A new method is described for extraction of metagenomic DNA from soil and sediments which is based on DNA adsorption to silica without the use of phenol, ethanol precipitation or a cesium chloride gradient. High-quality DNA was obtained, and PCR inhibition was overcome by adding bovine serum albumin and adjusting magnesium concentration. By using PCR-DGGE with Firmicutes and lactic acid bacteria-specific primers the extracted metagenomic DNA was shown to contain a mixture of bacterial genomes. This method can be used for screening bacterial diversity in soil and sediment samples.

Aims:  To identify lactic acid bacteria (LAB) colonies isolated from sediments of a coastal marsh by the reduction of 2,3,5‐triphenyltetrazolium chloride (TTC) in MRS medium. Methods and Results:  Single colonies isolated from sediments of a coastal marsh by enrichment in MRS broth were selected from MRS‐TTC plates and classified according to colony phenotype based on TTC reduction. A total of 37 colonies grouped in seven different phenotypes were identified by analysis of its 16S ribosomal gene sequence. Most isolates belonged to the Firmicutes phylum, mainly to orders Bacillales and Lactobacillales. LAB were represented by 20 isolates, 15 of which belong to the genus Weissella. Conclusions:  Enrichment in MRS was highly selective for the isolation of bacteria belonging to phylum Firmicutes. Several different phenotypes were developed by LAB and must be considered during LAB isolation based on TTC reduction. Significance and Impact of the Study:  To our knowledge, this is the first study aimed at determining a relationship between colony phenotype from TTC reduction and a partial identification of isolates based on 16S ribosomal gene sequence similarities. Besides, this is the first report of isolation of W. cibaria from environmental samples.

No abstract

Aim:  Development of a method for the isolation and purification of metagenomic RNA (mgRNA) from the ectopic bacterial flora of octopus. Methods and Results:  Modifications were made to the methods of Valenzuela‐Avendaño et al. (Plant Mol Biol Rep, 2005, 23, 199a) and Chomczynski and Sacchi (Anal Biochem, 1987, 162, 156) to develop a protocol based on chemical lysis with Trizol. This proposed protocol effectively isolated mgRNA. The resulting bacterial RNA transcripts were amplified with universal primers directed to the hypervariable regions of the 16S rRNA gene by complementary DNA synthesis. Protocol efficacy in the study of metabolically active bacterial flora was proven using DGGE, which produced a banding pattern that recovered sequences mainly related to the Vibrionaceae family. Conclusion:  The analysed samples were clearly complex, and the proposed protocol was proven to effectively isolate mgRNA from the metabolically active bacterial flora associated with octopus. Significance and Impact of the Study:  This is the first protocol proposed for the isolation of bacterial mgRNA that allows identification and study of metabolically active bacterial flora associated with octopus. This is an important step forward in understanding and controlling the microbial community of this economically important fishery resource, aimed at detecting its potentially pathogenic bacteria.

Coastal bodies of water formed by the combination of seawater, underground rivers and rainwater comprise the systems with the greatest solar energy flow and biomass production on the planet. These characteristics make them reservoirs for a large number species, mainly microorganisms. Bacteria of the genus Vibrio are natural inhabitants of these environments and their presence is determined by variations in the nutrient, temperature and salinity cycles generated by the seasonal hydrologic behavior of these lagoon systems. This study determined the diversity of the genus Vibrio in 4 coastal bodies of water on the Yucatan Peninsula (Celestun Lagoon, Chelem Lagoon, Rosada Lagoon and Sabancuy Estuary). Using the molecular technique of 454 pyrosequencing, DNA extracted from water samples was analyzed and 32,807 reads were obtained belonging to over 20 culturable species of the genus Vibrio and related genera. OTU (operational taxonomic unit) richness and Chao2 and Shannon Weaver diversity indices were obtained with the database from this technique. Physicochemical and environmental parameters were determined and correlated with Vibrio diversity measured in OTUs.

Background: Octopus is a fishery product of economic importance worldwide, the main species caught on the coast of the Gulf of Mexico and the Caribbean Sea are Octopus maya and O. vulgaris, the first represents up to 95 % of national production. Goals: Identify the bacterial flora associated with commercial Octopus maya captured in the Yucatan Peninsula, using PCR-DGGE. Methods: From the metagenomic DNAs (mDNAs) extracted from samples representative of the octopus muscle, PCR products were synthesized with universal primers for bacteria (gc338F and 518R) and specific primers for Phylum Firmicutes (FirF: 369 and gcFirR: 1244). They were separated by electrophoresis in denaturing gradient gels (DGGE). The fragmented DNAs were recovered by elution, amplified (338F / 518R and FirF: 369 / FirR: 1244), sequenced and analyzed phylogenetically. Results: The sequences amplified with universal primers, after the DNA fragmentation by DGGE were associated with Psychrobacter urativorans, Psychrobacter sp, Pseudomonas sp, Pseudoalteromonas sp, Shewanella sp, Shewanella baltica, Klebsiella oxytoca, Vibrio aestuarianus, Photobacterium sp, Flavobacterium sp, F. antarcticum, Bizionia sp, Flavobacteriaceae bacterium, Bacillus sp, C. divergens, Cetobacterium somerae, Psychrilyobacter atlanticus, Salinimicrobium sp as well as, Flavobacteriaceae not yet classified. In the sequences amplified with specific primers (Phylum Firmicutes) were identified: Carnobacterium sp, Lactococcus piscium Lactococcus sp, and Vagococcus sp Conclusion: The bacterial genus detected have been reported in samples from marine environments; therefore, can be part of the native microbial diversity associated with commercial O. maya captured in the Yucatan Peninsula, Mexico.

No abstract