mtDNA diversity in rhesus monkeys reveals overestimates of divergence time and paraphyly with neighboring species

Reconstructions of the human-African great ape phylogeny by using mitochondrial DNA (mtDNA) have been subject to considerable debate. One confounding factor may be the lack of data on intraspecific variation. To test this hypothesis, we examined the effect of intraspecific mtDNA diversity on the phylogenetic reconstruction of another Plio- Pleistocene radiation of higher primates, the fascicularis group of macaque (Macaca) monkey species. Fifteen endonucleases were used to identify 10 haplotypes of 40-47 restriction sites in M. mulatta, which were compared with similar data for the other members of this species group. Interpopulational, intraspecific mtDNA diversity was large (0.5%- 4.5%), and estimates of divergence time and branching order incorporating this variation were substantially different from those based on single representatives of each species. We conclude that intraspecific mtDNA diversity is substantial in at least some primate species. Consequently, without prior information on the extent of genetic diversity within a particular species, intraspecific variation must be assessed and accounted for when reconstructing primate phylogenies. Further, we question the reliability of hominoid mtDNA phylogenies, based as they are on one or a few representatives of each species, in an already depauperate superfamily of primates.      

Variation in the ribosomal internal transcribed spacers and 5.8S rDNA among five species of Acropora (Cnidaria; Scleractinia): patterns of variation consistent with reticulate evolution

The ITS sequences of Acropora spp. are the shortest so far identified in any metazoan and are among the shortest seen in eukaryotes; ITS1 was 70-80 bases, and ITS2 was 100-112 bases. The ITS sequences were also highly variable, but base composition and secondary structure prediction indicate that divergent sequence variants are unlikely to be pseudogenes. The pattern of variation was unusual in several other respects: (1) two distinct ITS2 types were detected in both A. hyacinthus and A. cytherea, species known to hybridize in vitro with high success rates, and a putative intermediate ITS2 form was also detected in A. cytherea; (2) A. valida was found to contain highly (29%) diverged ITS1 variants; and (3) A. longicyathus contained two distinct 5.8S rDNA types. These data are consistent with a reticulate evolutionary history for the genus Acropora.      

Chromatin decondensation and DNA synthesis in human sperm activated in vitro by using Xenopus laevis egg extracts

An in vitro sperm activation system was used to study nuclear swelling-chromatin decondensation and DNA synthesis; processes that occur in vivo following fertilization. Lysolecithin-permeabilized human sperm were incubated in Xenopus laevis egg extract and examined by using phase-contrast light microscopy, electron microscopy, and autoradiography. During a 3-hour incubation, the activated sperm nuclear chromatin underwent a decondensation-recondensation cycle during which DNA was synthesized. This also occurred when egg extract was given a 3-hour preincubation before the addition of the sperm, suggesting that the factor(s) required for initiating the decondensation-recondensation cycle is associated with the sperm. Because both nuclear swelling and DNA synthesis were found to be reproducible and quantifiable, we studied the effect of various agents on the two processes, characterizing the critical component(s) in the egg extract that induces these events. EGTA was found to have no effect on the induced nuclear swelling or DNA synthesis that occurs in the activated sperm. Freezing and thawing the extract or treating the extract with aphidicolin also had no effect on subsequent nuclear swelling; however, the DNA synthesis activity was blocked. Sperm incubated in extract treated with alkaline phosphatase (AP) had both nuclear swelling and DNA synthesis blocked. However, if the sperm were pretreated with DTT, and then incubated with the AP-treated extract, only the DNA synthesis activity of the extract was blocked. When the extract was treated with serine protease inhibitors (PMSF, soybean trypsin inhibitor, or alpha-2-macroglobulin), nuclear swelling occurred; however, DNA synthesis was blocked. These data suggest that phosphoproteins are involved in one or more of the activation events and that a serine protease(s) is involved in the synthesis of DNA.     

Measurement of normal and anomalous diffusion of dyes within protein structures fabricated via multiphoton excited cross-linking

We demonstrate microscale spatial and chemical control of diffusion within protein matrixes created through the use of nonlinear multiphoton excited photochemistry. The mobility of fluorescent dyes of different mass and composition within controlled cross-linked environments has been measured using two-photon excited fluorescence recovery after photobleaching (FRAP). The diffusion times for several rhodamine and sulforhodamine dyes within these fabricated structures were found to be approximately 3-4 orders of magnitude slower than in free solution. The precise diffusion times can be tuned by varying the laser exposure during the fabrication of the matrix, and the diffusion can be correlated with the mesh size determined by TEM and Flory-Rehner analysis. We find that the hydrophobic Texas Red dyes (sulforhodamines) exhibit diffusion that is highly anomalous, indicative of a strong interaction with the hydrophobic cross-linked protein matrix. These results suggests the use of these cross-linked protein matrixes as ideal model systems in which to systematically study anomalous diffusion. Finally, the diffusion can be tuned within a multilayered protein matrix, and this in conjunction with slow diffusion also suggests the use of these structures in controlled release applications.     

Serosurvey of Brucella spp. infection in the Kafue lechwe (Kobus leche kafuensis) of the Kafue flats in Zambia

One of the diseases of veterinary and public health importance affecting the Kafue lechwe (Kobus leche kafuensis) on the Kafue flats is brucellosis, for which only scant information is available. During the 2003 (October), 2004 (December), and 2008 (July-December) hunting seasons in the Kafue flats, we conducted a study to determine the seroprevalence of Brucella spp. in the Kafue lechwe and to evaluate serologic tests for detection of Brucella spp. antibodies in lechwe. The Rose Bengal Test (RBT), competitive enzyme-linked immunosorbent assay (cELISA), and fluorescence polarization assay (FPA) were used. A total of 121 Kafue lechwe were hunted for disease investigations in 2003, 2004, and 2008 in the Kafue Flat Game Management Area. Of these, 21.6%, (95% confidence interval [CI]: 14.2-29.1%) had detectable antibodies to Brucella spp. The Kafue lechwe in Lochnivar National Park had higher antibody results than those in Blue Lagoon National Park (odds ratio=3.0; 95% CI: 0.94-9.4). Infection levels were similar in females (21.6%) and males (21.7%). Results were similar among RBT, FPA, cELISA tests, suggesting that these could effectively be used in diagnosing brucellosis in the Kafue lechwe. Our study demonstrates the presence of Brucella infections in the Kafue lechwe in two national parks located in the Kafue flats and further highlights the suitability of serologic assays for testing the Kafue lechwe. Because the Kafue lechwe is the most hunted wildlife species in Zambia, hunters need to be informed of the public health risk of Brucella spp. infection.