Evidence of two genetic clusters of manatees with low genetic diversity in Mexico and implications for their conservation.

The Antillean manatee (Trichechus manatus manatus) occupies the tropical coastal waters of the Greater Antilles and Caribbean, extending from Mexico along Central and South America to Brazil. Historically, manatees were abundant in Mexico, but hunting during the pre-Columbian period, the Spanish colonization and throughout the history of Mexico, has resulted in the significantly reduced population occupying Mexico today. The genetic structure, using microsatellites, shows the presence of two populations in Mexico: the Gulf of Mexico (GMx) and Chetumal Bay (ChB) on the Caribbean coast, with a zone of admixture in between. Both populations show low genetic diversity (GMx: N(A) = 2.69; H(E) = 0.41 and ChB: N(A) = 3.0; H(E) = 0.46). The lower genetic diversity found in the GMx, the largest manatee population in Mexico, is probably due to a combination of a founder effect, as this is the northern range of the sub-species of T. m. manatus, and a bottleneck event. The greater genetic diversity observed along the Caribbean coast, which also has the smallest estimated number of individuals, is possibly due to manatees that come from the GMx and Belize. There is evidence to support limited or unidirectional gene flow between these two important areas. The analyses presented here also suggest minimal evidence of a handful of individual migrants possibly between Florida and Mexico. To address management issues we suggest considering two distinct genetic populations in Mexico, one along the Caribbean coast and one in the riverine systems connected to the GMx.

Molecular cloning and biochemical characterization of a family-9 endoglucanase with an unusual structure from the gliding bacteria Cytophaga hutchinsonii.

Cytophaga hutchinsonii was originally isolated from sugarcane piles. This microorganism therefore probably produces an array of enzymes allowing it to digest cellulosic substrates. C. hutchinsonii thus represents a rich source of potentially effective cellulase enzymes that can be harnessed for conversion of biomass to simple sugars. These sugars can then be used as feedstock for ethanol production or other chemical syntheses. In this study, we report the PCR cloning of an endoglucanase gene (Cel9A) from C. hutchinsonii using degenerated primers directed at the catalytic domain. Alignment of the amino acids sequence revealed that Cel9A has a gene structure totally different from the other known cellulose degraders. The most striking feature of this cloned protein is the absence of a cellulose-binding domain (CBD), which to date was believed to be imperative in cellulose hydrolysis. Consequently, the Cel9A gene, encoding beta-1,4 endoglucanase from C. hutchinsonii was overexpressed in Escherichia coli with a His-Tag based expression vector. The resulting polypeptide, with a molecular mass of 105 KDa, was purified from cell extracts by affinity chromatography on cellulose. Mature Cel9A was optimally active at pH 5.0 and 45 degrees C. The enzyme efficiently hydrolyzes carboxymethyl- cellulose (CMC). Analysis of CMC and filter paper hydrolysis suggests that Cel9A is a nonprocessive enzyme with endo-cellulase activities.

Phylogeography, phylogeny and hybridization in trichechid sirenians: implications for manatee conservation.

Abstract The three living species of manatees, West Indian (Trichechus manatus), Amazonian (Trichechus inunguis) and West African (Trichechus senegalensis), are distributed across the shallow tropical and subtropical waters of America and the western coast of Africa. We have sequenced the mitochondrial DNA control region in 330 Trichechus to compare their phylogeographic patterns. In T. manatus we observed a marked population structure with the identification of three haplotype clusters showing a distinct spatial distribution. A geographic barrier represented by the continuity of the Lesser Antilles to Trinidad Island, near the mouth of the Orinoco River in Venezuela, appears to have restricted the gene flow historically in T. manatus. However, for T. inunguis we observed a single expanding population cluster, with a high diversity of very closely related haplotypes. A marked geographic population structure is likely present in T. senegalensis with at least two distinct clusters. Phylogenetic analyses with the mtDNA cytochrome b gene suggest a clade of the marine Trichechus species, with T. inunguis as the most basal trichechid. This is in agreement with previous morphological analyses. Mitochondrial DNA, autosomal microsatellites and cytogenetic analyses revealed the presence of hybrids between the T. manatus and T. inunguis species at the mouth of the Amazon River in Brazil, extending to the Guyanas and probably as far as the mouth of the Orinoco River. Future conservation strategies should consider the distinct population structure of manatee species, as well as the historical barriers to gene flow and the likely occurrence of interspecific hybridization.