Expression and characterization of the periplasmic cobalamin-binding protein of Photobacterium damselae subsp. piscicida

Cobalamin (vitamin B₁₂) is an essential cofactor in a variety of enzymatic reactions and most prokaryotes contain transport systems to import vitamin B₁₂. A gene coding for a periplasmic cobalamin-binding protein of Photobacterium damselae subsp. piscicida was identified by in silico analysis of sequences from a genomic library. The open reading frame was composed of 834 bp encoding a protein of 277 amino acids. The protein showed 61% identity with the vitamin B₁₂-binding protein precursor of P. profundum , 53% identity with the corresponding protein of Vibrio parahaemolyticus and 43% identity with the periplasmic binding protein BtuF of Escherichia coli. The expression of the native protein was investigated in P. damselae subsp. piscicida , but BtuF was weakly expressed under normal conditions. To characterize the BtuF of P. damselae subsp. piscicida , the recombinant protein was expressed with a C-terminal His₆-tag and purified; the molecular weight was estimated to be approximately 30 kDa. The protein does not contain any free thiol group, consistent with the view that the two cysteine residues are involved in a disulphide bond. The purified BtuF binds cyanocobalamin with an affinity constant of 6 ± 2 μ m .     

Water pH and Prochilodus lineatus larvae survival

The present study aimed to determine the influence of water pH on survival of curimbatá, Prochilodus lineatus, larvae. Forty-five 2-l glass aquaria with artificial aeration and no water exchange were stocked at a density of 10 larvae l⁻¹. The pH values used as treatments were 3.7 ± 0.0, 4.0 ± 0.0, 4.4 ± 0.2, 4.6 ± 0.2, 4.8 ± 0.2, 5.1 ± 0.2, 5.3 ± 0.2, 5.6 ± 0.2, 7.2 ± 0.2, 8.7 ± 0.4, 9.0 ± 0.4, 9.2 ± 0.4, 9.4 ± 0.6, 9.7 ± 0.5 and 10.0 ± 0.5 in a completely random experimental design with three repetitions. Water pH was maintained by the addition of NaOH or H₂SO₄ solutions. After 72 h of experiment, no survival was registered at pH below 4.6, 1.5% survival at pH 4.6, and about 50% survival at pH 9.4 and above. Survival rates between 70 and 80% were registered at pH 4.8–5.6 and at pH 7.2, whereas over 90% survival was registered at pH between 8.7 and 9.2.     

Bioactivity Improvement of <Emphasis Type="Italic">Olea europaea</Emphasis> Leaf Extract Biotransformed by <Emphasis Type="Italic">Wickerhamomyces anomalus</Emphasis> Enzymes

Olive leaves represent a quantitatively significant by-product of agroindustry. They are rich in phenols, mainly oleuropein, which can be hydrolyzed into several bioactive compounds, including hydroxytyrosol. In this study, water extract from olive leaves ‘Biancolilla’ was analyzed for polyphenol profile, DPPH (2,2-diphenyl-1-picrylhydrazyl) radical scavenging activity and protective effect on differentiated Caco-2 cells. The efficacy of two enzymatic treatments in promoting the release of bioactive phenols was investigated: a) enzymatic extract from Wickerhamomyces anomalus, characterized by β-glucosidase and esterase activities; b) commercial β-glucosidase. Composition and bioactivity of the resulting extracts were compared. The results showed that the yeast-treated extract presented hydroxytyrosol content and DPPH radical scavenging activity comparable to those obtained using commercial β-glucosidase; however, it was showed the additional presence of hydroxycinnamic acids. In experiments on Caco-2 cells, the leaf extracts promoted the recovery of cell membrane barrier at different minimum effective concentrations. The high specificity of W. anomalus enzymatic extract may represent an effective tool for the release of bioactive phenols from olive by-products.     

Polyostotic cortical hyperostosis in an 8-week-old cat with a 3-year follow-up

A 2 month-old female cat, mixed breed, was referred for difficulty moving and severe enlargement of the mandible and limbs. Polyostotic cortical hyperostosis was diagnosed based on diagnostic imaging and histopathological changes of the mandible and limbs. Marked cortical bone thickening was detected on radiographs and CT scan images. The diaphyses of both radii and ulnae, together with the mandibular rami and bodies, were most severely affected. The many similarities shared with the human condition, Caffey's disease, are discussed.     

Effect of Combined Non‐ionized Ammonia and Dissolved Oxygen Levels on the Survival of Juvenile Dourado,Salminus brasiliensis (Cuvier)

The aim of this study was to assess the mean lethal concentration (LC₅₀) of dissolved oxygen in high ammonia concentration and also the LC₅₀ of ammonia under hypoxic conditions for juveniles of dourado, Salminus brasiliensis. In the first experiment, the non‐ionized ammonia (NH₃) concentrations were: 0.026, 0.447, 0.612, 0.909, and 1.334 mg/L, and the dissolved oxygen concentration was maintained at approximately 1.65 mg/L. In the second experiment, the dissolved oxygen concentrations were: 1.64, 1.99, 3.33, 5.10, and 7.77 mg/L, and the non‐ionized ammonia concentration was kept at approximately 0.927 mg/L. The mean lethal concentrations of non‐ionized ammonia varied from 0.584 to 0.577 mg/L, indicating that LC₅₀ values were almost unaffected by exposure time. The estimated LC₅₀ of dissolved oxygen varied from 4.02 to 5.02 mg/L, indicating a slight increase in the mean lethal concentrations as the exposure time increased. Results from this study indicate that interaction between these two parameters increases mortality and also suggest that dourado is susceptible to the combination of high ammonia with low dissolved oxygen concentrations.     

Effects of agroecosystem landscape complexity on small mammals: a multi-species approach at different spatial scales

Landscape Ecology - The effect of landscape complexity on biodiversity is an important topic in landscape ecology, and spatial scale is key to understand true species-landscape relationships. We...     

Trypan Blue/Giemsa Staining to Assess Sperm Membrane Integrity in Salernitano Stallions and its Relationship to Pregnancy Rates

Aim of this study was to test the reliability of Trypan blue/Giemsa staining to evaluate sperm membrane integrity, acrosomal intactness and morphology in stallion to verify whether it could be applied in vitro as useful tool for sperm fertilizing ability. Fertility data on inseminated mares were collected to evaluate the relationship of sperm quality to pregnancy rates. Forty‐one ejaculates were collected from 3 stallions of Salernitano Horse Breed and evaluated for gross appearance, volume, visual motility and membrane integrity with Trypan blue/Giemsa staining and thirty‐five mares were inseminated during the breeding season from April to July. Differences among stallions were found in volume, sperm concentration (p < 0.05) and visual motility (p < 0.01). A decrease in sperm motility, concentration (p < 0.05) and total sperm number was found in June–July (p < 0.01). Live sperm with intact acrosome (LSIA) and proximal droplets (PD) were lower (p < 0.01) in June–July, while acrosome reacted sperm (ARS) percentage increased (p < 0.05). No fertility differences were found among stallions with an average fertility per cycle of 44.6% and a pregnancy rate of 68.6%. Higher percentages of LSIA were found in the ejaculates used to inseminate mares that became pregnant vs those used in mares not pregnant (p < 0.05). The significance of LSIA as test variable to verify the reliability of Trypan blue/Giemsa staining was confirmed by Receiver operating characteristic ROC analysis and the sensitivity of the test was 85% at a cut‐off value of 48% LSIA. Trypan blue‐Giemsa showed to be an accurate method that can be applied on field to evaluate sperm membrane integrity and to identify poor‐quality ejaculates.     

Survey of owned feline and canine populations in apartments from a neighbourhood in Curitiba, Brazil

Animal population estimates are essential for public health services to ensure the success of zoonoses control programmes. Canine and feline populations vary among different regions mainly because of local human income status and type of human residence. Accordingly, the present study estimated the pet population size living in apartments of a vertical neighbourhood in Curitiba, Brazil. We chose a neighbourhood with a predominance of apartment buildings. All apartment buildings were visited, and questionnaires were completed by doormen or residents. Data were obtained from 120 of 173 apartment buildings. Survey questions included the number of apartments, residents, dogs and cats. Two thousand nine hundred and sixty six apartments with a total of 7429 residents were surveyed. The number of dogs and cats was 569 and 86 respectively. Thus, the human : dog and human : cat ratios were 13.05 : 1 and 86.38 : 1. These ratios were higher than those observed in other neighbourhoods in Curitiba. The present study indicates that the number of pets from apartments may be different from houses, and different among distinct areas within the same city.     

Bioavailability of pelargonidin-3-O-glucoside and its metabolites in humans following the ingestion of strawberries with and without cream

Plasma and urine were collected over a 24 h period after the consumption by humans of 200 g of strawberries, containing 222 micromol of pelargonidin-3- O-glucoside, with and without cream. The main metabolite, a pelargonidin- O-glucuronide, reached a peak plasma concentration ( C max) of 274 +/- 24 nmol/L after 1.1 +/- 0.4 h ( t max) when only strawberries were ingested. When the strawberries were eaten with cream, the C max was not statistically different but the t max at 2.4 +/- 0.5 h was delayed significantly ( p < 0.001). The pelargonidin- O-glucuronide, along with smaller quantities of other metabolites, was also excreted in urine in quantities corresponding to ca. 1% of anthocyanin intake. The quantities excreted over the 0-24 h collection period were not influenced significantly by cream. However, the 0-2 h excretion of anthocyanin metabolites was significantly lower when the strawberries were eaten with cream, whereas the reverse occurred during with the 5-8 h excretion period. In keeping with these observations, measurement of plasma paracetamol and breath hydrogen revealed that cream delayed gastric emptying and extended mouth to cecum transit time.