Unsaturated soil columns were used to examine the transport of the plasmid pLEPO1 and plant DNA (transplastomic tobacco DNA), both carrying an antibiotic resistance gene (
aadA gene), and the ...capacity of bacteria to incorporate the gene in their genome after its passage through the soil. Soil columns containing a top leaf layer had sterile water percolated through them at a rate of 0.5
mL
h
−1. DNA from column leachate water was extracted and analyzed. Quantitative measurements included total DNA concentrations in the water and the transformation frequencies of
Acinetobacter sp. BD413 by DNA in the column effluent. Qualitative measurements included the relative degradation of DNA after passage in the columns by agarose gel electrophoresis and the potential of effluent DNA to transform bacteria, leading to the production of antibiotic-resistant bacteria. The presence of
aadA gene in the leachate water of soil columns suggests the mobility of DNA in unsaturated soil medium. The extent of DNA degradation was found to be proportional to its residence time in the soil column while a fraction of DNA was always able to incorporate into the
Acinetobacter genome under all conditions studied. These results suggest that biologically active transgenic DNA might be transported downward by rain in unsaturated soils.
To evaluate the effects of management practices and seasons on a soil bacterial community and the composition of ammonia-oxidizing bacteria (AOB), molecular screenings were compared among ...Mediterranean (Sardinia) soils with different plant covers and different agricultural practices, namely cork oak forest, tilled/non-tilled vineyard, hay crop and pasture. We compared the fingerprints from both independent replicates and pooled samples to ascertain the best approach for studying the environmental effects on bacterial composition. The soil microbial biomass, which was estimated from the amounts of extracted soil dsDNA, was 2 to 3 folds higher in the spring than in the autumn; in the spring, it was negatively correlated with the intensity of land use. A 16S rDNA DGGE experiment confirmed that both the land use and season markedly affect the composition of the soil bacterial community. Tilled vineyard soil exhibited the lowest similarities in community structures, suggesting that tillage induced the most marked disturbance among the tested land management methods. Distinct AOB populations were found for each type of land use; among these types, the cork oak forest proved to be a protective habitat for AOB against environmental changes. Our results suggest that the comparative community level and group-specific fingerprinting enabled an accurate evaluation of multiple factors in soil bacterial structures when performed with both independent and pooled replicates.
Probes for the detection of Azospirillum strains were obtained from DNA fragments generated by random amplification of polymorphic DNA (RAPD) and tested to assess their specificity towards DNA ...extracted from pure cultures. The most specific probe, referred to as alpha 4, produced a hybridization signal only with amplified DNA of A. lipoferum ATCC29731. This strain was inoculated, together with two other Azospirillum strains, in soil microcosms of different complexity and its presence tested with the probe alpha 4. This probe confirmed its high specificity with amplified DNA extracted from the soil microcosm and in the presence of other A. lipoferum strains, indicating that the strategy for bacterial detection, based on RAPD markers, is useful for monitoring the presence of a particular strain under environment-like conditions. Other RAPD-derived probes, when tested on soil samples. did not show the same level of specificity as that shown on DNA from pure cultures. This result suggests that some precautions are necessary in the choice of a really specific RAPD marker. In a further development of this strategy, the alpha 4 probe was sequenced and two pairs of "nested" primers were designed, which enabled a diagnostic polymerase chain reaction from soil samples that was specific for the A. lipoferum species.
Summary
Background
Pyoderma gangrenosum (PG) is a rare skin disease characterized clinically by ulcers with undermined borders, and histologically by neutrophil‐rich infiltrates. PG may occur alone, ...in syndromic forms or associated with systemic diseases, such as inflammatory bowel disease and haematological or rheumatological disorders.
Objectives
To determine a specific genetic background related to autoinflammation for PG.
Methods
We assessed autoinflammation by evaluating the cytokine profile and genes involved in classic autoinflammatory diseases in 13 patients with PG and in seven patients with the syndromic form, known as PASH (pyoderma gangrenosum, acne and suppurative hidradenitis).
Results
In skin samples, the expression of interleukin (IL)‐1β and its receptors, IL‐17 and its receptor, and tumour necrosis factor‐α and its receptors were significantly higher in both PG (P = 0·001) and in PASH (P < 0·001) than in controls. The chemokines IL‐8; chemokine (C‐X‐C motif) ligand 1/2/3; chemokine (C‐X‐C motif) ligand 16; and RANTES (regulated on activation, normal T‐cell‐expressed and secreted) were also overexpressed. Cases of PG and PASH showed mutations in the autoinflammatory genes MEFV, NLRP3, NLRP12, NOD2, LPIN2 and PSTPIP1.
Conclusions
Overexpression of cytokines/chemokines, along with genetic changes, supports the hypothesis that PG and its syndromic form, PASH, are a spectrum of polygenic autoinflammatory conditions.
What's already known about this topic?
Pyoderma gangrenosum (PG) is a prototypic neutrophilic dermatosis manifesting as skin ulcers. It may also occur in the context of syndromes like PAPA (pyogenic arthritis, pyoderma gangrenosum and acne) and PASH (pyoderma gangrenosum, acne and suppurative hidradenitis).
Although an autoinflammatory origin has been demonstrated for its syndromic forms, to date a specific genetic background related to autoinflammation has not been proven for pyoderma gangrenosum.
What does this study add?
The clear‐cut increase in skin expression of interleukin (IL)‐1β and IL‐17 and the presence of mutations in the main genes involved in autoinflammation indicate that PG is a polygenic autoinflammatory condition, as previously demonstrated in PASH.
The involvement of proinflammatory cytokines could address the use of biological drugs blocking IL‐1 or IL‐17 in patients with refractory PG and in patients with PASH.
The Han:SPRD cy/+ strain develops a form of slowly progressive disease that appears similar in many respects to that seen in the autosomal dominant polycystic kidney disease (ADPKD) in humans. We ...have performed a total genome scan in an experimental backcross population derived from affected Han:SPRD cy/+ rat (PKD) and non-affected Wistar Ottawa Karlsburg rat (WOK) using 117 microsatellite markers. The genetic dissection of PKD allowed us to map on rat chromosome 5, a quantitative trait locus (QTL) controlling PKD, kidney mass and plasma urea concentration. The homology region is likely to reside on human chromosome 8. The gene responsible for PKD in Han:SPRD cy/+ rat is neither PKD1, localised on human chromosome 16, nor PKD2, localised on human chromosome 4. Therefore, we propose that this new locus be denoted PKDr1. The detection of the PKDr1 locus and associated QTL should accelerate research into the genetic causes of ADPKD.
The aim of this work was to study variations in the composition of eubacteria and ammonia-oxidizing populations of soil, both determined by denaturing gradient gel electrophoresis (DGGE), after the ...addition of a pharmaceutical fungal biomass, treated to degrade its DNA. This waste can be used as an amendment. The fungal biomass waste was added at three rates: 0.05, 0.1, and 1% per dry weight of soil. Control soil, without any amendment, was also investigated. Total DNA was extracted, purified, and amplified by using either universal (eubacteria) or specific (amoA) primers. Amplicons were separated by DGGE. Sequencing was also carried out to better assess the diversity of ammonia oxidizing bacteria. Changes in the composition of eubacterial community were detected after 3 days only in the soil treated with the highest dose, while the ammonia oxidizing population responded more promptly (after 1 day) with evident modifications at level of Nitrosolobus like sequences.
Molecular and biochemical investigation methods have been exploited to evaluate the effects of pig slurry, used for three years as a fertilizer in intensive agriculture, on the presence and activity ...of chemolithotrophic ammonia oxidizing bacteria, the biocatalysts of the first step in the nitrification process. The evaluation was carried out, on bulk soil, comparing data from the first and the third year of fertilization. Oligonucleotide sequences selected from the 16S rRNA genes of autotrophic ammonia oxidizers have been used as specific PCR amplification primers and probes. The potential nitrification activity (PNA) has been used to determine the effects of swine manure fertilization on the ammonia oxidizing activity. Members of the genera
Nitrosomonas and
Nitrosospira were always detectable, but the genus
Nitrosospira was more represented than
Nitrosomonas. The differences between the hybridization signals were correlated with the doses of pig slurry and the number of applications. The mineral (urea) and organic fertilizations showed similar effects on the ammonia oxidisers investigated by molecular and biochemical methods. It was found that additions of swine manure to soil plots increased both potential ammonia-oxidise activity and ammonia oxidiser sequences. Stimulation of the soil ammonia-oxidise activity was due mainly to its effects on the indigenous bacterial population.
In search of a possible involvement of the human herpesvirus type 6 (HHV-6) in human Hodgkin’s and non-Hodgkin’s lymphomas, we studied the levels of anti–HHV-6 antibodies in the sera of 94 cases by ...an immunofluorescence assay, as well as the presence of HHV-6 sequences in the affected tissues of 66 cases by polymerase chain reaction, using one set of primer oligonucleotides. Our results showed higher anti–HHV-6 antibody titers in human lymphomas than in normal blood donors, but the difference is statistically significant only when normal donors are compared with Hodgkin’s lymphoma cases. HHV-6 sequences were detected in 3 of 25 Hodgkin’s lymphomas and 0 of the 41 cases of non-Hodgkin’s lymphomas studied. The three cases positive for HHV-6 sequences belong to the nodular sclerosis-lymphocyte depletion histologic subtype and share remarkable similarities in their clinical features. Furthermore, Southern blot analysis of total genomic DNA obtained from the neoplastic tissues of two of the three patients showed the same restriction fragment length polymorphism. Our results suggest that: (1) the high level of anti–HHV-6 antibodies in Hodgkin’s disease is due to an activation of the immune system not related to the presence of HHV-6 sequences in affected lymph nodes; (2) the presence of HHV-6 sequences in human lymphoid tissues is not a frequent event, rather it is in fact a very rare event in non-Hodgkin’s lymphomas, while in Hodgkin’s cases it is more frequent than previously reported on the basis of Southern blot analysis; and (3) the presence of HHV-6 sequences in Hodgkin’s lymphomas may have a relation with the clinical presentation of the disease.
Congenital Central Hypoventilation Syndrome (CCHS) is a rare condition characterized by an alveolar hypoventilation due to a deficient autonomic central control of ventilation and a global autonomic ...dysfunction. Paired-like homeobox 2B (PHOX2B) mutations are found in most of the patients with CCHS. In recent years, the condition has evolved from a life-threatening neonatal onset disorder to include broader and milder clinical presentations, affecting children, adults and families. Genes other than PHOX2B have been found responsible for CCHS in rare cases and there are as yet other unknown genes that may account for the disease. At present, management relies on lifelong ventilatory support and close follow up of dysautonomic progression. BODY: This paper provides a state-of-the-art comprehensive description of CCHS and of the components of diagnostic evaluation and multi-disciplinary management, as well as considerations for future research.
Awareness and knowledge of the diagnosis and management of this rare disease should be brought to a large health community including adult physicians and health carers.
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