Upregulation of the Nifhdken Operon in Leptospirillum Species Subjected to Soluble Nitrogen Starvation

Background: Members of the genus Leptospirillum routinely dominate bioleaching systems. Understanding their metabolic complexity is necessary for the improvement of bioleaching technologies. Nitrogen is an essential element required for the cellular growth of all microorganisms. In this study Leptospirillum species were assessed for their nitrogen fixation capabilities under aerobic and anaerobic conditions, in the absence of soluble ammonia

Carbon fixation genes in biomining microorganisms

Background and aims: Studying metabolic pathways will help provide a better understanding of the role of different microorganisms within biomining environments. The majority of microorganisms involved in biomining are autotrophs which rely on atmospheric carbon fixation for growth. The aim of this study is to investigate genes involved with carbon fixation in a range of biomining microorganisms

The recovery of nucleic acid from biomining and acid mine drainage microorganisms

The cornerstone of biological molecular techniques is the extraction of the intra- or extra-cellular component of interest. However, inefficiencies in the extraction method can lead to results that are not representative of the microbial population under investigation. It is particularly difficult to extract clean and pure samples of the cellular component of interest from the microbial inhabitants of low-pH, sulfidic environments, such as those found in biomining or acid mine drainage (AMD). A number of nucleic acid (NA) extraction methods were tested using microorganisms commonly found in biomining and AMD environments, including archaea and Gram-negative and Gram-positive bacteria. The different stages in the methods of NA extraction were investigated separately, including the: (i) removal of cells from pyrite, (ii) cell lysis and nucleic acid extraction, (iii) nucleic acid precipitation and (iv) RNA purification using, as appropriate, microscopy, spectrophotometry, agarose gel electrophoresis of NA, PCR, quantitative-PCR and quantitative reverse transcriptase-PCR to assess the quality and quantity of the DNA and RNA.The relative percentage of NA recovered from each microorganism using the optimised method discussed in this paper returned the following percentage of NA per cell: At. ferrooxidans 91% ± 1.4%; At. caldus 91% ± 2.7%; L. ferriphilum 98% ± 1.2%; F. acidiphilum 83% ± 2.9%; and S. thermosulfidooxidans 79% ± 0.7%. Differences in lysis methods and NA precipitation greatly impacted the quality and quantity of the extracted NA. A method for the reliable, representative and reproducible extraction of NA from five strains of biomining and AMD microorganisms from pyrite and from liquid culture is described

Direct amplification of nodD from community DNA reveals the genetic diversity of Rhizobium leguminosarum in soil

Sequences of nodD, a gene found only in rhizobia, were amplified from total community DNA isolated from a pasture soil. The polymerase chain reaction (PCR) primers used, Y5 and Y6, match nodD from Rhizobium leguminosarum biovar trifolii, R. leguminosarum biovar viciae and Sinorhizobium meliloti. The PCR product was cloned and yielded 68 clones that were identified by restriction pattern as derived from biovar trifolii [11 restriction fragment length polymorphism (RFLP) types] and 15 clones identified as viciae (seven RFLP types). These identifications were confirmed by sequencing. There were no clones related to S. meliloti nodD. For comparison, 122 strains were isolated from nodules of white clover (Trifolium repens) growing at the field site, and 134 from nodules on trap plants of T. repens inoculated with the soil. The nodule isolates were of four nodD RFLP types, with 77% being of a single type. All four of these patterns were also found among the clones from soil DNA, and the same type was the most abundant, although it made up only 34% of the trifolii-like clones. We conclude that clover selects specific genotypes from the available soil population, and that R. leguminosarum biovar trifolii was approximately five times more abundant than biovar viciae in this pasture soil, whereas S. meliloti was rare

Akkermansia muciniphila and improved metabolic health during a dietary intervention in obesity: relationship with gut microbiome richness and ecology

Objective: Individuals with obesity and type 2 diabetes differ from lean and healthy individuals in their abundance of certain gut microbial species and microbial gene richness. Abundance of Akkermansia muciniphila, a mucin-degrading bacterium, has been inversely associated with bodyfat mass and glucose intolerance in mice, but more evidence is needed in humans. The impact of diet and weight loss on this bacterial species is unknown. Our objective was to evaluate the association between fecal A. muciniphila abundance, fecal microbiome gene richness, diet, host characteristics, and their changes after calorie restriction (CR). Design: The intervention consisted of a 6-week CR period followed by a 6-week weight stabilization (WS) diet in overweight and obese adults (N=49, including 41 women). Fecal A. muciniphila abundance, fecal microbial gene richness, diet and bioclinical parameters were measured at baseline and after CR and WS. Results: At baseline A. muciniphila was inversely related to fasting glucose, waist-to-hip ratio, and subcutaneous adipocyte diameter. Subjects with higher gene richness and A. muciniphila abundance exhibited the healthiest metabolic status, particularly in fasting plasma glucose, plasma triglycerides and body fat distribution. Individuals with higher baseline A. muciniphila displayed greater improvement in insulin sensitivity markers and other clinical parameters after CR. A. muciniphila was associated with microbial species known to be related to health. Conclusion: A. muciniphila is associated with a healthier metabolic status and better clinicaloutcomes after CR in overweight/obese adults, however the interaction between gut microbiota ecology and A. muciniphila has to be taken into account

Population dynamics of a low-grade chalcopyrite bioleaching column

Terminal Restriction Fragment Length Polymorphism (T-RFLP) was used to determine the diversity of the bacterial and archaeal populations in a bioleaching column charged with a lowgrade chalcopyrite ore and operated at 50 oC. Differing populations were identified in the leachate and the column solids but there was not discernible effect in respect of location in the column

Nucleic acid extraction from biomining microorganisms

Various methods of nucleic acid (NA) extraction were investigated with the aim of developing a quantitative method of NA extraction from five representative strains of biomining microorganisms. The process of removing cells from mineral surfaces, lysing microorganisms, precipitating NA and purifying RNA were analysed. The success of each method was examined spectrophotometrically, by agarose gel electrophoresis and PCR or quantitative real time PCR (qPCR). The most important step was shown to be cellular lysis, which principally impacted on thequantity of NA extracted from each strain. The quantity and quality of extracted NA was highlydependent on the method of NA precipitation. This study resulted in the development of a NA extraction method that reliably and reproducibly extracted NA from five strains of biomining microorganisms