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Lars Mølbak

Publications and source records attributed to Lars Mølbak.

8 recordsLinked to original sources
PubMed2006

Root growth and exudate production define the frequency of horizontal plasmid transfer in the Rhizosphere.

To identify the main drivers of plasmid transfer in the rhizosphere, conjugal transfer was studied in the rhizospheres of pea and barley. The donor Pseudomonas putida KT2442, containing plasmid pKJK5::gfp, was coated onto the seeds, while the recipient P. putida LM24, having a chromosomal insertion of dsRed, was inoculated into the growth medium. Mean transconjugant-to-donor ratios in vermiculite were 4.0+/-0.8 x 10(-2) in the pea and 5.9+/-1.4 x 10(-3) in the barley rhizospheres. In soil, transfer ratios were about 10 times lower. As a result of a 2-times higher root exudation rate in pea, donor densities in pea (1 x 10(6)-2 x 10(9) CFU g(-1) root) were about 10 times higher than in barley. No difference in recipient densities was observed. In situ visualization of single cells on the rhizoplane and macroscopic visualization of the colonization pattern showed that donors and transconjugants were ubiquitously distributed in the pea rhizosphere, while they were only located on the upper parts of the barley roots. Because the barley root elongated about 10 times faster than the pea root, donors were probably outgrown by the elongating barley root. Thus by affecting the cell density and distribution, exudation and root growth appear to be key parameters controlling plasmid transfer in the rhizosphere.

Gene Transfer, Horizontal↗
PubMed2006

Identification of a novel, invasive, not-yet-cultivated Treponema sp. in the large intestine of pigs by PCR amplification of the 16S rRNA gene.

Laser capture microdissection in combination with fluorescent in situ hybridization was used to identify an unknown species of spirochetes from the pig colonic mucosa. The 16S rRNA gene was PCR amplified, and the closest related type strain was Treponema bryantiiT (90.1%). The spirochete, here named "Candidatus Treponema suis," was associated with colitis, including invasion of the surface epithelium as well as superficial parts of the mucosa.

Animals↗
PubMed2006

Freezing at -800 degrees C distorts the DNA composition of bacterial communities in intestinal samples.

Terminal-restriction fragment length polymorphism (T-RFLP) was used to evaluate how to store intestinal specimens for bacterial community analysis. Bacterial communities are increasingly often described by means of DNA-based methods and it is common practice to store intestinal or faecal specimens either at -20 degrees C or -80 degrees C. In this study, samples of intestines from five different pigs were stored at -80 degrees C and -20 degrees C, respectively and a thawing and freezing procedure was carried out three times for each intestinal per pig per temperature. The cumulative sum of the T-RFLP peak heights (T-RF intensities) decreased as the temperature decreased. The composition of the bacterial community changed when stored at -80 degrees C compared to the samples stored at -20 degrees C. Thus it is recommended from this study that samples of intestinal content are stored at -20 degrees C before use for bacterial community analysis, instead of the current practice at -80 degrees C.

Animals↗
PubMed2005

Comparison of methods and animal models commonly used for investigation of fecal microbiota: effects of time, host and gender.

Denaturing gradient gel electrophoresis (DGGE), terminal restriction fragment length polymorphism (T-RFLP) and plating on selective agars were used to study variation in the fecal microbiota of rats over time as well as variation between individuals. Investigated rats were either conventional and specific pathogen free (SPF), or human flora associated (HFA). A higher variation (p<0.05) in fecal microbiota over time was observed for HFA than for SPF animals. Analysis of DGGE and T-RFLP profiles of fecal microbiota from SPF and HFA rats revealed that variation over time was less significant than variation between individuals, and that phylogenetic profiles clustered according to gender. These observations should be taken into account when designing future research addressing changes in fecal microbiota.

Adult↗
PubMed2005

Luteibacter rhizovicinus gen. nov., sp. nov., a yellow-pigmented gammaproteobacterium isolated from the rhizosphere of barley (Hordeum vulgare L.).

Three strains of Gram-negative, aerobic, yellow-pigmented, chemo-organotrophic bacteria, motile by a polar flagellum, were isolated from the rhizosphere of spring barley (Hordeum vulgare L.) at a research field near Copenhagen, Denmark. The three strains, LJ79, LJ96T and LJ99, formed visible colonies on one-tenth-strength tryptic soy broth supplemented with agar (1/10 TSBA) after incubation for 6 days at 15 degrees C. The strains hydrolysed starch, casein (skimmed milk), gelatin and various pentoses and hexoses and grew on MacConkey agar and full-strength TSBA. Growth on 1/10 TSBA occurred at 4-30 degrees C, pH 6-9 and 0-3 % (w/v) NaCl. The strains had identical 16S rRNA gene sequences and ERIC (enterobacterial repetitive intergenic consensus sequence) fingerprint profiles, but could be differentiated by their RAPD (random amplified polymorphic DNA) fingerprint patterns. Strain LJ96(T) had a DNA G+C content of 64.3 mol% and the major fatty acids were 15 : 0 iso (23.4 %), 17 : 1 iso omega9c (25.5 %) and 17 : 0 iso (18.1 %). Phylogenetic analysis of the 16S rRNA gene sequences of the three strains showed 96 % sequence similarity to Rhodanobacter lindaniclasticus LMG 18385T, 95 % to Frateuria aurantia DSM 6220T and 96 % to Fulvimonas soli LMG 19981T. Using LJ96T DNA as probe, DNA-DNA hybridizations documented the relationship of the three strains to a single species (87.4-98.7 % relatedness) and showed less than 30 % relatedness to Frateuria aurantia DSM 6220T and Fulvimonas soli DSM 14263T. Rhodanobacter lindaniclasticus LMG 18385T is not extant and the strain not available from any public strain collections, thus DNA-DNA hybridization could not include this strain. On the basis of genotypic and phenotypic characteristics, the three yellow-pigmented strains could also be differentiated from Frateuria aurantia, Fulvimonas soli and Rhodanobacter lindaniclasticus. The name Luteibacter rhizovicinus gen. nov., sp. nov. is proposed, with the type strain LJ96T (=DSM 16549T=ATCC BAA-1015T).

DNA, Bacterial↗
PubMed2005

Laser capture microdissection of bacterial cells targeted by fluorescence in situ hybridization.

Direct cultivation-independent sequence retrieval of unidentified bacteria from histological tissue sections has been limited by the difficulty of selectively isolating specific bacteria from a complex environment. Here, a new DNA isolation approach is presented for prokaryotic cells. By this method, a potentially pathogenic strain of the genus Brachyspira from formalin-fixed human colonic biopsies were visualized by fluorescence in situ hybridization (FISH) with a 16S rRNA-targeting oligonucleotide probe, followed by laser capture microdissection (LCM) of the targeted cells. Direct 16S rRNA gene PCR was performed from the dissected microcolonies, and the subsequent DNA sequence analysis identified the dissected bacterial cells as belonging to the Brachyspira aalborgi cluster 1. The advantage of this technique is the ability to combine the histological recognition of the specific bacteria within the tissue with molecular analysis of 16S rRNA gene or other genes of interest. This method is widely applicable for the identification of noncultivable bacteria and their gene pool from formalin-fixed paraffin-embedded tissue samples.

Brachyspira↗
PubMed2003

Plasmid transfer from Pseudomonas putida to the indigenous bacteria on alfalfa sprouts: characterization, direct quantification, and in situ location of transconjugant cells.

The transfer of the plasmids pJKJ5 and TOL (pWWO) from Pseudomonas putida to the indigenous bacterial community on alfalfa sprouts was studied. Tagging with fluorescent protein markers allowed direct quantification of the introduced donor bacteria and of indigenous bacteria that had received the plasmids. The sprouts were observed for 9 days; during this time alfalfa seeds, inoculated with donor bacteria, developed to edible and subsequently decaying sprouts. The first transconjugants were detected on day 6 after donor inoculation and occurred at frequencies of 3.4 x 10(-4) and 2.0 x 10(-6) transconjugant cells per donor cell for pKJK5::gfp and TOL::gfp, respectively. Confocal laser scanning microscopy revealed that the sprouts were heavily colonized with donors and that most transconjugants were located around the hypocotyl and root areas. Randomly selected members of the indigenous bacterial community from both inoculated and uninoculated sprouts, as well as a representative part of the community that had received the plasmids, were characterized by polymorphisms of PCR-amplified ribosomal DNA (rDNA) spacer regions between the 16S and 23S genes, followed by partial 16S rDNA sequencing. This showed that the initially dominating genera Erwinia and Paenibacillus were gradually replaced by Pseudomonas on the fully developed sprouts. Transconjugants carrying either of the investigated plasmids mainly belonged to the genera Pseudomonas and ERWINIA: The numbers of transconjugant cells did not reach detectable levels until 6 days after the onset of germination, at which point these species constituted the majority of the indigenous bacteria. In conclusion, the alfalfa sprouts provided an environment that allowed noteworthy frequencies of plasmid transfer from P. putida in the absence of selective pressure that could favor the presence of the investigated plasmids.

Bacteria↗