Extracellular alpha-galactosidase from Debaryomyces hansenii UFV-1 and its use in the hydrolysis of raffinose oligosaccharides

Raffinose oligosaccharides (RO) are the factors primarily responsible for flatulence upon ingestion of soybean-derived products. ROs are hydrolyzed by alpha-galactosidases that cleave alpha-1,6-linkages of alpha-galactoside residues. The objectives of this study were the purification and characterization of extracellular alpha-galactosidase from Debaryomyces hansenii UFV-1. The enzyme purified by gel filtration and anion exchange chromatographies presented an Mr value of 60 kDa and the N-terminal amino acid sequence YENGLNLVPQMGWN. The Km values for hydrolysis of pNP alphaGal, melibiose, stachyose, and raffinose were 0.30, 2.01, 9.66, and 16 mM, respectively. The alpha-galactosidase presented absolute specificity for galactose in the alpha-position, hydrolyzing pNPGal, stachyose, raffinose, melibiose, and polymers. The enzyme was noncompetitively inhibited by galactose (Ki = 2.7 mM) and melibiose (Ki = 1.2 mM). Enzyme treatments of soy milk for 4 h at 60 degrees C reduced the amounts of stachyose and raffinose by 100%.     

Ultrastructure of Anaplasma marginale with an inclusion appendage, isolated in Minas Gerais State, Brazil

This is the first report on the occurrence and isolation of a strain of Anaplasma marginale with an inclusion appendage in Brazil. The inclusion appendage presented longitudinal electron-dense striations and did not originate directly from the body of the rickettsia but from an electron-dense complex located at the junction of the inclusion membrane and inclusion appendage. The inclusion appendage remained in the host cell after the Anaplasma inclusion appeared to be leaving the red blood cell. Other ultrastructures of this rickettsia are described and its epidemiological importance is discussed.     

Outbreak of foetal infection with bovine pestivirus in a central Queensland beef herd

OBJECTIVE: To describe a significant outbreak of foetal infection and subsequent losses due to bovine pestivirus on a 5200 ha beef breeding and fattening property in central Queensland. DESCRIPTION OF THE HERD: The affected herd consisted of 656 cows, including 269 recently purchased cows, and 221 heifers that were joined in December/January 1995/96. There were approximately 2500 cattle on the property. INVESTIGATION: Following the purchase of 269 cows in October 1995, which were mingled with the existing cow herd, losses were experienced due to foetal infection with bovine pestivirus. These losses were recorded between 1996 and 1999 as: reduced pregnancy rates, losses between pregnancy testing (midpregnancy) and branding (calves averaged 3 months-of-age), losses due to pneumonia and ill-thrift between branding and approximately 12 months-of-age, and losses due to ill-thrift and the chronic wasting form of mucosal disease thereafter. All surviving calves were tested for bovine pestivirus in 1997 at an average of 10 months. Fifty-three calves were identified as persistently infected with bovine pestivirus. A further 110 calf losses could reasonably be attributed to bovine pestivirus infection. Persistently infected cattle were always unthrifty compared to their virus negative counterparts. Only one persistently infected calf was identified, on the basis of severe ill thrift, in the 1997 birth cohort and none in 1998. CONCLUSIONS: This outbreak of foetal infection with bovine pestivirus resulted in significant production losses. These losses were recorded over the three years subsequent to the outbreak. Significant numbers of persistently infected calves were not evident among calves born in the two years after this outbreak.     

Effects of Osmotic Stress on the Growth of Etiolated Mung Bean Seedlings

The alterations in growth and water status of dark‐grown mung bean tissues following sudden osmotic stress were investigated. Photographic measurements of hypocotyl elongation revealed that stress (?0.2 to ?0.8 MPa) induced growth cessation within one minute, with resumption occurring at lower rates after a latent period; periods which tended to increase in accordance with whether the seedlings were 4, 5 or 6 days old. Tissue water status was related to growth as water and osmotic potentials were highest in the expanded root region and the lowest in the growing hypocotyl region. These rankings were not verified under osmotic stress, and values were reduced in the course of 24 h. Turgor levels were similar in all tissues and were not altered by stress. The responses were not caused by substantial water loss, which was detected only after 24 h of stress, and could have resulted from the accumulation of osmotic solutes. The results indicate that plants exposed to sudden osmotic stress, besides exhibiting rapid growth cessation, show growth resumption at lower rates and no subsequent short‐term changes in any water status measure of the growing region. Because growth resumes without further detectable turgor increments, water status alterations are unlikely to be the cause of the observed immediate growth stoppage.     

Responses of elephantgrass to nutrient solution ph levels

Seedlings of elephantgrass (Pennisetumpurpureum, Schum.), cvs. Vruckwona and Napier, were grown in aerated nutrient solution and exposed to pH levels of 4.0, 5.0, 6.0, or 7.0. After 30 days under controlled conditions, the effects on growth were evaluated. Plants exposed to pH 5.0, as compared to those under pH 4.0, showed increases in total leaf area, plant height, number of leaves (NL), and leaf, root, and stem fresh weight. No significant differences were found in stubble fresh weight, maximum individual leaf area, leaf area ratio (LAR), leaf area:plant height ratio, and leaf, root, stem, and stubble dry weight and dry matter percentage. Seedlings exposed to pH 4.0, as compared to those under pH 5.0, exhibited higher leaf:stem ratio on a fresh weight basis (FWR) and a tendency for higher leaf :stem ratio on a dry matter basis (DWR). Cultivar Vruckwona yielded superior results than Napier for most studied parameters, except for NL, LAR, and DWR (in which there were no significant differences) and for stubble length and FWR (in which Napier performed better). The results suggest that reductions below 5.0 in the nutrient solution pH cause growth inhibition, affecting root, stem, and leaf FW to a greater extent. The magnitude of such effects varies among contrasting cultivars. In turn, pH increases above 5.0 induce no significant growth reduction or promotion.     

The Influence of Powdered Coconut Water (ACP‐318®) in In Vitro Maturation of Canine Oocytes

The objective of this study was to determine the influence of powdered coconut water (ACP‐318^®) diluted in high glucose (11.0 mm ) TCM199 in the achievement of nuclear in vitro maturation (IVM) of canine oocytes. Cumulus oocyte complexes (COCs) (n = 632) were randomly allocated into three experimental groups named as group 1 (control group), group 2 (5% powdered coconut water) and group 3 (10% powdered coconut water). The percentage of meiotic resumption (MR) (GVBD to MII) was 39.1% (81/207), 50.2% (108/215) and 46.6% (98/210) for groups 1, 2 and 3 respectively (p < 0.05). There were no differences in MR rates among groups 2 and 3. The medium with ACP‐318^® slightly enhanced the nuclear maturation of canine oocytes when a comparison was established with rates of maturation exhibited by oocytes in the experimental group 1 without ACP‐318^® (p < 0.05). The results suggest that oocytes’ nuclear morphology integrity and meiosis achievement were positively influenced when exposed to high glucose TCM199 supplemented with 5% powdered coconut water. Further investigation must be performed for a better understanding of powdered coconut water influence in cellular events during IVM of dog oocytes.     

Strictosidinic acid, isolated from Psychotria myriantha Mull. Arg. (Rubiaceae), decreases serotonin levels in rat hippocampus

Psychotria is a complex genus whose neotropical species are known by the presence of glucosidic monoterpene indole alkaloids. These compounds are able to display a large range of effects on the central nervous system, such as anxiolytic, antidepressant, analgesic, and impairment of learning and memory acquisition. The aims of this study were to investigate the effects displayed by strictosidinic acid, isolated from Psychotria myriantha Mull. Arg. (Rubiaceae) leaves, on monoamine levels in rat hippocampus and on monoamine oxidase activity. A significance (p<0.01) of 83.5% reduction in 5-HT levels was observed after intra-hippocampal injection (20 μg/μl). After treatment by intraperitoneal route (10 mg/kg), a 63.4% reduction in 5-HT levels and a 67.4% reduction in DOPAC values were observed. The results indicate that strictosidinic acid seems to act on 5-HT system in rat hippocampus, possibly inhibiting precursor enzymes of 5-HT biosynthesis. The decrease verified in DOPAC levels suggests a role of strictosidinic acid in the dopaminergic transmission, probably due to an inhibition of monoamine oxidase activity, confirmed by the enzymatic assay, which demonstrated an inhibitory effect on MAO A in rat brain mitochondria.     

Survival of <Emphasis Type="Italic">Curtobacterium flaccumfaciens</Emphasis> pv. <Emphasis Type="Italic">flaccumfaciens</Emphasis> in the soil under Brazilian conditions

Mustard clubroot, caused by Plasmodiophora brassicae, is a serious disease that affects Brassica juncea var. tumida Tsen, a mustard plant that is the raw material for a traditional fermented food manufactured in the Chongqing Municipality, People’s Republic of China. To find antagonistic bacteria for P. brassicae, 124 bacteria were obtained from the rhizosphere soil of B. juncea var. tumida grown in Fuling, Chongqing. Isolates were preliminarily chosen by evaluating the inhibition rate of the P. brassicae resting spore germination. The biocontrol effects of three antagonistic bacteria against clubroot on B. juncea var. tumida were evaluated in a greenhouse experiment. B18 showed the highest control efficiency, at 63.4% in the greenhouse test. In a field trial, B18 was also effective in controlling clubroot, but only at a 49.7% efficiency rate. According to 16S rDNA sequence analysis, strain B18 had a 100% sequence similarity with type strain Zhihengliuella aestuarii DY66^T (EU939716). Based on morphological, cultural, physiological and biochemical characteristics, the DNA G + C content, polar lipids, fatty acids, cell wall analysis, as well as DNA–DNA hybridization, strain B18 was identified as Z. aestuarii B18. Thus, the isolate B18 might have a potential biocontrol application for clubroot. We report for the first time that Z. aestuarii B18 can control clubroot.     

Antigenic characterization of morphologically distinct Anaplasma marginale isolates using a panel of monoclonal antibodies

The present study, describes the antigenic characterization of a Brazilian isolate of Anaplasma marginale with appendage (tail). A panel of monoclonal antibodies (McAbs) was produced and tested by the indirect fluorescent antibody test (IFAT), ELISA and Western blotting, and used to characterize two isolates of A. marginale (one with appendage and another without appendage). Among the clones produced, eight recognized antigenic proteins, with molecular weights varying from 18.4 to 66kDa. In Western blotting, the McAb reacted against a 45kDa antigen, which was shown, by the IFAT, to be located in the tail. Immunocytochemistry confirmed the tail specificity of the monoclonal reacting against the 45kDa antigen. The panel of McAb produced has a potential use in discriminating morphologically distinct A. marginale isolates. The present study, demonstrates the occurrence of antigenic diversity among Brazilian isolates of A. marginale.