Isolation of Mycobacterium spp. in milk from cows suspected or positive to tuberculosis

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

The comparison of the Butzler medium, filtration technique and their association with isolation of Campylobacter ssp

Do total de 622 amostras de fezes, sendo 200 de suínos, 220 de bezerros e 202 de cães, Campylobacter foi isolado em 178 amostras. Destas, o agente foi identificado em 64 (36%) amostras, isoladas somente em meio seletivo de Butzler (MSB); em 34 (19%) o microorganismo foi isolado a partir da técnica de filtração (TF) e em 80 (45%), através de ambos os procedimentos (MSB+TF). A comparação entre as proporções de positividade, pelas diferentes técnicas, revelou significância (c2 = 9.184; p > 0,001) sendo a MSB (36%) a mais eficiente comparada com a TF (19%) O uso associado de ambos os procedimentos proporcionou a positividade mais elevada de isolamento com 45%.

Isolation and HPLC quantitative analysis of flavonoid glycosides from Brazilian beverages (Maytenus ilicifolia and M-aquifolium)

Aqueous infusions of Brazilian Maytenus leaves are used as beverages, foodstuffs, and phytomedicines. Previously, we isolated two new flavonoid tetrasaccharides from the infusion of Maytenus aquifolium leaves that showed antiulcer activity. In this investigation a new flavonoid tetrasaccharide, kaempferol-3-O-alpha -L-rhamnopyranosyl (1-6)-O-[alpha -L-arabinopyranosyl (1 -->3)-O-alpha -L-rhamnopyranosyl (1-2)]-O-beta -D-galactopyranoside (3), was isolated, together with kaempferol tri- and disaccharides and quercetin trisaccharides from the aqueous infusion of Maytenus ilicifolia leaves. All structures were elucidated by ES-MS and NMR spectroscopic methods. The quantitative analysis of the flavonoid glycosides from Maytenus ilicifolia and M. aquifolium has been performed by HPLC.

Isolation of Candida spp. from mastitic bovine milk in Brazil

The purpose of this study was to isolate yeast (Candida) from the quarter milk of cow udders from 37 dairy farms in Brazil and to identify the different species involved in mastitis. The samples were collected between October 2002 and February 2003. Two-hundred-and-sixty milk samples from cows with clinical and subclinical mastitis were examined. Milk samples were plated onto Blood agar, Mac Conkey agar and Sabouraud dextrose agar. Forty-five (17.3%) samples were positive for the genus Candida. The Candida species isolated were C. krusei (44.5%), C. rugosa (24.5%), C. albicans (8.9%), C. guilliermondii (8.9%), and others (13.2%). We also isolated Escherichia coli (26.5%), coagulase-positive Staphylococcus (25.0%), Streptococcus spp. (8.1%), Enterobacter spp. (8.1%), and other fungi (8.1%), among others.

Semi-selective culture medium for Curtobacterium flaccumfaciens pv. flaccumfaciens isolation from bean seeds

Curtobacterium wilt has become an important disease of beans in several localities in the country. Its causal agent, Curtobacterium flaccumfacciens pv. flaccumfaciens (Cff), survives and is disseminated through seeds. To date, few studies have been conducted with the objective of developing an effective and low-cost culture medium to isolate this bacterium from bean seeds, for health analysis purposes. Usually, the culture media employed for coryneform bacteria isolation contain specific carbon sources and antimicrobial products not available in the Brazilian market. A culture medium known as MSCFF was developed (peptone - 5 g, meat extract - 3 g, sucrose - 5 g, agar 15 g, skim milk powder* - 5 g. Congo red* - 0.05 g-, chlorothalonil* - 0.01 g, thiophanate methyl* - 0.01 g, nalidixic acid* - 0.01 g, nitrofurantoin* - 0.01 g. oxacillin* 0.001 g, sodium azide* - 0.001 g and distilled water q.s. 1L; *added after autoclaving the basal medium), which has the ability to inhibit growth of a large amount of saprophytic bacteria, but with low supressivity to Cff isolates. The MSCFF medium was highly effective for Cff isolation from naturally infected bean seeds and could be used for routine detection of this bacterium in bean seeds.

Isolation and in vitro hydrolysis of lentil protein fractions by trypsin

The albumin and globulin fractions from lentil seeds were isolated and characterised by gel filtration. The latter was shown to be homogeneous and the former heterogeneous on PAGE. The aminoacid analysis revealed high values of amidic amino acids for both fractions with great differences in the sulphur-containing amino acids. Native albumin, globulin and salt-soluble proteins were markedly resistant to trypsin hydrolysis compared to casein. The SDS-PAGE of native salt-soluble proteins indicated that the globulin fragments (20 to 30 kD) were slowly digested in the presence of albumin. The heating increased the hydrolysis of the proteins in the order: salt-soluble, albumin and globulin. The facilitated hydrolysis of the heated salt-soluble fraction seemed to be due to protein-protein interactions induced by heat.

Isolation and characterization of microsatellite loci in the woolly mouse opossum, Micoureus paraguayanus (Marsupialia : Didelphimorphia)

Five microsatellite loci were isolated and characterized within the woolly mouse opossum (Micoureus paraguayanus), a Neotropical marsupial, using an enrichment cloning procedure. Between four and seven alleles were detected per locus, with expected heterozygosity ranging from 0.358 to 0.560. These microsatellites should provide useful markers in a variety of genetic analyses to examine parentage, inbreeding, population structure and population dynamics in fragmented forest habitats.

Isolation and structure elucidation of two new flavonoid glycosides from the infusion of Maytenus aquifolium leaves. Evaluation of the antiulcer activity of the infusion

Droplet countercurrent chromatography and high-performance liquid chromatography fractionation of the aqueous infusion from Maytenus aquifolium Martius leaves afforded two flavonoid tetrasaccharides: quercetin 3-O-alpha-L-rhamnopyranosyl(1-->6)-O-[beta-D-glucopyranosyl(1-->3)-O-alpha-L-rhamnopranosyl( 1-->2)-O-beta-D-galactopyranoside and kaempferol 3-O-alpha-L-rhamnopyranosyl(1-->6)-O-[beta-D-glucopyranosyl( 1-->3)-O-alpha-L-rhamnopyranosy(1-->2-2)-O-beta-D-galactopyranoside. All structures were elucidated by spectroscopic methods. Pharmacological essays of the infusion showed antiulcer activity in rats.

ISOLATION AND IN-VITRO HYDROLYSIS OF GLOBULIN G1 FROM LENTILS (LENS-CULINARIS, MEDIK)

The major globulin fraction from lentil seeds was investigated with respect td in vitro hydrolysis by trypsin and chymotrypsin. Globulin was isolated by a NaCl-ascorbate extraction procedure and purified by DEAE-cellulose chromatography and gelfiltration chromatography on Sepharose CL-6B. The purity and identification of the protein were performed by PAGE. The native globulin, with a molecular weight of 375 kD, was resolved by SDS-PAGE into twelve polypeptides with molecular weights ranging from 61 to 14.5 kD. Native and heated globulin GI was hydrolyzed with trypsin and chymotrypsin. SDS-PAGE indicated that native globulin was more resistant to digestion than heated protein. Amino acid analysis of the major globulin revealed that glutamic acid was present in the largest concentration, followed by aspartic acid, arginine and leucine. As is also the case for other legumin-like globulins, lentil GI was deficient in sulfur-containing amino acids.

Isolation of Campylobacter jejuni from viscera and bile secretion of broiler chickens with diarrhea

Since chickens are important reservoirs of Campylobacter jejuni and their meat is the most frequent route of transmission for human campylobacteriosis, the purpouse of the present study was to investigate the presence of Campylobacter in viscera of chickens with diarrhea, evaluating the frequency of isolation of this microorganism from organs considered to be preferential for its isolation. A total of 107 visceral samples from chickens with diarrhea from different farms in the Ribeirao Preto region, SP, were examined for the presence of Campylobacter jejuni. The material consisted of 73 livers and 34 spleens, plus 29 bile secretion samples. The frequency of Campylobacter jejuni isolation was 54.79% for the liver samples, 35,29% for the spleens and 6,89% for the bile secretion samples. The data suggest that, under the conditions of the present study, the liver may be the organ of choice for the isolation of Campylobacter in the presence of diarrhea and liver involvement in chickens.