Biodiversity of Actinomycetes associated with Caribbean sponges and their potential for natural product discovery.

Marine actinomycetes provide a rich source of structurally unique and bioactive secondary metabolites. Numerous genera of marine actinomycetes have been isolated from marine sediments as well as several sponge species. In this study, 16 different species of Caribbean sponges were collected from four different locations in the coastal waters off Puerto Rico in order to examine diversity and bioactive metabolite production of marine actinomycetes in Caribbean sponges. Sediments were also collected from each location, in order to compare actinomycete communities between these two types of samples. A total of 180 actinomycetes were isolated and identified based on 16S rRNA gene analysis. Phylogenetic analysis revealed the presence of at least 14 new phylotypes belonging to the genera Micromonospora, Verruscosispora, Streptomyces, Salinospora, Solwaraspora, Microbacterium and Cellulosimicrobium. Seventy-eight of the isolates (19 from sediments and 59 from sponges) shared 100 % sequence identity with Micromonospora sp. R1. Despite having identical 16S rRNA sequences, the bioactivity of extracts and subsequent fractions generated from the fermentation of both sponge- and sediment-derived isolates identical to Micromonospora sp. R1 varied greatly, with a marked increase in antibiotic metabolite production in those isolates derived from sponges. These results indicate that the chemical profiles of isolates with high 16S rRNA sequence homology to known strains can be diverse and dependent on the source of isolation. In addition, seven previously reported dihydroquinones produced by five different Streptomyces strains have been purified and characterized from one Streptomyces sp. strain isolated in this study from the Caribbean sponge Agelas sceptrum.

Novel linear megaplasmid from Brevibacterium sp. isolated from extreme environment.

Brevibacterium sp. Ap13, isolated from flamingo's feces in Laguna Aparejos, a high-altitude lake located at approximately 4,200 m in the northwest of Argentina was previously found to be resistant to multiple antibiotics, and was therefore screened for plasmids that may be implicated in antibiotic resistance. Brevibacterium sp. Ap13 was found to contain two plasmids of approximately 87 and 436 kb, designated pAP13 and pAP13c, respectively. Only pAP13 was stably maintained and was extensively characterized by pulsed-field gel electrophoresis to reveal that this plasmid is linear and likely has covalently linked terminal proteins associated with its 5' ends. This is the first report of a linear plasmid in the genus Brevibacterium and may provide a new tool for genetic manipulation of this commercially important genus.

First report of linear megaplasmids in the genus Micrococcus.

High-altitude wetlands (above 4200m) in the northwest of Argentina are considered pristine and extreme environments. Micrococcus sp. A1, H5, and V7, isolated from such environments, were shown to contain linear megaplasmids, designated pLMA1, pLMH5, and pLMV7, respectively. As known from linear plasmids of other actinomycetes, all three plasmids were resistant to lambda exonuclease treatment, which is consistent with having terminal proteins covalently attached to their 5' DNA ends. Electrophoretic mobility, Southern analysis, and restriction endonuclease patterns revealed pLMA1 and pLMH5 being indistinguishable plasmids, even though they were found in different strains isolated from two distant wetlands - Laguna Azul and Laguna Huaca Huasi. Analysis of 16S rDNA sequences of Micrococcus sp. A1, H5, and V7 suggested a close relationship to Micrococcus luteus. Typing of isolates was performed using fingerprint patterns generated by BOX-PCR. Plasmid-deficient strains, generated from Micrococcus sp. A1, showed a significantly decreased resistance level for erythromycin.

Screening of heavy metal-tolerant actinomycetes isolated from the Salí River.

Fifty-three strains of actinomycetes resistant to heavy metals were isolated from the Salí River in northwest Argentina. Screening procedures that involve solid and liquid synthetic media containing Cd(2+), Cu(2+), or Hg(2+) allowed the selection of six strains. These strains showed a quantitative sorption of Cd(2+) and Cu(2+) by more than 98% of the initial metal concentrations (0.1, 0.5, and 1.0 mM) tested.