Marielos Mora

Characterization of a photosynthetic Euglena strain isolated from an acidic hot mud pool of a volcanic area of Costa Rica.

Abstract Conspicuous green patches on the surface of an acidic hot mud pool located near the Rincón de la Vieja volcano (northwestern Costa Rica) consisted of apparently unialgal populations of a chloroplast-bearing euglenoid. Morphological and physiological studies showed that it is a non-flagellated photosynthetic Euglena strain able to grow in defined mineral media at temperatures up to 40 degrees C and exhibiting higher thermotolerance than Euglena gracilis SAG 5/15 in photosynthetic activity analyses. Molecular phylogeny studies using 18S rDNA and GapC genes indicated that this strain is closely related to Euglena mutabilis, another acid-tolerant photosynthetic euglenoid, forming a clade deeply rooted in the Euglenales lineage. To our knowledge this is the most thermotolerant euglenoid described so far and the first Euglenozoan strain reported to inhabit acidic hot aquatic habitats.

Comparison of Midgut Bacterial Diversity in Tropical Caterpillars (Lepidoptera: Saturniidae) Fed on Different Diets

ABSTRACT As primary consumers of foliage, caterpillars play essential roles in shaping the trophic structure of tropical forests. The caterpillar midgut is specialized in plant tissue processing; its pH is exceptionally alkaline and contains high concentrations of toxic compounds derived from the ingested plant material (secondary compounds or allelochemicals) and from the insect itself. The midgut, therefore, represents an extreme environment for microbial life. Isolates from different bacterial taxa have been recovered from caterpillar midguts, but little is known about the impact of these microorganisms on caterpillar biology. Our long-term goals are to identify midgut symbionts and to investigate their functions. As a first step, different diet formulations were evaluated for rearing two species of tropical saturniid caterpillars. Using the polymerase chain reaction (PCR) with primers hybridizing broadly to sequences from the bacterial domain, 16S rRNA gene libraries were constructed with midgut DNA extracted from caterpillars reared on different diets. Amplified rDNA restriction analysis indicated that bacterial sequences recovered from the midguts of caterpillars fed on foliage were more diverse than those from caterpillars fed on artificial diet. Sequences related to Methylobacterium sp., Bradyrhizobium sp., and Propionibacterium sp. were detected in all caterpillar libraries regardless of diet, but were not detected in a library constructed from the diet itself. Furthermore, libraries constructed with DNA recovered from surface-sterilized eggs indicated potential for vertical transmission of midgut symbionts. Taken together, these results suggest that microorganisms associated with the tropical caterpillar midgut may engage in symbiotic interactions with these ecologically important insects.

Silicate Dissolution in Las Pailas Thermal Field: Implications for Microbial Weathering in Acidic Volcanic Hydrothermal Spring Systems

A longitudinal field microcosm study was conducted in the Las Pailas hot spring system, located on the SW flank of Rincon de la Vieja, Costa Rica, in order to investigate initial microbial attachment and colonization, as well as chemical (abiotic) and biological silicate weathering under hydrothermal conditions. Solution chemistry was pH = 2.42–3.96, T = 43–89.3°C, Si = 4.45–8.19 mmol L−1, Fe = 1.50–6.95 mmol L−1and PO3− 4 = below detection limits-4.9 μmol L−1. Microcosms consisted of washed, sonicated primary silicate samples in polycarbonate vessels. The vessels were enclosed either by mesh to observe water/rock/microbial interactions or by 0.2–0.45 μm filters to observe water/rock interactions. Microcosms were incubated for periods of 6 h, 24 h, or 2 mo, fixed in the field, then analyzed in the laboratory. Scanning electron microscopy (SEM) analysis revealed that microbial attachment to mineral samples occurred in as little as 6 h. Microbial colonization and the development of minor etch pits associated with microorganisms occurred within 24 h. The most significant differences in chemical vs. biological weathering were observed after 2 mo. SEM analysis of these incubated surfaces showed that volumetric losses to mineral samples were more than one order of magnitude greater for samples that had been colonized by microorganisms and thus weathered biologically. With time, preferential colonization of anorthoclase mineral samples with Fe-oxides and apatite inclusions occurred. Subsequent weathering, therefore, may be a metabolic strategy by microorganisms to access mineral-bound PO3− 4, which is otherwise scarce in solution. Results from this study suggest that microorganisms may play a significant role in weathering in some hydrothermal systems.

Evidence for Widespread Associations between Neotropical Hymenopteran Insects and Actinobacteria

The evolutionary success of hymenopteran insects has been associated with complex physiological and behavioral defense mechanisms against pathogens and parasites. Among these strategies are symbiotic associations between Hymenoptera and antibiotic-producing Actinobacteria, which provide protection to insect hosts. Herein, we examine associations between culturable Actinobacteria and 29 species of tropical hymenopteran insects that span five families, including Apidae (bees), Vespidae (wasps), and Formicidae (ants). In total, 197 Actinobacteria isolates were obtained from 22 of the 29 different insect species sampled. Through 16S rRNA gene sequences of 161 isolates, we show that 91% of the symbionts correspond to members of the genus Streptomyces with less common isolates belonging to Pseudonocardia and Amycolatopsis. Electron microscopy revealed the presence of filamentous bacteria with Streptomyces morphology in brood chambers of two different species of the eusocial wasps. Four fungal strains in the family Ophiocordycipitacea (Hypocreales) known to be specialized insect parasites were also isolated. Bioassay challenges between the Actinobacteria and their possible targeted pathogenic antagonist (both obtained from the same insect at the genus or species level) provide evidence that different Actinobacteria isolates produced antifungal activity, supporting the hypothesis of a defensive association between the insects and these microbe species. Finally, phylogenetic analysis of 16S rRNA and gyrB demonstrate the presence of five Streptomyces lineages associated with a broad range of insect species. Particularly our Clade I is of much interest as it is composed of one 16S rRNA phylotype repeatedly isolated from different insect groups in our sample. This phylotype corresponds to a previously described lineage of host-associated Streptomyces. These results suggest Streptomyces Clade I is a Hymenoptera host-associated lineage spanning several new insect taxa and ranging from the American temperate to the Neotropical region. Our work thus provides important insights into the widespread distribution of Actinobacteria and hymenopteran insects associations, while also pointing at novel resources that could be targeted for the discovery of active natural products with great potential in medical and biotechnological applications.