Distribution and habitat use patterns of the endangered Central American clouded oncilla (Leopardus pardinoides oncilla) in Costa Rica.

Montane cloud forests are highly threatened ecosystems that are vulnerable to climate change. These complex habitats harbor many species that suffer the negative consequences of this global phenomenon, such as shifts in their distribution and habitat use. The Central American clouded oncilla (Leopardus pardinoides oncilla) is the smallest and most endangered wild cat in Mesoamerica and is primarily reported in cloud forests throughout its distribution. The species is poorly understood, with no studies conducted in Central America assessing its habitat preferences. To bridge this knowledge gap, we sampled two mountain ranges in Costa Rica with camera traps and conducted an occupancy analysis to understand the anthropogenic and environmental features that influence oncilla habitat use within them. Additionally, we conducted spatial predictions of habitat use across its northern and southern range in Costa Rica to identify priority conservation areas for the species. We found that Central American clouded oncilla habitat use is driven primarily by environmental factors. Our results showed that oncillas select habitats with denser tree cover at high elevations, closer to permanent water sources, which may provide them with high prey density and a favorable habitat structure for their survival. Spatial predictions identified two main regions as conservation priority areas where threat mitigation efforts and monitoring should be implemented: the Caribbean slope of the Talamanca mountains, and the Arenal-Monteverde forest complex. The occupancy modeling approach turned out to be very useful to assess the spatial associations of the species with the environment and mapping the conservation priority areas. Future research and mitigation actions should focus on potential threats that could negatively impact Central American clouded oncilla populations and habitat use, including the role of mesopredators and feral species.

Ecological modeling, biogeography, and phenotypic analyses setting the tiger cats' hyperdimensional niches reveal a new species.

Recently, the tiger-cat species complex was split into Leopardus tigrinus and Leopardus guttulus, along with other proposed schemes. We performed a detailed analysis integrating ecological modeling, biogeography, and phenotype of the four originally recognized subspecies-tigrinus, oncilla, pardinoides, guttulus-and presented a new multidimensional niche depiction of the species. Species distribution models used > 1400 records from museums and photographs, all checked for species accuracy. Morphological data were obtained from institutional/personal archives. Spotting patterns were established by integrating museum and photographic/camera-trap records. Principal component analysis showed three clearly distinct groups, with the Central American specimens (oncilla) clustering entirely within those of the Andes, namely the pardinoides group of the cloud forests of the southern Central-American and Andean mountain chains (clouded tiger-cat); the tigrinus group of the savannas of the Guiana Shield and central/northeastern Brazil (savanna tiger-cat); and the guttulus group in the lowland forests of the Atlantic Forest domain (Atlantic Forest tiger-cat). This scheme is supported by recent genetic analyses. All species displayed different spotting patterns, with some significant differences in body measurements/proportions. The new distribution presented alarming reductions from the historic range of - 50.4% to - 68.2%. This multidimensional approach revealed a new species of the elusive and threatened tiger-cat complex.

Methods used by dental practice-based research network dentists to diagnose dental caries.

OBJECTIVES: To (1) identify the methods that dentists in The Dental Practice-based Research Network (DPBRN) use to diagnose dental caries; (2) quantify their frequency of use, and (3) test the hypothesis that certain dentist and dental practice characteristics are significantly associated with their use. METHODS: A questionnaire about methods used for caries diagnosis was sent to DPBRN dentists who reported doing some restorative dentistry; 522 dentists participated. Questions included the use of dental radiographs, the dental explorer, laser fluorescence, air-drying and fiber-optic devices and magnification as used when diagnosing primary, secondary/recurrent or non-specific caries lesions. Variations on the frequency of their use were tested using multivariate analysis and Bonferroni tests. RESULTS: Overall, the dental explorer was the instrument most commonly used to detect primary occlusal caries and caries at the margins of existing restorations. In contrast, laser fluorescence was rarely used to help diagnose occlusal primary caries. For proximal caries, radiographs were used to help diagnose 75%-100% of lesions by 96% of the DPBRN dentists. Dentists who use radiographs most often to assess proximal surfaces of posterior teeth were significantly more likely to also report providing a higher percentage of patients with individualized caries prevention (p = .040) and seeing a higher percentage of pediatric patients (p = .001). CONCLUSION: The use of specific diagnostic methods varied substantially. The dental explorer and radiographs are still the most commonly used diagnostic methods.

Mapping Ligand Interactions of Bromodomains BRD4 and ATAD2 with FragLites and PepLites─Halogenated Probes of Druglike and Peptide-like Molecular Interactions.

The development of ligands for biological targets is critically dependent on the identification of sites on proteins that bind molecules with high affinity. A set of compounds, called FragLites, can identify such sites, along with the interactions required to gain affinity, by X-ray crystallography. We demonstrate the utility of FragLites in mapping the binding sites of bromodomain proteins BRD4 and ATAD2 and demonstrate that FragLite mapping is comparable to a full fragment screen in identifying ligand binding sites and key interactions. We extend the FragLite set with analogous compounds derived from amino acids (termed PepLites) that mimic the interactions of peptides. The output of the FragLite maps is shown to enable the development of ligands with leadlike potency. This work establishes the use of FragLite and PepLite screening at an early stage in ligand discovery allowing the rapid assessment of tractability of protein targets and informing downstream hit-finding.

Combination therapy with Pretarget CC49 radioimmunotherapy and gemcitabine prolongs tumor doubling time in a murine xenograft model of colon cancer more effectively than either monotherapy.

Pretarget radioimmunotherapy (RIT) is a multistep strategy for cancer therapy designed to reduce nontarget organ exposure by uncoupling the tumor targeting moiety from the radioactive ligand. Using this approach, we and others have demonstrated objective responses to therapy among patients with non-Hodgkin's lymphoma, with less hematological toxicity than is typically seen at equivalent doses of conventional RIT in the same patient population. In the present study, we show that combination therapy with gemcitabine (200 mg/kg on days -1 and +1) and Pretarget RIT (400 micro Ci (90)Y-labeled DOTA-biotin on day +1) is superior to Pretarget monotherapy (400 or 800 micro Ci (90)Y) as well as to gemcitabine monotherapy in nude mice bearing established human LS174T colon cancer xenografts. For the targeting moiety, we used a murine anti-TAG-72 (CC49) single-chain Fv-streptavidin (scFvSA) fusion protein that has been shown to be safe and well-tolerated in humans. The median number of days to tumor volume doubling in the gemcitabine-only studies (200 mg/kg) was 10.4 +/- 5.5 days; in the Pretarget 400 micro Ci dose-only studies, tumor doubling time was 6.7 +/- 4.9 days; and in combination therapy studies, it was 23.9 +/- 7.2 days (P

Weather and Prey Predict Mammals' Visitation to Water.

Throughout many arid lands of Africa, Australia and the United States, wildlife agencies provide water year-round for increasing game populations and enhancing biodiversity, despite concerns that water provisioning may favor species more dependent on water, increase predation, and reduce biodiversity. In part, understanding the effects of water provisioning requires identifying why and when animals visit water. Employing this information, by matching water provisioning with use by target species, could assist wildlife management objectives while mitigating unintended consequences of year-round watering regimes. Therefore, we examined if weather variables (maximum temperature, relative humidity [RH], vapor pressure deficit [VPD], long and short-term precipitation) and predator-prey relationships (i.e., prey presence) predicted water visitation by 9 mammals. We modeled visitation as recorded by trail cameras at Sevilleta National Wildlife Refuge, New Mexico, USA (June 2009 to September 2014) using generalized linear modeling. For 3 native ungulates, elk (Cervus Canadensis), mule deer (Odocoileus hemionus), and pronghorn (Antilocapra americana), less long-term precipitation and higher maximum temperatures increased visitation, including RH for mule deer. Less long-term precipitation and higher VPD increased oryx (Oryx gazella) and desert cottontail rabbits (Sylvilagus audubonii) visitation. Long-term precipitation, with RH or VPD, predicted visitation for black-tailed jackrabbits (Lepus californicus). Standardized model coefficients demonstrated that the amount of long-term precipitation influenced herbivore visitation most. Weather (especially maximum temperature) and prey (cottontails and jackrabbits) predicted bobcat (Lynx rufus) visitation. Mule deer visitation had the largest influence on coyote (Canis latrans) visitation. Puma (Puma concolor) visitation was solely predicted by prey visitation (elk, mule deer, oryx). Most ungulate visitation peaked during May and June. Coyote, elk and puma visitation was relatively consistent throughout the year. Within the diel-period, activity patterns for predators corresponded with prey. Year-round water management may favor species with consistent use throughout the year, and facilitate predation. Providing water only during periods of high use by target species may moderate unwanted biological costs.

The importance and benefits of species.

Humans depend on biodiversity in myriad ways, yet species are being rapidly lost due to human activities. The ecosystem services approach to conservation tries to establish the value that society derives from the natural world such that the true cost of proposed development actions becomes apparent to decision makers. Species are an integral component of ecosystems, and the value they provide in terms of services should be a standard part of ecosystem assessments. However, assessing the value of species is difficult and will always remain incomplete. Some of the most difficult species' benefits to assess are those that accrue unexpectedly or are wholly unanticipated. In this review, we consider recent examples from a wide variety of species and a diverse set of ecosystem services that illustrate this point and support the application of the precautionary principle to decisions affecting the natural world.

Single-chain Fv-streptavidin substantially improved therapeutic index in multistep targeting directed at disialoganglioside GD2.

UNLABELLED: Multistep targeting can improve the therapeutic index of antibody-based targeting, particularly relevant to pediatric tumors where acute toxicity and late effects of treatment are major concerns. Neuroblastoma is uniquely suited for such investigations because of its abundance of surface ganglioside GD2. METHODS: 5F11scFv (scFv = single-chain variable fragment) was constructed from the variable regions of the heavy (V(H)) and kappa-light (V(L)) chain complementary DNA (cDNA) of anti-GD2 IgM hybridoma 5F11 and ligated to full-length streptavidin cDNA for expression in Escherichia coli. Purified 5F11-scFv-streptavidin (5F11-scFv-SA) was a homotetramer and showed comparable avidity to 5F11 IgM and a 30-fold improvement over monomeric scFv. Biodistribution of 5F11-scFv-SA was studied in nude mice xenografted with neuroblastoma LAN-1. Twenty-four hours after intravenous injection of 300-900 microg 5F11-scFv-SA, 150-450 microg of a thiogalactoside-containing clearing agent, (Gal-NAc)(16)-alpha-S-C(5)H(10)-NH-LC-N-Me-biotin (molecular weight, 8652), were administered intravenously, followed by approximately 2.5 microg (1.85-3.7 MBq) (111)In-1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid-biotin ((111)In-DOTA-biotin) intravenously 4 h later and clocked as time 0. RESULTS: Tumor uptake (percentage of injected dose per gram [%ID/g]) at 2 h was 7 %ID/g and decayed with a half-life of 72 h, whereas blood %ID/g rapidly decreased to <1/500 of that of tumor after the first 24 h. The tumor-to-nontumor (T/NT) ratio at 72 h was high (median, 106; range, 3.4 [kidney] to 1660 [blood]). When the area under the radioactivity curve was computed, the T/NT organ ratio was favorable (4.8 for kidney and 162 for blood). When human and murine tumors were surveyed, the T/NT ratio of (111)In-DOTA-biotin uptake correlated with their levels of GD2 expression as assayed by flow cytometry. Biotinylated polypeptides (bovine serum albumin and vasointestinal peptides) achieved selective tumor targeting when the multistep strategy was applied. CONCLUSION: Improvement in the T/NT ratio using pretargeting strategy may increase the efficacy and safety of scFv-based approaches in cancer therapy. Additionally, since biotinylated polypeptides can be rendered tumor selective, a large repertoire of agents can potentially be explored.

Nested species subsets, gaps, and discrepancy.

The nested-subset hypothesis of Patterson and Atmar states that species composition on islands with less species richness is a proper subset of those on islands with greater species richness. The sum of species absences, referred to as gaps, was suggested as a metric for nestedness, and null models have been used to test whether or not island species exhibited nestedness. Simberloff and Martin stated that finding examples of non-nested faunas was difficult. We revisit previous analyses of nested faunas and introduce a new metric we call "discrepancy" which we recommend as a measure for nestedness. We also recommend that the sample spaces conserve both row sums (number of species per site) and column sums (number of sites per species) derived from the incidence matrix. We compare our results to previous analyses.

Null matrices and the analysis of species co-occurrences.

Patterns in species occurrences on islands have been analyzed by several authors. At issue is the number of non-occurring pairs of species (also known as checkerboards). Previous authors have suggested that if the number of checkerboards differs from what is expected by chance, then island communities might have been structured by competition. Investigators have pursued this problem by first generating random (or null) matrices and then testing a metric derived from the collection of null matrices against the metric calculated from the actual species co-occurrence matrix. The random matrices were constrained by requiring the number of species on each island, and the number of islands on which each species occurred to be equal to their observed values. We show that results from previous studies are generally flawed. We present a fast, efficient algorithm to generate null matrices for any set of fixed row and column sums, and propose a modification of a previously proposed metric as a test statistic. We evaluated the efficacy of our construction method for null creation and our metric using incidence matrices from the avifauna of Vanuatu (formerly New Hebrides).

Mild and ligand-free Pd(II)-catalyzed conjugate additions to hindered γ-substituted cyclohexenones.

Ligand-free cationic Pd(II) catalyst with NaNO3 as an additive is a highly active catalytic system for conjugate additions to sterically hindered γ-substituted cyclohexenones. More challenging γγ- and ßγ-substrates also react well to produce products with quaternary centers in good dr. The conjugate additions occur in a diastereoselective fashion under mild, practical and air-stable conditions, using readily available commercial reagents.

The molecular deposition of transgenically modified starch in the starch granule as imaged by functional microscopy.

The molecular deposition of starch extracted from normal plants and transgenically modified potato lines was investigated using a combination of light microscopy, environmental scanning electron microscopy (ESEM) and confocal laser scanning microscopy (CLSM). ESEM permitted the detailed (10 nm) topographical analysis of starch granules in their hydrated state. CLSM could reveal internal molar deposition patterns of starch molecules. This was achieved by equimolar labelling of each starch molecule using the aminofluorophore 8-amino-1,3,6-pyrenetrisulfonic acid (APTS). Starch extracted from tubers with low amylose contents (suppressed granule bound starch synthase, GBSS) showed very little APTS fluorescence and starch granules with low molecular weight amylopectin and/or high amylose contents showed high fluorescence. Growth ring structures were sharper in granules with normal or high amylose contents. High amylose granules showed a relatively even distribution in fluorescence while normal and low amylose granules had an intense fluorescence in the hilum indicating a high concentration of amylose in the centre of the granule. Antisense of the starch phosphorylating enzyme (GWD) resulted in low molecular weight amylopectin and small fissures in the granules. Starch granules with suppressed starch branching enzyme (SBE) had severe cracks and rough surfaces. Relationships between starch molecular structure, nano-scale crystalline arrangements and topographical-morphological features were estimated and discussed.

Streptavidin in antibody pretargeting. 4. Site-directed mutation provides evidence that both arginine and lysine residues are involved in kidney localization.

The in vivo application of the protein streptavidin is limited by its propensity to localize to kidney, particularly when it is used as a carrier of radionuclides in Targeted Radionuclide Therapy. Our previous studies demonstrated that modification of recombinant "core" streptavidin (rSAv) by reaction of lysine residues with succinic anhydride and arginine residues with 1,2-cyclohexanedione dramatically decreases the kidney concentrations over that obtained with wild-type rSAv. In this investigation, we explored the role of lysine and arginine residues in kidney localization further by evaluating site-directed mutants of rSAv. In the five mutants studied, the four lysine residues found in each subunit of rSAv were replaced (independently) with an alanine (K80A, K121A, K132A, K134A), and a specific arginine was replaced with a histidine (R59H). The rSAv mutants were prepared from a "core" rSAv construct produced by expression in E. coli that had 124 amino acids (residues 13-136). Another rSAv construct that had 127 amino acids (residues 13-139), used in most of our previous studies, was also included for comparison. As an additional comparison, succinylated rSAv was prepared and evaluated. The rSAv proteins were radioiodinated and injected into athymic mice that were on a biotin-free diet for 5-7 days prior, and biodistribution data were obtained (for most proteins) at 1, 4, 24, and 48 h postinjection. The data obtained show large differences in kidney localizations of the wild-type rSAv and some rSAv mutants. The largest difference in the kidney concentration was noted for the rSAv-K134A mutant (1.90 +/- 0.22%ID/g; 24 h pi) as compared to the wild-type rSAv (31.83 +/-5.26%ID/g) at the same time point. The concentration of rSAv-K134A mutant in kidney was slightly lower than that obtained with succinylated rSAv. At the 24 h time point, the kidney concentrations of the rSAv-R59H mutant (8.95 +/- 2.94%ID/g) and the rSAv-K121A mutant (11.86 +/- 1.61%ID/g) were lower than wild-type rSAv, but the rSAv mutants rSAv-K80A (27.95 +/- 1.82%ID/g) and rSAv-K132A (32.50 +/- 10.09%ID/g) were essentially the same. The data suggests that specific lysine and arginine residues are involved in kidney localization. Possible mechanisms for the observed kidney localization are discussed.