Clearing and dissecting insects for internal skeletal morphological research with particular reference to bees

ABSTRACT A detailed protocol for chemical clearing of bee specimens is presented. Dry specimens as well as those preserved in liquid media can be cleared using this protocol. The procedure consists of a combined use of alkaline solution (KOH or NaOH) and hydrogen peroxide (H2O2), followed by the boiling of the cleared specimens in 60–70% EtOH. Clearing is particularly useful for internal skeletal morphological research. This procedure allows for efficient study of internal projections of the exoskeleton (e.g., apodemes, furcae, phragmata, tentoria, internal ridges and sulci), but this process makes external features of the integument, as some sutures and sulci, readily available for observation as well. Upon completion of the chemical clearing process the specimens can be stored in glycerin. This procedure was developed and evaluated for the preparation of bees and other Apoidea, but modifications for use with other insect taxa should be straightforward after some experimentation on variations of timing of steps, concentration of solutions, temperatures, and the necessity of a given step. Comments on the long-term storage, morphological examination, and photodocumentation of cleared specimens are also provided.

Variability of apparently homogeneous soilscapes in São Paulo state, Brazil: II. quality of soil maps

The quality of semi-detailed (scale 1:100.000) soil maps and the utility of a taxonomically based legend were assessed by studying 33 apparently homogeneous fields with strongly weathered soils in two regions in São Paulo State: Araras and Assis. An independent data set of 395 auger sites was used to determine purity of soil mapping units and analysis of variance within and between mapping units and soil classification units. Twenty three soil profiles were studied in detail. The studied soil maps have a high purity for some legend criteria, such as B horizon type (> 90%) and soil texture class (> 80%). The purity for the "trophic character" (eutrophic, dystrophic, allic) was only 55% in Assis. It was 88% in Araras, where many soil units had been mapped as associations. In both regions, the base status of clay-textured soils was generally better than suggested by the maps. Analysis of variance showed that mapping was successful for "durable" soil characteristics such as clay content (> 80% of variance explained) and cation exchange capacity (≥ 50% of variance explained) of 0-20 and 60-80 cm layers. For soil characteristics that are easily modified by management, such as base saturation of the 0-20 cm layer, the maps had explained very little (< 15%) of the total variance in the study areas. Intermediate results were obtained for base saturation of the 60-80 cm layer (56% in Assis; 42% in Araras). Variance explained by taxonomic groupings that formed the basis for the legend of the soil maps was similar to, often even smaller than, variance explained by mapping units. The conclusion is that map boundaries have been very carefully located, but descriptions of mapping units could be improved. In future mappings, this could possibly be done at low cost by (a) bulk sampling to remove short range variation and enhance visualization of spatial patterns at distances > 100 m; (b) taking advantage of correlations between easily measured soil characteristics and chemical soil properties and, (c) unbending the link between legend criteria and a taxonomic system. The maps are well suited to obtain an impression of land suitability for high-input farming. Additional field work and data on former land use/management are necessary for the evaluation of chemical properties of surface horizons.

Computer-assisted cartography using topographic properties: precision and accuracy of local soil maps in central Mexico

Map units directly related to properties of soil-landscape are generated by local soil classes. Therefore to take into consideration the knowledge of farmers is essential to automate the procedure. The aim of this study was to map local soil classes by computer-assisted cartography (CAC), using several combinations of topographic properties produced by GIS (digital elevation model, aspect, slope, and profile curvature). A decision tree was used to find the number of topographic properties required for digital cartography of the local soil classes. The maps produced were evaluated based on the attributes of map quality defined as precision and accuracy of the CAC-based maps. The evaluation was carried out in Central Mexico using three maps of local soil classes with contrasting landscape and climatic conditions (desert, temperate, and tropical). In the three areas the precision (56 %) of the CAC maps based on elevation as topographical feature was higher than when based on slope, aspect and profile curvature. The accuracy of the maps (boundary locations) was however low (33 %), in other words, further research is required to improve this indicator.

Optimization of the sampling scheme for maps of physical and chemical properties estimated by kriging

The sampling scheme is essential in the investigation of the spatial variability of soil properties in Soil Science studies. The high costs of sampling schemes optimized with additional sampling points for each physical and chemical soil property, prevent their use in precision agriculture. The purpose of this study was to obtain an optimal sampling scheme for physical and chemical property sets and investigate its effect on the quality of soil sampling. Soil was sampled on a 42-ha area, with 206 geo-referenced points arranged in a regular grid spaced 50 m from each other, in a depth range of 0.00-0.20 m. In order to obtain an optimal sampling scheme for every physical and chemical property, a sample grid, a medium-scale variogram and the extended Spatial Simulated Annealing (SSA) method were used to minimize kriging variance. The optimization procedure was validated by constructing maps of relative improvement comparing the sample configuration before and after the process. A greater concentration of recommended points in specific areas (NW-SE direction) was observed, which also reflects a greater estimate variance at these locations. The addition of optimal samples, for specific regions, increased the accuracy up to 2 % for chemical and 1 % for physical properties. The use of a sample grid and medium-scale variogram, as previous information for the conception of additional sampling schemes, was very promising to determine the locations of these additional points for all physical and chemical soil properties, enhancing the accuracy of kriging estimates of the physical-chemical properties.

Comparison between detailed digital and conventional soil maps of an area with complex geology

Since different pedologists will draw different soil maps of a same area, it is important to compare the differences between mapping by specialists and mapping techniques, as for example currently intensively discussed Digital Soil Mapping. Four detailed soil maps (scale 1:10.000) of a 182-ha sugarcane farm in the county of Rafard, São Paulo State, Brazil, were compared. The area has a large variation of soil formation factors. The maps were drawn independently by four soil scientists and compared with a fifth map obtained by a digital soil mapping technique. All pedologists were given the same set of information. As many field expeditions and soil pits as required by each surveyor were provided to define the mapping units (MUs). For the Digital Soil Map (DSM), spectral data were extracted from Landsat 5 Thematic Mapper (TM) imagery as well as six terrain attributes from the topographic map of the area. These data were summarized by principal component analysis to generate the map designs of groups through Fuzzy K-means clustering. Field observations were made to identify the soils in the MUs and classify them according to the Brazilian Soil Classification System (BSCS). To compare the conventional and digital (DSM) soil maps, they were crossed pairwise to generate confusion matrices that were mapped. The categorical analysis at each classification level of the BSCS showed that the agreement between the maps decreased towards the lower levels of classification and the great influence of the surveyor on both the mapping and definition of MUs in the soil map. The average correspondence between the conventional and DSM maps was similar. Therefore, the method used to obtain the DSM yielded similar results to those obtained by the conventional technique, while providing additional information about the landscape of each soil, useful for applications in future surveys of similar areas.

Genetic linkage maps of 'Pêra' sweet orange and 'Cravo' mandarin with RAPD markers

The objective of this work was to construct linkage maps of 'Pêra' sweet orange [Citrus sinensis (L.) Osbeck] and 'Cravo' mandarin (Citrus reticulata Blanco) using RAPD markers and the pseudo-testcross strategy. The parents were chosen according to the resistance/susceptibility to citrus variegate chlorosis (CVC). The segregation of 176 markers was analyzed in 94 progeny of F1 hybrids, which were obtained from controlled crossings. The linkage map of 'Pêra' sweet orange had 117 markers defined by 12 linkage groups, which spanned 612.1 cM. Only six markers could not be linked to the linkage group and 48.7% of the markers showed segregation distortion. The linkage map of 'Cravo' mandarin had 51 markers defined by 12 linkage groups, which spanned 353.3 cM. Only two markers did not link to the groups and 15.7% showed segregation distortion. The construction of linkage maps is relevant to future mapping studies of the inheritance of CVC, citrus canker and leprosis.

Chemotaxonomy of three genera of the annonaceae family using self-organizing maps and 13C NMR data of diterpenes

The Annonaceae family is distributed throughout Neotropical regions of the world. In Brazil, it covers nearly all natural formations particularly Annona, Xylopia and Polyalthia and is characterized chemically by the production of sources of terpenoids (mainly diterpenes), alkaloids, steroids, polyphenols and, flavonoids. Studies from 13C NMR data of diterpenes related with their botanical occurrence were used to generate self-organizing maps (SOM). Results corroborate those in the literature obtained from morphological and molecular data for three genera and the model can be used to project other diterpenes. Therefore, the model produced can predict which genera are likely to contain a compound.

Cadastral maps of irrigated areas by center pivots in the State of Minas Gerais, using CBERS-2B/CCD satellite imaging

Medium-resolution satellite images have been widely used for the identification and quantification of irrigated areas by center pivot. These areas, which present predominantly circular forms, can be easily identified by visual analyses of these images. In addition to identifying and quantifying areas irrigated by center pivot, other information that is associated to these areas is fundamental for producing cadastral maps. The goal of this work was to generate cadastral mapping of areas irrigated by center pivots in the State of Minas Gerais, Brazil, with the purpose of supplying information on irrigated agriculture. Using the satellite CBERS2B/CCD, images were used to identify and quantify irrigated areas and then associate these areas with a database containing information about: irrigated area, perimeter, municipality, path row, basin in which the pivot is located, and the date of image acquisition.3,781 center pivots systems were identified. The smallest area irrigated was 4.6 hectares and the largest one was 192.6 hectares. The total estimated value of irrigated area was 254,875 hectares. The largest number of center pivots appeared in the municipalities of Unaí and Paracatu, with 495 and 459 systems, respectively. Cadastral mapping is a very useful tool to assist and enhance information on irrigated agriculture in the State of Minas Gerais.

Determination of management zones from normalized and standardized equivalent produtivity maps in the soybean culture

Through the site-specific management, the precision agriculture brings new techniques for the agricultural sector, as well as a larger detailing of the used methods and increase of the global efficiency of the system. The objective of this work was to analyze two techniques for definition of management zones using soybean yield maps, in a productive area handled with localized fertilization and other with conventional fertilization. The sampling area has 1.74 ha, with 128 plots with site-specific fertilization and 128 plots with conventional fertilization. The productivity data were normalized by two techniques (normalized and standardized equivalent productivity), being later classified in management zones. It can be concluded that the two methods of management zones definition had revealed to be efficient, presenting similarities in the data disposal. Due to the fact that the equivalent standardized productivity uses standard score, it contemplates a better statistics justification.

Comparison measures of maps generated by geostatistical methods

This study uses several measures derived from the error matrix for comparing two thematic maps generated with the same sample set. The reference map was generated with all the sample elements and the map set as the model was generated without the two points detected as influential by the analysis of local influence diagnostics. The data analyzed refer to the wheat productivity in an agricultural area of 13.55 ha considering a sampling grid of 50 x 50 m comprising 50 georeferenced sample elements. The comparison measures derived from the error matrix indicated that despite some similarity on the maps, they are different. The difference between the estimated production by the reference map and the actual production was of 350 kilograms. The same difference calculated with the mode map was of 50 kilograms, indicating that the study of influential points is of fundamental importance to obtain a more reliable estimative and use of measures obtained from the error matrix is a good option to make comparisons between thematic maps.

Comparison of maps of spatial variability of soil resistance to penetration constructed with and without covariables using a spatial linear model

A study about the spatial variability of data of soil resistance to penetration (RSP) was conducted at layers 0.0-0.1 m, 0.1-0.2 m and 0.2-0.3 m depth, using the statistical methods in univariate forms, i.e., using traditional geostatistics, forming thematic maps by ordinary kriging for each layer of the study. It was analyzed the RSP in layer 0.2-0.3 m depth through a spatial linear model (SLM), which considered the layers 0.0-0.1 m and 0.1-0.2 m in depth as covariable, obtaining an estimation model and a thematic map by universal kriging. The thematic maps of the RSP at layer 0.2-0.3 m depth, constructed by both methods, were compared using measures of accuracy obtained from the construction of the matrix of errors and confusion matrix. There are similarities between the thematic maps. All maps showed that the RSP is higher in the north region.

Soybean yield maps using regular and optimized sample with different configurations by simulated annealing

ABSTRACT This study aimed to compare thematic maps of soybean yield for different sampling grids, using geostatistical methods (semivariance function and kriging). The analysis was performed with soybean yield data in t ha-1 in a commercial area with regular grids with distances between points of 25x25 m, 50x50 m, 75x75 m, 100x100 m, with 549, 188, 66 and 44 sampling points respectively; and data obtained by yield monitors. Optimized sampling schemes were also generated with the algorithm called Simulated Annealing, using maximization of the overall accuracy measure as a criterion for optimization. The results showed that sample size and sample density influenced the description of the spatial distribution of soybean yield. When the sample size was increased, there was an increased efficiency of thematic maps used to describe the spatial variability of soybean yield (higher values of accuracy indices and lower values for the sum of squared estimation error). In addition, more accurate maps were obtained, especially considering the optimized sample configurations with 188 and 549 sample points.

Loss of Ovarian Function Results in Increased Loss of Skeletal Muscle in Arthritic Rats

Objective We studied the effects of loss of ovarian function (ovariectomy) onmuscle mass of gastrocnemius and themRNA levels of IGF-1, atrogin-1, MuRF-1, andmyostatin in an experimental model of rheumatoid arthritis in rats. Methods We randomly allocated 24 female Wistar rats (9 weeks, 195.3±17.4 grams) into four groups: control (CT-Sham; n = 6); rheumatoid arthritis (RA; n = 6); ovariectomy without rheumatoid arthritis (OV; n = 6); ovariectomy with rheumatoid arthritis (RAOV; n = 6). We performed the ovariectomy (OV and RAOV) or Sham (CTSham or RA) procedures at the same time, fifteen days before the rheumatoid arthritis induction. The RA and RAOV groups were immunized and then were injected with Met- BSA in the tibiotarsal joint. After 15 days of intra-articular injections the animals were euthanized. We evaluated the external manifestations of rheumatoid arthritis (perimeter joint) as well as animal weight, and food intake throughout the study. We also analyzed the cross-sectional areas (CSA) of gastrocnemius muscle fibers in 200 fibers (H&E method). In the gastrocnemius muscle, we analyzed mRNA expression by quantitative real time PCR followed by the Livak method (ΔΔCT). Results The rheumatoid arthritis induced reduction in CSA of gastrocnemius muscle fibers. The RAOV group showed a lower CSA of gastrocnemius muscle fibers compared to RA and CT-Sham groups. Skeletal muscle IGF-1 mRNA increased in arthritics and ovariectomized rats. The increased IGF-1 mRNA was higher in OV groups than in the RA and RAOV groups. Antrogin-1 mRNA also increased in the gastrocnemius muscle of arthritic and ovariectomized rats. However, the increased atrogin-1 mRNA was higher in RAOV groups than in the RA and OV groups. Gastrocnemius muscle MuRF-1 mRNA increased in the OVand RAOVgroups, but not in the RA and Shamgroups. However, the RAOV group showed higher MuRF-1 mRNA than the OV group. The myostatin gene expression was similar in all groups. Conclusion Loss of ovarian function results in increased loss of skeletal musclerelated ubiquitin ligases atrogin-1 and MuRF-1 in arthritic rats.

Standardization of metachromatic staining method of myofibrillar ATPase activity of myosin to skeletal striated muscle of mules and donkeys

This study aims at standardizing the pre-incubation and incubation pH and temperature used in the metachromatic staining method of myofibrillar ATPase activity of myosin (mATPase) used for asses and mules. Twenty four donkeys and 10 mules, seven females and three males, were used in the study. From each animal, fragments from the Gluteus medius muscle were collected and percutaneous muscle biopsy was performed using a 6.0-mm Bergström-type needle. In addition to the metachromatic staining method of mATPase, the technique of nicotinamide adenine dinucleotide tetrazolium reductase (NADH-TR) was also performed to confirm the histochemical data. The histochemical result of mATPase for acidic pre-incubation (pH=4.50) and alkaline incubation (pH=10.50), at a temperature of 37ºC, yielded the best differentiation of fibers stained with toluidine blue. Muscle fibers were identified according to the following colors: type I (oxidative, light blue), type IIA (oxidative-glycolytic, intermediate blue) and type IIX (glycolytic, dark blue). There are no reports in the literature regarding the characterization and distribution of different types of muscle fibers used by donkeys and mules when performing traction work, cargo transportation, endurance sports (horseback riding) and marching competitions. Therefore, this study is the first report on the standardization of the mATPase technique for donkeys and mules.

Infestation maps and spatial stability of main weed species in maize culture

A study on the spatial distribution of the major weeds in maize was carried out in 2007 and 2008 in a field located in Golegã (Ribatejo region, Portugal). The geo-referenced sampling focused on 150 points of a 10 x 10 m mesh covering an area of 1.5 ha, before herbicide application and before harvest. In the first year, 40 species (21 botanical families) were identified at seedling stage and only 22 during the last observation. The difference in species richness can be attributed to maize monoculture favouring reduction in species number. Three of the most representative species were selected for the spatial distribution analysis: Solanum nigrum, Chenopodium album and Echinochloa crus-galli. The three species showed an aggregated spatial pattern and spatial stability over both years, although the herbicide effect is evident in the distribution of some of them in the space. These results could be taken into account when planning site-specific treatments in maize.