Short communication: High frequency of ß-lactam-resistant Staphylococcus aureus in artisanal coalho cheese made from goat milk produced in northeastern Brazil.

Reports of ß-lactam-resistant Staphylococcus aureus in artisanal goat cheese are increasing, and this phenomenon is relevant to public health. The objective of the present study was to determine the prevalence of S. aureus strains carrying the blaZ and mecA resistance genes, as well as the genes encoding the staphylococcal enterotoxins SEA, SEB, SEC, SED, SEE, and TSST-1 in artisanal coalho cheese made from goat milk produced in northeastern Brazil. We used biochemical and molecular tests to characterize 54 S. aureus isolates found in artisanal coalho cheese collected from commercial establishments producing animal products in 11 municipalities of Pernambuco State, Brazil. A PCR analysis revealed that 42.6% (23/54) of the isolates were positive for the blaZ gene, and 7.4% (4/54) were resistant to methicillin by phenotypic testing. We did not detect mecA or any genes encoding enterotoxins. The presence of S. aureus carriers of the blaZ gene and the identification of methicillin-resistant S. aureus strains are of concern for the health of consumers of this type of cheese.

Flavonol glycosides from Costus spicatus.

Two flavonol diglycosides, tamarixetin 3-O-neohesperidoside, kaempferide 3-O-neohesperidoside and the known quercetin 3-O-neohesperidoside, together with six other known flavonoids were isolated from the leaves of Costus spicatus and their structures were elucidated by a combination of spectroscopic and chemical methods. The flavonol diglycosides were evaluated for inhibitory activity of nitric oxide production by activated macrophages (Fig. 1).

A furostanol glycoside from rhizomes of Costus spicatus.

A new furostanol glycoside was isolated from the rhizomes of Costus spicatus. Its structure was established as (3 beta,22 alpha,25R)-26-(beta -D-glucopyranosyloxy)-22-methoxyfurost-5-en-3-yl O-D-apio-beta-D-furanosyl-(1-->2)-O-[6-deoxy-alpha-L-mannopyranosy l-(1-->4)]- beta-D-glucopyranoside. The structural identification was performed using detailed analysis of 1H and 13C NMR spectra including 2D NMR spectroscopic techniques (COSY, HETCOR and COLOC) and chemical conversions.

Anti-inflammatory and immunologically active polysaccharides of Periandra mediterranea.

Three polysaccharides, glucans with mean M(r)'s of 1.5 x 10(5), 3.6 x 10(4) and 2.1 x 10(4), were isolated from dried roots of Periandra mediterranea by fractionation on Sephacryl S-300 HR and Sephadex G-25. Chemical and spectroscopic studies indicated that they have a highly branched glucan type structure composed of alpha-(1-->4) linked D-glucopyranose residues with both (3-->4) and (4-->6) branching points. The polysaccharides enhance phagocytosis in vivo, and exhibit anti-inflammatory activity.

An anti-inflammatory and immunomodulatory polysaccharide from Orbignya phalerata.

A polysaccharide, a glucan with mean M(r) of 1.0 x 10(6) (MP1), was isolated from the mesocarp of fruits of Orbignya phalerata. Chemical and spectroscopic studies indicated that MP1 has a highly branched glucan type structure composed of alpha-(1-->4) linked D-glucopyranose residues with (3-->4), (4-->6), and with (3-->6) branching points. MP1 enhanced phagocytosis in vivo and exhibited anti-inflammatory activity.

Biochanin A triglycoside from Andira inermis.

A new isoflavonol triglycoside, biochanin A 7-O-beta-D-apiofuranosyl-(1-->5)-beta-D-apiofuranosyl-(1-->6 )-beta-D- glucopyranoside (1), was isolated from Andira inermis roots in addition to the known compounds genistein 7-O-beta-D-apiofuranosyl-(1-->6)-beta-D-glucopyranoside and lanceolarin.

Immunologically active polysaccharides from Centrosema pubescens.

Two polysaccharides with mean M(r)s of 2.0 x 10(6) and 3.75 x 10(5), were isolated from powdered seeds of Centrosema pubescens by fractionation on Sephacryl S-300 HR. Chemical and spectroscopic studies indicated that they have a backbone chain composed of beta-(1-->4)-linked D-galactopyranose residues having branches composed of alpha-(1-->5)-linked L-arabinofuranose residues at position 6 of D-galactose of the backbone chain. The polysaccharides showed reticuloendothelial system-potentiating activity in a carbon clearance test.

Flavonol glycosides from leaves of Costus spiralis.

A new flavonol diglycoside, 3,5-dihydroxy-7,4'-dimethoxyflavone 3-O-neohesperidoside (1), together with four known flavonol 3-O-neohesperidosides were isolated from the leaves of Costus spiralis and their structures were elucidated by a combination of spectroscopic methods and chemical reactions.

[Leishmania (Leishmania) chagasi infection in children from an endemic area of visceral leishmaniasis in the São Luís Island-MA, Brazil]. / Infecção por Leishmania (Leishmania) chagasi em crianças de uma área endêmica de leishmaniose visceral americana na Ilha de São Luis-MA, Brasil.

A prospective study was undertaken in 648 children with less than 6 years of age in the municipality of Raposa, Maranhão, Brazil, from June 1997 to June 1998, to evaluate the characteristics of the infection by L.(L.)chagasi and verify if there is an association between malnutrition and asymptomatic infection. A standardized questionnaire was used containing socioeconomic, environmental and behavioral data. Montenegro skin reaction (IDRM) with L. amazonensis and Enzyme Linked Immunosorbent Assay (ELISA) test to detect infection, and anthropometric examination were performed. Initial and final prevalence and incidence of infection were 18.6%, 20.6% and 10.8% as measured by IDRM and 13.5%, 34.4% and 28% according to ELISA. The prevalence of chronic malnutrition was 26%. No association was detected between malnutrition and asymptomatic infection by L. (L.) chagasi. More effective control measures are needed in these areas since asymptomatic infection seems to be on the increase.

Ocorrência da infecção por Campylobacter fetus subsp. venerealis e Tritrichomonas foetus em búfalos no estado de Pernambuco, Brasil / Infection occurrence of Campylobacter fetus subsp. venerealis and Tritrichomonas foetus in buffaloes in the state of Pernambuco, Brazil

Objetivou-se com estudo determinar a ocorrência da infecção por Campylobacter fetus subsp. venerealis e Tritrichomonas foetus em búfalos no estado de Pernambuco, Brasil. Foram coletadas 133 amostras biológicas (muco cervicovaginal e raspado prepucial) de animais, procedentes de oito propriedades, de diferentes regiões do estado. O material biológico coletado foi transferido para solução salina tamponada (PBS) e, posteriormente, inoculado em meios de transporte específicos, Lander para diagnóstico de C. fetus subsp. venerealis e Diamond para T. foetus. Para o diagnóstico das infecções por Campylobacter fetus subsp. venerealis e Tritrichomonas foetus, as amostras foram submetidas à reação em cadeia da polimerase (PCR) e cultivadas em meio ágar Columbia acrescido de antibiótico e Diamond, respectivamente. Para pesquisa de C. fetus subsp. venerealis, observou-se uma ocorrência de 1,8% (2/113) de animais positivos no exame microbiológico com confirmação pela PCR. Em relação à procedência, observou-se que 100% das amostras positivas pertenciam a dois machos do mesmo rebanho. Nenhum animal foi positivo na pesquisa de T. foetus. Este é o primeiro registro da infecção por C. fetus subsp. venerealis em búfalos no Brasil. Apesar da baixa ocorrência, recomenda-se adoção de medidas de controle, com o intuito de se evitar a disseminação do agente para outros rebanhos.(AU)

The objective this study was to determine the occurrence of infection with Campylobacter fetus subsp. venerealis and Tritrichomonas foetus in buffaloes in the State of Pernambuco, Brazil. Biological samples were collected (cervico vaginal mucus and shaved prepucial) of 113 animals, coming from 8 properties in different regions of the state. The biological material collected was transferred into phosphate buffered saline (PBS) and inoculated in the specific transport, Lander for diagnosis of C. fetus subsp. venerealis and Diamond for T. fetus subsequently. For the diagnosis of infection by Campylobacter fetus subsp. venrealis and Tritrichomonas foetus the samples were submitted to Polymerase Chain Reaction (PCR) grown in Columbia agar plus antibiotics and Diamond, respectively. There was an occurrence of 1.8% (2/113) of positive animals in the microbiological examination with confirmation by PCR, for C. fetus subsp. venerealis. We observed that 100% of positive samples were from two (2) males from the same herd. No animals were positive for T. foetus. This is the first report of infection with C. fetus subsp. venerealis in buffaloes in Brazil. Despite rare occurrence, control measures are recommended in order to prevent the spread of the agent to other herds.(AU)

Occurrence of Mycoplasma bovigenitalium and Ureaplasma diversum in dairy cattle from to Pernambuco state, Brazil / Ocorrência de Mycoplasma bovigenitalium e Ureaplasma diversum em bovinos leiteiros do estado de Pernambuco, Brasil

The objective of this study was to conduct an investigation of Mycoplasma bovigenitalium and Ureaplasma diversum infections in cattle in the microregion of the Ipanema Valley, state of Pernambuco, Brazil. Vaginal swabs were collected from 355 breeding cows in reproductive age and were analyzed by multiplex PCR (mPCR) and culture. An epidemiological investigation of risk factors was performed for Mollicutes. mPCR analysis showed that, 9.29% (33/355) of the cows were positive for M. bovigenitalium and 21.69% (77/355) for U. diversum; coinfection was observed in 2.81% (10/355) of the cows. The microbiological isolation showed, 81.81% (27/33) of Mycoplasma spp. and 24.67% (19/77) of Ureaplasma spp.. The risk factors related to Mollicutes infection identified were semi-intensive breeding system (OR= 4.6), pasture rent (OR= 3.6), non-isolation of animals with reproductive disorders (OR= 3.2), and natural mounting and artificial insemination (OR= 3.5). There was a significant association between Mollicutes infection and abortions in the first gestational third (P= 0.001). This is the first record of M. bovigenitalium and U. diversum infection in cows in the semiarid region of the state of Pernambuco, Brazil. Preventive measures directed to the identified risk factors can decrease the occurrence of Mollicutes in these herds.(AU)

O objetivo deste estudo foi realizar uma investigação de Mycoplasma bovigenitalium e Ureaplasma diversum em bovinos leiteiros da microrregião do Vale do Ipanema, estado de Pernambuco, Brasil. Foram coletados suabes vaginais de 355 vacas em idade reprodutiva. As amostras foram analisadas por multiplex PCR (mPCR) e cultura. Foi realizada uma investigação dos fatores de risco para Mollicutes. Na mPCR, 9,29% (33/355) das vacas foram positivas para M. bovigenitalium e 21,69% (77/355) para U. diversum; coinfecção foi observada em 2,81% (10/355) das vacas. O isolamento microbiológico mostrou crescimento de Mycoplasma spp. em 81,81% (27/33) das amostras e em 24,67% (19/77) para Ureaplasma spp. Os fatores de risco relacionados à infecção por Mollicutes identificados foram sistema de produção semi-intensivo (OR= 4,6), aluguel de pastagem (OR= 3,6), não isolamento de animais com desordens reprodutivas (OR= 3,2) e monta natural e inseminação artificial (OR= 3,5). Houve uma associação significativa entre a infecção por Mollicutes e abortos no primeiro terço gestacional (P=0,001). Este é o primeiro relato da infecção por M. bovigenitalium e U. diversum em vacas na região semiárida do estado de Pernambuco, Brasil. As medidas preventivas direcionadas aos fatores de risco identificados podem diminuir a ocorrência de Mollicutes nesses rebanhos.(AU)

In vitro and in vivo antitumor activity of crude extracts obtained from Brazilian Chromobacterium sp isolates.

Natural products produced by microorganisms have been an important source of new substances and lead compounds for the pharmaceutical industry. Chromobacterium violaceum is a Gram-negative ß-proteobacterium, abundant in water and soil in tropical and subtropical regions and it produces violacein, a pigment that has shown great pharmaceutical potential. Crude extracts of five Brazilian isolates of Chromobacterium sp (0.25, 2.5, 25, and 250 µg/mL) were evaluated in an in vitro antitumor activity assay with nine human tumor cells. Secondary metabolic profiles were analyzed by liquid chromatography and electrospray ionization mass spectrometry resulting in the identification of violacein in all extracts, whereas FK228 was detected only in EtCE 308 and EtCE 592 extracts. AcCE and EtCE 310 extracts showed selectivity for NCI/ADR-RES cells in the in vitro assay and were evaluated in vivo in the solid Ehrlich tumor model, resulting in 50.3 and 54.6% growth inhibition, respectively. The crude extracts of Chromobacterium sp isolates showed potential and selective antitumor activities for certain human tumor cells, making them a potential source of lead compounds. Furthermore, the results suggest that other compounds, in addition to violacein, deoxyviolacein and FK228, may be involved in the antitumor effect observed.

In vitro and in vivo antitumor activity of crude extracts obtained from Brazilian Chromobacterium sp isolates

Natural products produced by microorganisms have been an important source of new substances and lead compounds for the pharmaceutical industry. Chromobacterium violaceum is a Gram-negative β-proteobacterium, abundant in water and soil in tropical and subtropical regions and it produces violacein, a pigment that has shown great pharmaceutical potential. Crude extracts of five Brazilian isolates of Chromobacterium sp (0.25, 2.5, 25, and 250 µg/mL) were evaluated in an in vitro antitumor activity assay with nine human tumor cells. Secondary metabolic profiles were analyzed by liquid chromatography and electrospray ionization mass spectrometry resulting in the identification of violacein in all extracts, whereas FK228 was detected only in EtCE 308 and EtCE 592 extracts. AcCE and EtCE 310 extracts showed selectivity for NCI/ADR-RES cells in the in vitro assay and were evaluated in vivo in the solid Ehrlich tumor model, resulting in 50.3 and 54.6% growth inhibition, respectively. The crude extracts of Chromobacterium sp isolates showed potential and selective antitumor activities for certain human tumor cells, making them a potential source of lead compounds. Furthermore, the results suggest that other compounds, in addition to violacein, deoxyviolacein and FK228, may be involved in the antitumor effect observed.

Acylated and deacylated saponins of Quillaja saponaria mixture as adjuvants for the FML-vaccine against visceral leishmaniasis.

The adjuvant of the FML-vaccine against murine and canine visceral leishmaniasis, the Riedel de Haen saponin mixture, was fractionated by ion exchange chromatography on DEAE-cellulose to afford one TLC homogeneous Quillaja saponaria Molina QS21 saponin fraction (18.0%), a mixture of two deacylsaponins (19.4%), sucrose (39.9%), sucrose and glucose (19.7%), rutin (0.8%) and quercetin (2.2%), that were identified by comparison of 1H and 13C NMR spectroscopy. The QS21 shows the typical aldehyde group in C-23 (65% equatorial) and a normonoterpene moiety acylated in C-28. The deacylsaponins show the aldehyde group but do not have the normonoterpene moiety. Balb/c mice were vaccinated with 150 microg of FML antigen of Leishmania donovani and 100 microg of each obtained fraction and further challenged by infection with 10(8) amastigotes of Leishmania chagasi. The safety analysis and the effect on humoral and cellular immune responses and in clinical signs showed that the QS21 saponin and the deacylsaponins are the most active adjuvant compounds of the Riedel the Haen saponin mixture. Both induced the highest and non-significantly different increases in DTH, CD4+ T lymphocytes in spleen, IFN-gamma in vitro, body weight gain and the most pronounced reduction of parasite burden in liver (95% for QS21 and 86% for deacylsaponins; p>0.05). While the QS21 showed mild toxicity, significant adjuvant effect on the anti-FML humoral response before and after infection, and decrease in liver relative weight, the deacylsaponins showed no toxicity, less haemolysis and antibody and DTH responses increased mainly after infection, still inducing a stronger Leishmania-specific in vitro splenocyte proliferation. Our results confirm in the Riedel de Haen saponin extract the presence of deacylsaponins normonoterpene-deprivated which are non-toxic and capable of inducing a specific and strong immunoprotective response in vaccination against murine visceral leishmaniasis.

A new steroidal saponin from the rhizomes of Costus spicatus.

A new steroidal saponin has been isolated from the rhizomes of Costus spicatus and its structure was elucidated as (3 beta, 22 alpha, 25R) -26-(beta-D-glucopyranosyloxy)-2-methoxyfurost-5-en-3-yl O-D-apio-beta-D-furanosyl-(1-->4)-O-[alpha-L-rhamnopyranosyl-(1--> 2)]- beta-D-glucopyranoside by means of IR, MS, NMR and chemical evidence.

Protective vaccination against murine visceral leishmaniasis using aldehyde-containing Quillaja saponaria sapogenins.

The presence of aldehyde groups at C-23 and C-24 of the triterpen aglycon moiety was disclosed in 1H NMR spectra of both the Riedel de Haen saponin (R) (delta 9.336) and Quillaja saponaria QuilA saponin (delta 9.348). The sign of the C-28 acylated linked moiety (delta 176) was present in both saponins, while the delta 171 at C-28 (carboxy group) corresponding to the deacylated saponin, was only detected in the QuilA preparation, indicating 50% of hydrolysis of the ester moiety, probably due to the storage in aqueous solution. The normoterpen moiety was present in both saponins (signals at delta 14-18). The chemical removal of saponin glicidic moieties gave rise to their sapogenin fractions. Their 1H NMR spectra showed the presence of two signals (delta 9.226 and 9.236) for sapogenin R and two signals (delta 9.338 and 9.352) for the QuilA sapogenin. The intensity of the signals suggested two conformational isomers of sapogenin R in the ratio 53% of equatorial aldehyde group to 47% of axial aldehyde group, and two conformational isomers of QuilA sapogenin in the ratio 76% of equatorial aldehyde group to 24% of axial aldehyde group. The chemical treatment abolished the saponin slight in vivo toxicity, reduced their hemolytic potential, did not affect their aldehyde contents, but gave rise to an enriched axial aldehyde-containing sapogenin R with enhanced potential on antibody humoral response (anti-IgM, IgG, IgG1, IgG2a, IgG2b and IgG3) and to an enriched equatorial aldehyde-containing QuilA-sapogenin that induced a mainly cellular specific immune response (increased intradermal response to leishmanial antigen and IFNgamma sera levels) and effective protection against murine infection by L. donovani (77% reduction in liver parasitic load). Our results suggest that the Riedel de Haen saponin is probably a Quillaja saponaria saponin.