent in women with persistent HPV16 infection (34%) as compared in women who cleared the HPV16 infection (19%) (p value = 0.036).ConclusionsThe vaginal microbiota showed a higher rate of transitioning between bacterial profiles in women with persistent HPV16 infection as compared to women with transient infection. This indicate an instability in the microenvironment in women with persistent HPV infection and development of CIN2+.
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| 10. |
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