Fractional extraction of compounds from grape seeds by supercritical fluid extraction and analysis for antimicrobial and agrochemical activities

White grape seeds were subjected to sequential supercritical fluid extraction. By increasing the polarity of the supercritical fluid using methanol as a modifier of CO(2), it was possible to fractionate the extracted compounds. Two fractions were obtained; the first, which was obtained with pure CO(2), contained mainly fatty acids, aliphatic aldehydes, and sterols. The second fraction, obtained with methanol-modified CO(2), had phenolic compounds, mainly catechin, epicatechin, and gallic acid. The fractions were bioassayed. Antimicrobial activities were checked on human pathogens, and a high degree of activity was obtained with the lipophilic fraction. Agrochemical activities on phytopathogenic fungi and activities on the etiolated wheat coleoptile bioassay were also checked. The more polar fraction was active in the latter bioassay.     

Presence of encysted immature nematodes in a released whooping crane (Grus americana).

Numerous nematode cysts were observed throughout the mesentery and on the surface of gastrointestinal organs in a whooping crane (Grus americana) that was found dead in a central Florida marsh. Morphology of the excysted nematodes most closely resembled third-stage larvae in the order Spirurida but were not similar to any species previously reported in whooping cranes. Evidence presented suggests that the larvae may be Physocephalus sexalatus, a swine spirurid in the subfamily Ascaropsinae that is commonly found encapsulated in birds, amphibians, and reptiles. We suspect that the whooping crane may potentially serve as a transport host for this parasite.     

Prevalence of resistance to 11 antimicrobials among Campylobacter coill isolated from pigs on 80 grower-finisher farms in Ontario.

We carried out a cross-sectional study to investigate antimicrobial resistance patterns of Campylobacter coli isolated from Ontario grower-finisher pigs. From January to June 2004, 1200 samples were collected from 80 farms by obtaining a constant number (15) of fecal samples per farm. Susceptibility of the isolates to 11 antimicrobial drugs was determined by the agar-dilution technique. The overall prevalence of resistance to 1 or more antimicrobials among the isolates was 99.2%. High levels of resistance were observed for azithromycin, clindamycin, erythromycin, streptomycin, and tetracycline: 91.7%, 82.5%, 81.4%, 70.7%, and 63.7%, respectively. For sulfamethoxazole, ampicillin, and nalidixic acid, resistance was observed in 40.3%, 26.6%, and 22.7% of the isolates, respectively. Although at very low levels, resistance was observed for ciprofloxacin (a fluoroquinolone), chloramphenicol, and gentamicin: in 2.4%, 1.7%, and 0.2%, respectively. Many of the isolates (29.7%) were resistant to 5 antimicrobials, the most common being azithromycin, clindamycin, erythromycin, streptomycin, and tetracycline. Isolates from the same farm showed at least 5 patterns of resistance. Results from this study indicate high levels of resistance to the antimicrobial drugs most commonly used in the Canadian swine industry (macrolides, lincosamides, and tetracyclines) among C. coli isolated from grower-finisher pigs in Ontario. Macrolides and fluoroquinolones are the drugs most commonly used to treat severe human campylobacteriosis. Fortunately, at present, there is little resistance to fluoroquinolones among C. coli from pigs in Ontario.     

Maritime Halophyte Species from Southern Portugal as Sources of Bioactive Molecules

Extracts of five halophytes from southern Portugal (Arthrocnemum macrostachyum, Mesembryanthemum edule, Juncus acutus, Plantago coronopus and Halimione portulacoides), were studied for antioxidant, anti-inflammatory and in vitro antitumor properties. The most active extracts towards the 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical were the methanol extracts of M. edule (IC₅₀ = 0.1 mg/mL) and J. acutus (IC₅₀ = 0.4 mg/mL), and the ether extracts of J. acutus (IC₅₀ = 0.2 mg/mL) and A. macrostachyum (IC₅₀ = 0.3 mg/mL). The highest radical scavenging activity (RSA) against the 2,2′-azino-bis (3-ethylbenzothiazoline-6-sulphonic acid) (ABTS) radical was obtained in the ether extract of J. acutus (IC₅₀ = 0.4 mg/mL) and H. portulacoides (IC₅₀ = 0.9 mg/mL). The maximum total phenolic content (TPC) was found in the methanol extract of M. edule (147 mg gallic acid equivalents (GAE)/g) and in the ether extract of J. acutus (94 mg GAE/g). Significant decreases in nitric oxide (NO) production were observed after incubation of macrophages with lipopolysaccharide (LPS) and the chloroform extract of H. portulacoides (IC₅₀ = 109 µg/mL) and the hexane extract of P. coronopus (IC₅₀ = 98.0 µg/mL). High in vitro cytotoxic activity and selectivity was obtained with the ether extract of J. acutus. Juncunol was identified as the active compound and for the first time was shown to display selective in vitro cytotoxicity towards various human cancer cells.     

Evaluation of biochemical analytes in vitreous humor collected after death in West Indian manatees

OBJECTIVE: To evaluate biochemical analytes in vitreous humor collected after death in West Indian manatees (Trichechus manatus latirostris). DESIGN: Prospective study. ANIMALS: 73 West Indian manatees of various ages and both sexes. PROCEDURE: The condition of manatee carcasses submitted for routine postmortem examination was categorized as fresh, moderately autolyzed, or severely autolyzed. The eyes were removed, frozen for shipping, and thawed on arrival at the laboratory. Vitreous humor was extracted, and various biochemical analytes were measured. Values for each analyte were compared with published reference ranges for corresponding biochemical analytes in serum. Values were also compared among the carcass condition groups. RESULTS: Creatinine and urea nitrogen concentrations in vitreous humor were significantly higher in severely autolyzed carcasses than in fresh carcasses. Potassium concentrations in vitreous humor were significantly higher in moderately autolyzed carcasses than in fresh carcasses but were highly variable in severely autolyzed carcasses. These data were consistent with those reported in other species. CONCLUSIONS AND CLINICAL RELEVANCE: Results indicate that measurement of biochemical analytes in vitreous humor of manatees is feasible and has potential for assisting in the postmortem diagnosis of certain metabolic, renal, and nutritional diseases; determining severity of autolysis at time of necropsy; and determining the time of occurrence of human-induced trauma.     

Perineal analgesic actions of epidural clonidine in cattle

Objective To determine the analgesic, sedative, motor, cardiac and respiratory effects of epidural clonidine in cattle. Study design Prospective randomized study. Animal population Six healthy male cattle weighing between 236 and 365 kg. Methods To investigate the effect of epidural clonidine, the animals received 2 and 3 µg kg^?1 of clonidine diluted to 8 mL with 0.9% saline. Two treatments were utilized as controls. The animals from the first control treatment received 2% lidocaine (0.4 mg kg^?1) and those from the second received an equal volume of 0.9% saline. Each animal received each treatment in random order. Evaluations of analgesia, sedation, muscle relaxation, heart rate, respiratory rate and rectal temperature were obtained at 0 (basal), 2, 5, 10, 15 and 30 minutes after epidural injection, and then at 30‐minute intervals until loss of analgesia occurred. All the animals received a standard noxious stimulus consisting of needle insertion into the skin and deep muscle; a 4‐point scale was used to score the response. A second scale was used to score sedation and a third for muscle relaxation. Results Both doses of clonidine were effective in producing analgesia of the tail, perineum, and upper hindlimb. Complete analgesia was present before (mean ± SE = 9 ± 4 vs. 19 ± 9 minutes) and lasted longer (311 ± 33 vs. 192 ± 27 minutes) for the 3 µg kg^?1versus the 2 µg kg^?1 dose, respectively. A dose‐dependent sedative effect of clonidine was also observed, with a peak effect between 60 and 180 minutes. No effects on heart or respiratory rates were observed with either dose of clonidine. Conclusions Epidural administration of 2 and 3 µg kg^?1 of clonidine in cattle in this study provided bilateral perineal analgesia/anesthesia with a dose‐dependent onset and duration of action. Clinical relevance Further studies are required to determine whether the analgesia is sufficient for surgery.     

Contrasting effects of heat shock during in vitro maturation on development of in vitro‐fertilized and parthenogenetic bovine embryos

This study investigated the influence of heat shock during in vitro maturation on embryo development following in vitro fertilization (IVF) or parthenogenesis (Part). Immature bovine cumulus–oocyte complexes were exposed to heat shock (41.0°C) during the first 12 hr of in vitro maturation (IVM), followed by 12 hr at 38.5°C. Control group consisted of in vitro maturation for 24 hr at 38.5°C. Oocytes were in vitro‐fertilized or activated with ionomycin and cultured in vitro for 192 hr post‐in vitro insemination or parthenogenetic activation (hpia). There was an interaction (p < .01) between temperature of IVM and method of oocyte activation (IVF or Part) for cleavage at 48 hpia. Heat shock had a negative impact (p < .01) on cleavage of IVF embryos, whereas no (p > .05) effect was found in the Part embryos. Embryo development towards blastocyst stage at 168 and 192 hpia decreased in both IVF and Part embryos derived from heat‐shocked oocytes. Heat shock increased (p < .05) the apoptotic index in Part blastocysts, but no effect (p > .05) was found in IVF counterparts. Heat shock also down‐regulated the expression of AQP3 (p < .01) and up‐regulated the expression of HSP70.1 (p < .01) in Part blastocysts, whereas it down‐regulated the expression of ATP1A1 (p < .05) in IVF blastocysts. In conclusion, the effects of heat shock during IVM on early embryo cleavage and blastocyst apoptosis are influenced by the method of oocyte activation and expression of some genes can be disturbed in embryos derived from heat‐shocked oocytes.     

First description of a natural infection with spleen and kidney necrosis virus in zebrafish

Zebrafish has become a popular research model in the last years, and several diseases affecting zebrafish research facilities have been reported. However, only one case of naturally occurring viral infections was described for this species. In 2015, infectious spleen and kidney necrosis virus (ISKNV) was detected in zebrafish from a research facility in Spain. Affected fish showed lethargy, loss of appetite, abnormal swimming, distention of the coelomic cavity and, in the most severe cases, respiratory distress, pale gills and petechial haemorrhages at the base of fins. Cytomegaly was the most relevant histopathological finding in organs and tissues, sometimes associated to degenerative and necrotic changes. ISKNV belongs to the relatively newly defined genus Megalocytivirus, family Iridoviridae, comprising large, icosahedral cytoplasmic DNA viruses. This is the first case of naturally occurring Megalocytivirus infection in zebrafish research facilities, associated with morbidity. The virus has been identified based on both pathologic and genetic evidence, to better understand the pathogenesis of the infection in zebrafish and the phylogenetic relationship with other iridoviruses. Given the ability of megalocytiviruses to cross‐species boundaries, it seems necessary to implement stringent biosecurity practices as these infections may invalidate experimental data and have major impact on laboratory and cultured fish.