Volatile Organic Compounds,Indole, and Biogenic Amines Assessment in Two Mediterranean Irciniidae (Porifera,Demospongiae)

Solid phase microextraction (SPME) coupled to gas chromatography-mass spectrometry (GC-MS) was employed for the headspace determination of the volatile organic fraction emitted by two of the most common Mediterranean demosponges, Ircinia variabilis and Sarcotragus spinosulus, and of indole and some biogenic amines released by sponges in an aqueous medium. A total of 50/30 µm divinylbenzene/carboxen/polydimethylsiloxane and 75 µm carboxen/polydimethylsiloxane fibers were used for the headspace extraction of low molecular weight sulfur compounds from a hermetically sealed vial containing sponge fragments, while the direct immersion determination of indole and biogenic amines was performed. The biogenic amines were extracted after in-solution derivatization with isobutyl chloroformate. All analytical parameters (linearity, limits of detection, and quantification, precision, and recovery) were evaluated for indole and biogenic amines. SPME-GC-MS proved to be a reliable means of highlighting the differences between molecules released by different sponges, principally responsible for their smell. The combined approaches allowed the identification of several volatile compounds in the headspace and other molecules released by the sponges in an aqueous medium, including indole and the BAs cadaverine, histamine, isobutylamine, isopentylamine, propylamine, 2-phenylethylamine, putrescine and tryptamine. The results obtained represent a further contribution to the picture of odoriferous molecules secreted by sponges.     

Canine visceral leishmaniasis in urban and rural areas of Northeast Brazil

The purpose of this study was to determine the clinical and laboratory profiles of canine leishmaniasis in two distinct areas. Dogs from urban and rural areas were examined. The population studied in the metropolitan area included 54 dogs. Of these, 20 (37%) animals did not present with any signs suggestive of visceral leishmaniasis (VL). Among these, only eight were confirmed negative by ELISA (rK39 and CE) and 12 dogs, clinically negative for leishmaniasis, were seropositive by ELISA (rK39 and CE). Thinness, conjunctivitis and onychogryphosis were the most frequent clinical signs in the urban areas, followed by crusty lesions, alopecia, ulcerated lesions, hyperkeratosis and exfoliation. In the metropolitan area human VL cases occurred mainly in 1991, 1992, 1999 and 2000. In the rural areas the ELISA rK39 test detected a seroprevalence of 11.3% and ELISA CE (Leishmania crude extract) of 20.6%. Thirty-nine dogs were examined 6 months after the first visit. Serological exams using rK39 antigen showed seroconversion of only one dog, whereas Leishmania CE showed seroconversion of 13 (33.4%) dogs. In this rural environment 83.3% of the positive dogs were asymptomatic. Lutzomyia intermedia and Lu. longipalpis were the most predominant sandfly vector species. Amastigotes were identified in spleen and liver fragments of symptomatic necropsied animals. PCR amplification of DNA isolated from promastigote culture indicated that the species was Leishmania chagasi. This finding suggests that delayed diagnosis and euthanasia of potentially infectious animals may occur with an increased transmission risk to sandflies and subsequently to humans.     

A Comparison of 18Oδ Composition of Water Extracted from Suction Lysimeters, Centrifugation, and Azeotropic Distillation

The representativeness of soil pore water extracted by suction lysimeters in ground-water monitoring studies is a problem that often confounds interpretation of measured data. Current soil water sampling techniques cannot delineate from which soil volume a pore water sample is extracted, neither macroscopic, microscopic, or preferential flowpath. This research was undertaken to compare δ¹⁸O and Br^? values of extracted suction lysimeters samples from intact soil cores with samples obtained by the direct extraction methods of centrifugation and azeotropic distillation. Also, the study was concerned with determining what portion of soil pore water is sampled by each method and explaining differences in concentrations of the extracted water from each method to allow a determination of the accuracy and viability of the three methods of extraction. Intact soil cores (30 cm diameter by 40 cm height) were extracted from two different sites. Site 1 was rapid infiltration basin number 50, near Altamonte Springs in Seminole County, Florida. Site 2 was the Missouri Management System Evaluation Area (MSEA) near Centralia in Boone County, Missouri. Isotopically (¹⁸Oδ) labeled water and bromide concentrations within water samples taken by suction lysimeters was compared with samples obtained by methods of centrifugation and azeotropic distillation. The ¹⁸Oδ water was analyzed by mass spectrometry while bromide concentration, applied in the form of KBr was measured using standard IC procedures. Water collected by centrifugation and azeotropic distillation data were about 0.25‰ more negative than that collected by suction lysimeter values from a sandy soil and about 2–7‰ more negative from a well structured soil. Results indicate that the majority of soil water in well-structured soil is strongly bound to soil grain surfaces and is not easily sampled by suction lysimeters. In cases where a sufficient volume of water has passed through the soil profile and displaced previous pore water, suction lysimeters will collect a representative sample of soil pore water from the sampled depth interval. It is suggested that for stable isotope studies monitoring precipitation and soil water, suction lysimeter be installed at shallow depths (10 cm). Samples should also be coordinated with precipitation events. The data also suggest that each extraction method samples a separate component of soil-pore water. Centrifugation can be used with success, particularly for efficient sampling of large areas. Azeotropic distillation is more appropriate when strict qualitative and quantitative data on sorption desorption, and various types of kinetic studies may be needed.     

Genetic diversity and structure of Ethiopian,Yemen and Brazilian C<Emphasis Type="Italic">offea arabica</Emphasis> L. accessions using microsatellites markers

Genetic diversity among 115 coffee accessions from the Coffea Germplasm Collection of IAC was assessed using SSR markers. The germplasm represents 73 accessions of Coffea arabica derived from spontaneous and subspontaneous plants in Ethiopia and Eritrea, species center of origin and diversity, 13 commercial cultivars of C. arabica developed by the Breeding Program of IAC, 1 accession of C. arabica cv. ‘Geisha’, 13 accessions of C. arabica from Yemen, 5 accessions of C. eugenioides, 4 accessions of C. racemosa and 6 accessions of C. canephora. Genetic analysis was performed using average number of alleles per locus (A), proportion of polymorphic loci (P), Shannon’s genetic index (H′ and G′_ST) and clustering analysis. All evaluated species were distinguished by a cluster analysis based on Jaccard’s coefficient. Differentiation between the cultivated plants of C. arabica and accessions derived from spontaneous and subspontaneous plants was observed. Spontaneous and subspontaneous accessions from Ethiopia were separated according to the geographical origin: east and west of the Great Rift Valley. Cultivated plants showed a low genetic diversity with a division in two groups: accessions from Yemen (H′=0,028) and Brazilian commercial cultivars (H′=0,030). The results agreed with previously reported narrow genetic basis of cultivated plants of C. arabica and supported the hypotheses about domestication of the species. This study also showed a significant genetic diversity among accessions from Ethiopia and Eritrea present in the Germplasm Collection of IAC. This diversity is specially observed in accessions from Sidamo (H′=0,143), Kaffa (H′=0,142) and Illubabor (H′=0,147) indicating their importance as source of genetic variability for coffee breeding programs.     

Microbial community development and unseen diversity recovery in inoculated sterile soil

Soil is considered as one of the most biodiverse environments on Earth; yet, the taxonomy, occurrence, and role of its different microbial populations are largely unknown. Here, two sterilized soils (from England and Italy) were inoculated with a subsample of their initial microbial communities and/or those from the other soil to study their microbial community evolution. This approach compared two driving factors (original community and soil physico-chemical characteristics) for microbial community definition. After 2 months of incubation and based on metagenomic datasets, the two inoculated communities (from an English grassland and an Italian forest) possessed similar functional and taxonomical structures when inoculated in the same sterile soil. For example, the newly colonized Italian soil was dominated by Actinobacteria related organisms (>66 % of the detected community) with a functional distribution independent of the inoculated soil origin. In addition, some of the organisms that dominated the different inoculated communities after 2 months were similar for a given sterile soil whether they came from the English grassland or the Italian forest, and they had not been detected in the original microbial community from either soil. Thus, similar microorganisms with low representation from the two distinct communities emerged in each sterilized soil, thus increasing the microbial diversity recovered from the microbial community of the donor soil. So far, these observations support the idea that different temperate soil microbial communities have different evenness due to environmental physico-chemical variations, yet have similar community composition (richness), and thus develop similarly when colonizing the same habitat.     

Evaluation of transcutaneous Doppler ultrasonography for the measurement of blood flow in the femoral artery of pigs

OBJECTIVE: To compare measurements of blood flow in the common femoral artery obtained by duplex Doppler ultrasonography (DDU) and a reference ultrasonic transit-time flow (TTF) method and to examine the impact of Doppler spectral waveform measurement techniques on volumetric estimates. ANIMALS: 5 healthy female pigs. PROCEDURES: Femoral arterial blood flow was measured simultaneously in anesthetized pigs by use of a TTF probe (left femoral artery) and transcutaneous DDU (right femoral artery). A range of flow states was induced pharmacologically by using xylazine, bradykinin, dobutamine, and isoflurane. Volumetric blood flow was calculated from DDU waveforms, using the product of the flow velocity integral (FVI), the cross-sectional vessel area, and heart rate. Three calculations of FVI were obtained by manually tracing the Doppler spectral envelopes at the outer envelope, the modal, and the inner envelope of the spectral dispersion pattern. Data analysis included calculation of Pearson correlation coefficients and Bland-Altman limits of agreement. RESULTS: Blood flow measured by DDU was more closely correlated with TTF measurements when the modal or inner envelope tracing method was used (r, 0.76 and 0.78; limits of agreement, -100 to 54.2 and -48.5 to 770 mL/min, respectively). Limits of agreement for the outer envelope tracing method were -238.5 to 64 mL/min. CONCLUSIONS AND CLINICAL RELEVANCE: Transcutaneous DDU is a reliable noninvasive technique for measuring blood flow in the femoral artery of pigs over a range of flow states. Tracing the inner envelope of the Doppler spectral dispersion pattern provided the best estimate of blood flow in this study.     

Longitudinal study of bovine rotavirus group A in newborn calves from vaccinated and unvaccinated dairy herds

Reports of rotavirus excretion in calves usually result from cross-sectional studies, and in face of the conflicting results regarding protection of calves born to vaccinated dams against diarrhea, the aim of the present study was to evaluate rotavirus excretion in dairy calves born to vaccinated or unvaccinated dams, to identify the genotypes of bovine rotavirus group A (RVA) strains isolated from these animals as well as to investigate characteristics of the disease in naturally occurring circumstances throughout the first month of life. Five hundred fifty-two fecal samples were taken from 56 calves, 28 from each farm and, in the vaccinated herd, 11/281 samples (3.91%) taken from six different calves tested positive for RVA while in the unvaccinated herd, 3/271 samples (1.11%) taken from 3 different calves tested positive. The genotyping of the VP7 genes showed 91.2% nucleotide sequence identity to G6 genotype (NCDV strain), and for the VP4 gene, strains from the vaccinated herd were 96.6% related to B223 strain, while strains from the unvaccinated herd were 88% related to P[5] genotype (UK strain). Genotypes found in this study were G6P[11] in the vaccinated herd and G6P[5] in the unvaccinated herd. All calves infected with rotavirus presented an episode of diarrhea in the first month of life, and the discrepancy between the genotypes found in the commercial vaccine (G6P[1] and G10P[11]) and the rotavirus strains circulating in both vaccinated and unvaccinated herds show the importance of keeping constant surveillance in order to avoid potential causes of vaccination failure.     

Residue study of ivermectin in plasma,milk, and mozzarella cheese following subcutaneous administration to buffalo (Bubalus bubalis)

The distribution of ivermectin in buffalo plasma and milk after administration of a single subcutaneous dose (0.2 mg kg(-)(1) b.w.) was studied. Ivermectin reached the maximal concentration in plasma (28.5 +/- 1.7 ng mL(-)(1)) and milk (23.6 +/- 2.6 ng mL(-)(1)) after 2.4 +/- 0.32 and 2.8 +/- 0.44 days, respectively. The drug showed a parallel disposition in milk and plasma, with a ratio of 1.12 +/- 0.16. Ivermectin concentrations were detected in mozzarella cheese obtained from milk collected on days 1, 3, 4, and 20 following administration. The highest values (81.4 +/- 3.26 ng g(-)(1)) were found in the cheese produced on day 3 and were 4-fold higher than those present in the milk.     

Increasing levels of supplementation for crossbred steers on pasture during the dry period of the year

The aim of this study was to evaluate the effect of increasing concentrate supplementation levels on the intake, nutrient digestibility, and performance of crossbred steers during the dry period of the year. The experiment was developed on Princesa do Mateiro farm, in the municipality of Ribeirão do Largo, located in the southwest region of Bahia State, Brazil. Forty uncastrated male crossbred (½ Holstein-Zebu) steers with an average body weight (BW) of 232.55?±?24.97 kg were distributed into four treatments in a completely randomized design with ten replicates. The animals were managed in an experimental area formed by Brachiaria brizantha cv. Marandu, in an intermittent grazing system. Treatments consisted of the following supplementation levels: 0.2% BW, with 60% crude protein (CP); 0.3% BW, with 40% CP; 0.4% BW, with 30% CP; and 0.5% BW, with 24% CP. The intakes of forage dry matter in kg/day and %BW and neutral detergent fiber corrected for ash and protein (NDFap) in %BW decreased linearly, whereas the intake of non-fibrous carbohydrates corrected for ash and protein in kg/day and average daily gain increased linearly. Therefore, the use of supplementation at 0.5% BW (24% crude protein) to provide gains of up to 0.500 kg/day is recommended for grazing steers during the post-weaning period in the dry season of the year.