Prospecting environmental mycobacteria: combined molecular approaches reveal unprecedented diversity

Background: Environmental mycobacteria (EM) include species commonly found in various terrestrial and aquatic environments, encompassing animal and human pathogens in addition to saprophytes. Approximately 150 EM species can be separated into fast and slow growers based on sequence and copy number differences of their 16S rRNA genes. Cultivation methods are not appropriate for diversity studies; few studies have investigated EM diversity in soil despite their importance as potential reservoirs of pathogens and their hypothesized role in masking or blocking M. bovis BCG vaccine. Methods: We report here the development, optimization and validation of molecular assays targeting the 16S rRNA gene to assess diversity and prevalence of fast and slow growing EM in representative soils from semi tropical and temperate areas. New primer sets were designed also to target uniquely slow growing mycobacteria and used with PCR-DGGE, tag-encoded Titanium amplicon pyrosequencing and quantitative PCR. Results: PCR-DGGE and pyrosequencing provided a consensus of EM diversity; for example, a high abundance of pyrosequencing reads and DGGE bands corresponded to M. moriokaense, M. colombiense and M. riyadhense. As expected pyrosequencing provided more comprehensive information; additional prevalent species included M. chlorophenolicum, M. neglectum, M. gordonae, M. aemonae. Prevalence of the total Mycobacterium genus in the soil samples ranged from 2.3×107 to 2.7×108 gene targets g−1; slow growers prevalence from 2.9×105 to 1.2×107 cells g−1. Conclusions: This combined molecular approach enabled an unprecedented qualitative and quantitative assessment of EM across soil samples. Good concordance was found between methods and the bioinformatics analysis was validated by random resampling. Sequences from most pathogenic groups associated with slow growth were identified in extenso in all soils tested with a specific assay, allowing to unmask them from the Mycobacterium whole genus, in which, as minority members, they would have remained undetected

Histomorphometric analysis of inflammatory response and necrosis in re-implanted central incisor of rats treated with low-level laser therapy

Low-level laser therapy is a tool employed in the management of post-operative inflammation process and in the enhancement of reparative process. The aim of the study was to perform histological evaluation of dental and periodontal ligament of rats central upper-left incisor teeth re-implanted and irradiated with low-level laser (InGaAl, 685 nm, 50 J/cm2) 15, 30, and 60 days after re-implantation. Seventy-two male rats had the central upper left incisor removed and kept for 15 min on dry gauze before replantation. Laser was irradiated over the root surface and empty alveolus prior replantation and over surrounding mucosa after the re-implantation. After histological procedures, all slices were analyzed regarding external resorption area and histological aspects. We observed an increase of root resorption (p < 0.05) in the control group compared to the laser group at 15, 30, and 60 days. These results showed that the laser groups developed less root resorption areas than the control group in all experimental periods. Additionally, histological analysis revealed less inflammatory cells and necrotic areas in laser groups

Differential, Positional-Dependent Transcriptional Response of Antigenic Variation (var) Genes to Biological Stress in Plasmodium falciparum

1% of the genes of the human malaria causing agent Plasmodium falciparum belong to the heterogeneous var gene family which encodes P. falciparum erythrocyte membrane protein 1 (PFEMP1). This protein mediates part of the pathogenesis of the disease by causing adherence of infected erythrocytes (IE) to the host endothelium. At any given time, only one copy of the family is expressed on the IE surface. The cues which regulate the allelic exclusion of these genes are not known. We show the existence of a differential expression pattern of these genes upon exposure to biological stress in relation to their positional placement on the chromosome – expression of centrally located var genes is induced while sub-telomeric copies of the family are repressed - this phenomenon orchestrated by the histone deacetylase pfsir2. Moreover, stress was found to cause a switch in the pattern of the expressed var genes thus acting as a regulatory cue. By using pharmacological compounds which putatively affect pfsir2 activity, distinct changes of var gene expression patterns were achieved which may have therapeutic ramifications. As disease severity is partly associated with expression of particular var gene subtypes, manipulation of the IE environment may serve as a mechanism to direct transcription towards less virulent genes

A prospective clinical trial on the influence of a triamcinolone/demeclocycline and a calcium hydroxide based temporary cement on pain perception

<p>Abstract</p> <p>Introduction</p> <p>The aim of this clinical trial was to compare the degree of short term post-operative irritation after application of a triamcinolone/demeclocycyline based or a calcium hydroxide based provisional cement.</p> <p>Methods</p> <p>A total of 109 patients (55 female and 54 male; mean age: 51 ± 14 years) with primary or secondary dentinal caries were randomly assigned to the two treatment groups of this biomedical clinical trial (phase III). Selection criteria were good systemic health and treated teeth, which were vital and showed no symptoms of pulpitis. Up to three teeth were prepared for indirect metallic restorations, and the provisional restorations were cemented with a triamcinolone/demeclocycyline (Ledermix) or a calcium hydroxide (Provicol) based material. The intensity of post-operative pain experienced was documented according to the VAS (4, 12, 20, 24, and 82 h) and compared to VAS baseline.</p> <p>Results</p> <p>A total of 159 teeth were treated (Ledermix: 83 teeth, Provicol: 76 teeth). The minor irritation of the teeth, experienced prior to treatment, was similar in both groups; however, 4 h after treatment this value was significantly higher in the Provicol group than in the Ledermix group (p < 0.005, t-test). After 12 h, the difference was no longer significant. The number of patients taking analgesics for post-treatment pain was higher in the Provicol group (n = 11/53) than in the Ledermix group (n = 3/56).</p> <p>Conclusions</p> <p>The patients had no long term post-operative pain experience in both groups. However, within the first hours after cementation the sensation of pain was considerably higher in the Provicol group than in the Ledermix group.</p