Isolation and 16S DNA characterization of soil microorganisms from tropical soils capable of utilizing the herbicides hexazinone and tebuthiuron.

Six non-fermentative bacteria were isolated from Colombian (South America) and Hawaiian (USA) soils after enrichment with minimal medium supplemented with two herbicides, hexazinone (Hex) and tebuthiuron (Teb). Microscopic examination and physiological tests were followed by partial 16S DNA sequence analysis, using the first 527 bp of the 16S rRNA gene for bacterial identification. The isolated microorganisms (and in brackets, the herbicide that each degraded) were identified as: from Colombia. Methylobacterium organophilum [Teb], Paenibacillus pabuli [Teb], and Micrmbacterium foliorum [Hex]; and from Hawaii, Methylobacterium radiotolerans [Teb], Paenibacillus illinoisensis [Hex], and Rhodococcus equi [Hex]. The findings further explain how these herbicides, which have potential for illicit coca (Erythroxylum sp.) control, dissipate following their application to tropical soils.

Identification of Erythroxylum taxa by AFLP DNA analysis.

Erythroxylum coca, indigenous to the Andean region of South America, is grown historically as a source of homeopathic medicine. However, in the last century, cultivation of E. coca and several closely-related species for the production of illicit cocaine has become a major global problem. Two subspecies, E. coca var. coca and E. coca var. ipadu, are almost indistinguishable phenotypically; a related cocaine-bearing species also has two subspecies (E. novogranatense var. novogranatense and E. novogranatense var. truxillense) that are phenotypically similar, but morphologically distinguishable. The purpose of this research was to discover unique AFLP DNA patterns ("genetic fingerprinting") that characterize the four taxa and then, if successful, to evaluate this approach for positive identification of the various species of coca. Of seven different AFLP primer pairs tested, a combination of five proved optimal in differentiating the four taxa as well as a non-cocaine-bearing species, E. aerolatum. This method of DNA fragment separation was selective, and faster, for coca identification, compared with analyses based on flavonoid chemotaxonomy. Using the 5-primer AFLP approach, 132 known and unknown coca leaf accessions were evaluated. Of these, 38 were collected in 1997-2001 from illicit coca fields in Colombia, and all were genetically differentiated from coca originating in Peru and Bolivia. Based on the DNA profiling, we believe that the Colombian coca now represents a hybridization of E. coca var. ipadu. Geographical profiling within Colombia also seems feasible as new coca production areas are developed or new types of coca are introduced within traditional growing areas.

Impact of four pesticides on the growth and metabolic activities of two photosynthetic algae.

The acute toxicity was determined for soil algae Chlorella kesslerei and Anabaena inaequalis, exposed to pesticides lindane, pentachlorophenol (PCP), isoproturon (IPU), and methyl parathion (MP). Toxicity markers included growth inhibition, chlorophyll biosynthesis, and total carbohydrate content, as a function of dose and time. Concentration response functions (EC50) were estimated by probit data transformation and weighted linear regression analyses. Lindane's toxicity to Chlorella increased sharply with time (EC50 = 7490, 10.3, 0.09 mg L(-1); 24, 48, 72 h), but remained nearly constant through 72 h with Anabaena (8.7-6.7 mg L(-1); 24-72 h). PCP at low concentrations stimulated algal growth and chlorophyll a production, an effect reversed at higher doses. Anabaena was less tolerant of PCP and MP than was Chlorella. The 96-h static EC50 values for Chlorella were: 0.003, 34, 0.05, and 291 mg L(-1) for lindane, PCP, isoproturon, and MP, respectively; for Anabaena, these were 4.2, 0.13, 0.21, and 19 mg L(-1). Carbohydrate production responses were similar to those of cell density (growth) and chlorophyll biosynthesis, with MP having the lowest adverse impact. The overall relative toxicity among the four tested pesticides was: for Chlorella, lindane > IPU >> PCP >> MP; and for Anabaena, PCP > IPU > lindane > MP. The results confirm that toxicants such as these pesticides may affect individual (though related) species to significantly different degrees.