Papers by Keyword: 16S rRNA Gene

Abstract: Twelve probiotic Lactobacillus strains for poultry were characterised by amplified ribosomal DNA restriction analysis (ARDRA) using Sau3AI, TaqI, HaeIII and AluI restriction endonucleases. Species-specific and strain-specific restriction patterns were observed from the bacterial strains. Numerical analysis of composite analysis of ARDRA exhibited D value of 0.8456. Whereas, the caculated D values of ARDRA patterns generated by Sau3AI, TaqI, HaeIII and AluI were 0.8309, 0.8382,0.8088 and 0.8088, repectively. Composite analysis of ARDRA was the most discriminative method when compared to the individual analysis. ARDRA could distinguished L. reuteri C 10 and L. panis C 17 into single strains. The 16S rRNA gene restriction patterns were also able to group L. gallinarum I 16 and I 26 into single strains. Lactobacillus brevis I 12, I 23, I 25, I 211 and I 218 seem to be multiple clones of the same bacterial strain as are L. reuteri C 1 and C 16. ARDRA is a valuable fingerprinting method to discriminate probiotic Lactobacillus strains.

Abstract: Thermophilic bacteria live at temperatures above 45⁰ C. Many investigations focused on their potential as sources of highly active enzymes ‘termostable enzyme’ and other products such as antibiotics and compatible solutes. Lake Linow is an active volcanic lake located in Tomohon City, North Sulawesi, Indonesia. Lake Linow becomes the habitat of thermophilic bacteria. A study has been conducted to obtain isolates of thermophilic bacteria and to identifikasi berdasarkan gen 16 s RNA. Bacterial DNA extraction procedure using the Presto TM Mini gDNA Bacteria Kit Geneaid protocol, with modifications. Amplification of 16s RNA gene using PCR method. Visualization of 16 s RNA amplicon genes with automatic electrophoresis capiler Qiaxel, Qiagen. Sequencing was carried out using Singapore's First BASE Sequencing service. The results showed that IL2 isolates and IL3 isolates could live up to 700C. Alignment analysis results using NCAST BLBI IL2 isolates showed 99% similarity with Bacillus thuringiensis strain H2682 (accession number CP009720.1). While isolate of IL3 thermophilic bacteria showed 94% similarity with Bacillus licheniformis strain 14DA11 (accession number CP023168.1). The results of phylogeny reconstruction with neighbor joining method, gene sequence 16S rRNA isolate IL2 showed the closest relation with Bacillus thuringiensis strain HD1011 (accession number CP009335.1). While IL3 isolate showed the closest relation with Bacillus licheniformis strain 14DA11 (accession number CP023168.1).

Abstract: Relatively little is known about the microbial communities present in natural environments that meet physico-chemical conditions for the development of potential leaching microorganisms such as thermal ecosystems from the Chilean Altiplano. Thermophilic leaching enrichments were obtained and identified from a high altitude solfataric pound in Lirima hot springs in the Chilean Altiplano. This ecosystem is characterized by hot underground freshwaters, enriched in sulfur compounds showing pH from neutral to acidic. Microbial diversity has been scarcely explored here, and preliminary results demonstrate that hydrothermal pounds are represented by thermophilic anaerobic and acidophilic taxa. Thermophilic leaching cultures in shake flasks were obtained using ferrous iron and pyrite as energy source. The presence of Bacteria and Archaea in oxidizing enrichments was determined by PCR amplification of 16S rRNA genes. A preliminary analysis of microbial diversity using massive sequencing revealed that Bacteria were more abundant than Archaea in both enrichments. Specifically for the iron-oxidizing culture, the majority of the sequences clustered within the Proteobacteria phylum (79%). Among Proteobacteria, the proportion of Betaproteobacteria (42.2%) and Gammaproteobacteria (21.27%) was much higher than that of Alphaproteobacteria (15.5%). Within the Betaproteobacteria class, the most frequent genus was Leptothrix-like. Similar results were obtained for the pyrite oxidizing culture. Interestingly, this study shows the presence of microorganisms close to the Leptothrix genus under low pH conditions (1.7-2.8) and their capacity to grow at high temperatures with ferrous iron or pyrite as sole energy source

Abstract: Bioleaching processes are usually open systems where introduced and native microorganisms survive to changes in pH, temperature, salt and metal concentration, among others. Spatial and temporal description of the microbial community could be relevant for better comprehension of copper extraction process and help in the development of operative procedures to improve the metal extraction. We performed metagenomics and high-throughput sequencing of 16S rRNA genes analyses on samples from Escondida mine bioleaching heap and laboratory columns tests. Archaeal community structure in samples was assessed using three pairs of Archaea-specific primers, and results were highly depending on the primers pairs used. Similarly, three pairs of Bacteria-specific primers were used to assess the bacterial community. Moreover, according to the metagenomics analysis, At. thiooxidans, F. acidarmanus, Leptospirillum spp., Acidiphilium sp. JA12-A1, Acidiphilium spp., At. ferrivorans, and Leptospirillum ferriphilum were the most representative microorganisms. The repercussion of the different methodologies and outputs in the characterization of the bioleaching microbial community is discussed. A better understanding of the microbial community in bioleaching processes could improve the analysis regarding environmental changes in the heap process, its metallurgical performance and, can be used to assist in the decision-making process.

Abstract: Acid mine drainage (AMD) presents numerous problems to the aquatic life and surrounding ecosystems. However, little is known about the geographic distribution, diversity, composition, structure and function of AMD microbial communities. In this study, an AMD-specific microarray was used to analyze nine AMD microbial communities, and showed that those nine AMD microbial communities had high variations measured by the number of detected genes, overlapping genes between samples, unique genes, and diversity indices. Statistical analyses indicated that the concentrations of Fe, S, Ca, Mg, Zn, Cu and pH had strong impacts on both phylogenetic and functional diversity, composition, and structure of AMD microbial communities. This study provides insights into our understanding of the geographic distribution, diversity, composition, structure and functional potential of AMD microbial communities and key environmental factors shaping them.

Abstract: In this study, API 50CH strips and 16S rRNA gene sequence analysis were used to determine the species and genus of strain LD33 isolated from traditional dairy products, and the phylogenetic tree was constructed by MEGA4 software with Neighbor-Joining method analysis, the results showed that LD33 had a closest relationship with Enterococcus faecalis ATCC19433T (DQ411814), there was 100% sequence similarity between them.

Abstract: The methods of culture-dependent and denaturing gradient gel electrophoresis (DGGE) based on the sequence of 16S rRNA V3 region gene were described to comparatively characterize the microbial population and community structure of cutlassfish (Trichiurus haumela) under the cold storage. The results showed that 13 kinds of bacteria were identified by the traditional culture-dependent methods, the dominant bacteria belonged to Shewanella putrefaciens and Pseudomonas fluorescens. To determine the community profiles of the samples on variable V3 region, the bacteria of 16S rRNA gene were amplified by PCR and 11 distinct PCR products were separated by DGGE fingerprinting technology. From the sequence analysis, Psychrobacter sp. was found to be the predominant bacteria in the initial stage of the storage. The proportion of Shewanella sp., Pseudomonas sp. increased gradually with the extension of storage time, and they took the place of Psychrobacter sp. to be the dominant bacteria. Thereinto, both Pseudomonas fluorescens and Vibrio sp. took high proportions in the process of storage due to the deterioration of cutlassfish (Trichiurus haumela).

Abstract: The largest lignite mining area in Europe is located 150 km southeast of Berlin. Acidic lakes exist in this area, known to be caused by marcasite oxidation. Thirty-two samples from the open-pit brown coal-mine Jaenschwalde were analyzed for microorganisms. Cell numbers determined after separation from sand particles revealed concentrations of 102 to 107 microorganisms per g sample. In samples exposed to the air within an hour, up to 4x107 cells were counted. Measurement of metabolic activity by microcalorimetry showed for such samples up to 50 µW per g sand, whereas in heap samples (with low moisture) low or even no activity was measurable. DNA extraction was successful for 28 samples. In 26 samples microbial 16S rRNA genes were amplified by PCR. Acidithiobacillus ferrooxidans and At. thiooxidans specific amplificates were detected by nested PCR in 23 and 10 cases, respectively. A specific signal indicating Leptospirillum ferrooxidans was obtained with nine samples. Random samples were sequenced and showed 96 to 99 % identity with published data of all three species. Surprisingly, in four samples archaeal 16S rRNA genes were amplified by PCR. Sequencing of two samples showed 99 % identity with unidentified or uncultured archaea found in NCBI-databases. Molecular biology results for At. ferrooxidans as well as for At. thiooxidans were supported by successful isolations of pure cultures in 23 cases. Cultivation of the archaea failed so far. These data indicate that iron- and sulfur-oxidizing microorganisms occur at these sites in large numbers. If in addition the evidence for archaea can become verified, a screening for hot spots as the sites of their occurrence would become interesting.

Abstract: The distribution and diversity of acidophilic microbe at the ore surface of Zijinshan commercial low-grade copper bioleaching heap operated at pH 0.8 were investigated. Samples taken from -1m, -2 m, -3m from the top of heap surface were investigated by 16S rRNA gene clone library. Phylogenetic analyses of 16S rRNA fragments revealed that the retrieved bacterial sequences mainly related to genus Acidithiobacillus (64.6%), genus Leptospirillum (27.3%), genus Sulfobacillus (3.1%) and genus Ferrimicrobium(1.6%). For archaea, only Ferroplasma acidiphilum was detected. Bacterial diversity in the heap was increased from surface layer to underground. The proportion of genus Leptospirillum was sharply reduced (from 48.5% to 5%) from higher depth to lower depth and reverse correlation of increased A.ferrooxidans (from 0.9% to 15%) and S. thermotolerans (from undetectable to 7%) were found in the heap. Sulfur oxidizers including A.albertensis, A.caldus and A.thiooxidans also vertically increased from higher depth to lower depth (from 50.5% to 80%). These results indicated that genus Acidithiobacillus especially sulfur oxidizers A.albertensis, A.caldus and A.thiooxidans may play very important roles in the commercial low-grade copper bioleaching heap.