OPHTHALMIC EXAMINATION FINDINGS AND INTRAOCULAR PRESSURE MEASUREMENTS IN SIX SPECIES OF ANURA.

A complete ophthalmic exam, including intraocular pressure (IOP) measurement, is key to diagnosing ocular diseases such as uveitis and glaucoma in frogs. We performed complete ophthalmic anterior segment examinations and IOP rebound tonometry measurements using two different settings (other "p" and canine "d") for six anuran species. The objectives were to describe common ocular abnormalities found in these species, to compare IOP values between different tonometer settings, and to compare IOPs between species. Examinations revealed abnormalities including cataracts (11/98 total eyes), lenticular sclerosis (10/98) and lipid keratopathy (9/98). IOP was measured with the TonoVet® and the ranges (oculus uterque, OU mm Hg other "p" setting, canine "d" setting) were giant waxy monkey tree frogs (Phyllomedusa bicolor) (3.5-7.6; 6.5-11.7; n = 5), mission golden-eyed tree frogs (Trachycephalus resinifictrix) (7.0-9.7, 13.2-15.7; n = 6), boreal toads (Bufo boreas boreas) (0.8-5.5, 5.7-10.5; n = 13), Mexican giant tree frogs (Pachymedusa dacnicolor) (3.8-5.0, 8.3-11.8; n = 3), Lake Titicaca frogs (Telmatobius culeus) (8.8-10.5, 14.0-17.2; n = 8), and mossy tree frogs (Theloderma corticale) (9.7-11.0, 15.7-17.0; n = 5). The TonoVet canine "d" setting IOP measurements were statistically higher (P = 0.01) than the other "p" setting measurements for all species except the giant waxy monkey tree frogs. IOP was significantly lower for giant waxy monkey tree frog eyes with cataracts (P < 0.05) with the other "p" setting. IOP did not statistically differ in eyes with lenticular sclerosis. IOP can be measured by rebound tonometer in anurans, but more research is needed for species-specific references using consistent settings.

Post-publication sharing of data and tools.

Despite existing guidelines on access to data and bioresources, good practice is not widespread. A meeting of mouse researchers in Rome proposes ways to promote a culture of sharing.

Cutaneous Bacteria of Confiscated Telmatobius culeus in Lima, Peru.

The Lake Titicaca frog is endangered due to threats such as water pollution, introduced species, and overharversting for markets, where people consume them as frog juice. This study, conducted June to November 2012, aimed to determinate the bacteria microflora living on the skin of frogs confiscated from the La Parada market, Lima, Peru, and housed individually in the Laboratory of Wildlife at the Faculty of Veterinary Medicine and Zootechnic of the Universidad Peruana Cayetano Heredia in Lima, Peru. Samples collected with sterile swabs and cultured on blood, tryptic soy, and MacConkey agars were investigated using commercially available test kits, to investigate the commonly encountered bacterial and potentially zoonotic microorganisms associated with their consumption. We found three species of zoonotic concern in the genus Vibro: Vibrio alginolyticus, Vibro cholerae, and Vibro fluvialis. Other Gram-negative species cultured included two different colonies of Aeromonas hydrophila, or Aeromonas caviae or Aeromonas sobria; Pseudomona luteola; one example of Weeksella virosa or Empedobacter brevis; and Citrobacter freundii. Gram-positive bacteria detected were Staphylococcus spp., Micrococcus spp., and Erysipelothrix rhusiopathiae. We recommend against the consumption of this frog due to the pathogens it may carry that could cause serious illness among consumers and in vendors who handle animals.

Chytrid Fungus, Batrachochytrium dendrobatidis , in Wild Populations of the Lake Titicaca Frog, Telmatobius culeus, in Peru.

The Lake Titicaca frog (Telmatobius culeus) is critically endangered, primarily from overexploitation. However, additional threats, such as chytrid fungus ( Batrachochytrium dendrobatidis ), are poorly studied. We found moderate levels of chytrid infection using quantitative PCR. Our results enhance our understanding of chytrid tolerance to high pH and low water temperature.

Batrachochytrium dendrobatidis in Confiscated Telmatobius in Lima, Peru.

The Peruvian Andes are the home of 27 species of frogs of the genus Telmatobius, many of which are critically endangered. Illegal trade of adult frogs for purported medical properties likely represents the major threat facing these species. This activity, besides reducing their populations, may contribute to the dissemination of the fungus Batrachochytrium dendrobatidis (Bd), which causes chytridiomycosis, a disease posing a threat to many populations of amphibians. We screened frogs confiscated by the Administration of Forestry and Wildlife in Lima, Peru, for Bd. We used real-time PCR to diagnose Bd at the Laboratory of Wildlife, Faculty of Veterinary Medicine and Zootecnics, Universidad Peruana Cayetano Heredia, in Lima and Pisces Molecular Laboratory in Boulder, Colorado, US. Of 62 samples collected during this study, 60% (37) were PCR positive for Bd, confirming that illegal trade of amphibians can pose a risk for disseminating Bd.

Miniaturized sealed-tube allele-specific PCR.

DNA diagnostics has been progressively moving from expensive, low-throughput, multi-step methods towards inexpensive, robust, and high-throughput methods. Here we describe the further validation and refinement of a recently described novel genotyping method that has the latter characteristics. An evolved form of allele-specific PCR, the method generates a fluorescent signal through the use of universal labeled primers, which can be quantified directly from microplates using standard plate readers. We have applied the method successfully to a test set of 12 novel single nucleotide polymorphisms (SNPs) on a panel of 47 individuals using low reaction volumes. We demonstrate that the method is extremely accurate, robust, and can be optimized in a simple and predictable manner. By conducting the assay in closed-tube format, the potential for contamination is reduced to a minimum. By virtue of its simplicity, the method is versatile and cost-effective with potential for use in industrial-scale genetic studies or in the clinical diagnostic setting.

Alternative splicing modulates Ubx protein function in Drosophila melanogaster.

The Drosophila Hox gene Ultrabithorax (Ubx) produces a family of protein isoforms through alternative splicing. Isoforms differ from one another by the presence of optional segments-encoded by individual exons-that modify the distance between the homeodomain and a cofactor-interaction module termed the "YPWM" motif. To investigate the functional implications of Ubx alternative splicing, here we analyze the in vivo effects of the individual Ubx isoforms on the activation of a natural Ubx molecular target, the decapentaplegic (dpp) gene, within the embryonic mesoderm. These experiments show that the Ubx isoforms differ in their abilities to activate dpp in mesodermal tissues during embryogenesis. Furthermore, using a Ubx mutant that reduces the full Ubx protein repertoire to just one single isoform, we obtain specific anomalies affecting the patterning of anterior abdominal muscles, demonstrating that Ubx isoforms are not functionally interchangeable during embryonic mesoderm development. Finally, a series of experiments in vitro reveals that Ubx isoforms also vary in their capacity to bind DNA in presence of the cofactor Extradenticle (Exd). Altogether, our results indicate that the structural changes produced by alternative splicing have functional implications for Ubx protein function in vivo and in vitro. Since other Hox genes also produce splicing isoforms affecting similar protein domains, we suggest that alternative splicing may represent an underestimated regulatory system modulating Hox gene specificity during fly development.

Models for financial sustainability of biological databases and resources.

Following the technological advances that have enabled genome-wide analysis in most model organisms over the last decade, there has been unprecedented growth in genomic and post-genomic science with concomitant generation of an exponentially increasing volume of data and material resources. As a result, numerous repositories have been created to store and archive data, organisms and material, which are of substantial value to the whole community. Sustained access, facilitating re-use of these resources, is essential, not only for validation, but for re-analysis, testing of new hypotheses and developing new technologies/platforms. A common challenge for most data resources and biological repositories today is finding financial support for maintenance and development to best serve the scientific community. In this study we examine the problems that currently confront the data and resource infrastructure underlying the biomedical sciences. We discuss the financial sustainability issues and potential business models that could be adopted by biological resources and consider long term preservation issues within the context of mouse functional genomics efforts in Europe.

Sharing genomes: an integrated approach to funding, managing and distributing genomic clone resources.

National biological resource centres have a vital role in archiving and distributing biological reagents that result from large-scale genome programmes. These reagents are invaluable to the research community as they enable independent validation of results disclosed in peer review and provide tools that facilitate the next steps in discovery science. Here we address the crucial issues of open access, quality of materials, integration with public databases and sustainability of resources.