Enhanced Endothelium-dependent Relaxation Of Rat Pulmonary Artery Following β-adrenergic Overstimulation: Involvement Of The No/cgmp/vasp Pathway.

The aim of this study was to investigate whether β-adrenoceptor (β-AR) overstimulation induced by in vivo treatment with isoproterenol (ISO) alters vascular reactivity and nitric oxide (NO) production and signaling in pulmonary arteries. Vehicle or ISO (0.3mgkg(-1)day(-1)) was administered daily to male Wistar rats. After 7days, the jugular vein was cannulated to assess right ventricular (RV) systolic pressure (SP) and end diastolic pressure (EDP). The extralobar pulmonary arteries were isolated to evaluate the relaxation responses, protein expression (Western blot), NO production (diaminofluorescein-2 fluorescence), and cyclic guanosine 3',5'-monophosphate (cGMP) levels (enzyme immunoassay kit). ISO treatment induced RV hypertrophy; however, no differences in RV-SP and EDP were observed. The pulmonary arteries from the ISO-treated group showed enhanced relaxation to acetylcholine that was abolished by the NO synthase (NOS) inhibitor N(ω)-nitro-l-arginine methyl ester (l-NAME); whereas relaxation elicited by sodium nitroprusside, ISO, metaproterenol, mirabegron, or KCl was not affected by ISO treatment. ISO-treated rats displayed enhanced endothelial NOS (eNOS) and vasodilator-stimulated phosphoprotein (VASP) expression in the pulmonary arteries, while phosphodiesterase-5 protein expression decreased. ISO treatment increased NO and cGMP levels and did not induce eNOS uncoupling. The present data indicate that β-AR overactivation enhances the endothelium-dependent relaxation of pulmonary arteries. This effect was linked to an increase in eNOS-derived NO production, cGMP formation and VASP content and to a decrease in phosphodiesterase-5 expression. Therefore, elevated NO bioactivity through cGMP/VASP signaling could represent a protective mechanism of β-AR overactivation on pulmonary circulation.

Chromobacterium Amazonense Sp. Nov. Isolated From Water Samples From The Rio Negro, Amazon, Brazil.

The taxonomic position of a bacterium isolated from water samples from the Rio Negro, in Amazon, Brazil, was determined by using a polyphasic approach. The organism formed a distinct phyletic line in the Chromobacterium 16S rRNA gene tree and had chemotaxonomic and morphological properties consistent with its classification in this genus. It was found to be closely related to Chromobacterium vaccinii DSM 25150(T) (98.6 % 16S rRNA gene similarity) and shared 98.5 % 16S rRNA gene similarity with Chromobacterium piscinae LGM 3947(T). DNA-DNA relatedness studies showed that isolate CBMAI 310(T) belongs to distinct genomic species. The isolate was readily distinguished from the type strain of these species using a combination of phenotypic and chemotaxonomic properties. Thus, based on genotypic and phenotypic data, it is proposed that isolate CBMAI 310(T) (=DSM 26508(T)) be classified in the genus Chromobacterium as the type strain of a novel species, namely, Chromobacterium amazonense sp. nov.

Biofilms Of Enterococcus Faecalis And Enterococcus Faecium Isolated From The Processing Of Ricotta And The Control Of These Pathogens Through Cleaning And Sanitization Procedures.

The biofilm formation of Enterococcus faecalis and Enterococcus faecium isolated from the processing of ricotta on stainless steel coupons was evaluated, and the effect of cleaning and sanitization procedures in the control of these biofilms was determined. The formation of biofilms was observed while varying the incubation temperature (7, 25 and 39°C) and time (0, 1, 2, 4, 6 and 8days). At 7°C, the counts of E. faecalis and E. faecium were below 2log10CFU/cm(2). For the temperatures of 25 and 39°C, after 1day, the counts of E. faecalis and E. faecium were 5.75 and 6.07log10CFU/cm(2), respectively, which is characteristic of biofilm formation. The tested sanitation procedures a) acid-anionic tensioactive cleaning, b) anionic tensioactive cleaning+sanitizer and c) acid-anionic tensioactive cleaning+sanitizer were effective in removing the biofilms, reducing the counts to levels below 0.4log10CFU/cm(2). The sanitizer biguanide was the least effective, and peracetic acid was the most effective. These studies revealed the ability of enterococci to form biofilms and the importance of the cleaning step and the type of sanitizer used in sanitation processes for the effective removal of biofilms.

Bp-13 Pla2: Purification And Neuromuscular Activity Of A New Asp49 Toxin Isolated From Bothrops Pauloensis Snake Venom.

A new PLA2 (Bp-13) was purified from Bothrops pauloensis snake venom after a single chromatographic step of RP-HPLC on μ-Bondapak C-18. Amino acid analysis showed a high content of hydrophobic and basic amino acids and 14 half-cysteine residues. The N-terminal sequence showed a high degree of homology with basic Asp49 PLA2 myotoxins from other Bothrops venoms. Bp-13 showed allosteric enzymatic behavior and maximal activity at pH 8.1, 36°-45°C. Full Bp-13 PLA2 activity required Ca(2+); its PLA2 activity was inhibited by Mg(2+), Mn(2+), Sr(2+), and Cd(2+) in the presence and absence of 1 mM Ca(2+). In the mouse phrenic nerve-diaphragm (PND) preparation, the time for 50% paralysis was concentration-dependent (P < 0.05). Both the replacement of Ca(2+) by Sr(2+) and temperature lowering (24°C) inhibited the Bp-13 PLA2-induced twitch-tension blockade. Bp-13 PLA2 inhibited the contractile response to direct electrical stimulation in curarized mouse PND preparation corroborating its contracture effect. In biventer cervicis preparations, Bp-13 induced irreversible twitch-tension blockade and the KCl evoked contracture was partially, but significantly, inhibited (P > 0.05). The main effect of this new Asp49 PLA2 of Bothrops pauloensis venom is on muscle fiber sarcolemma, with avian preparation being less responsive than rodent preparation. The study enhances biochemical and pharmacological characterization of B. pauloensis venom.

Saphenous vein graft bypass in the treatment of giant cavernous sinus aneurysms: report of two cases

Two cases of giant intracavernous aneurysms treated by high flow bypass with saphenous vein graft between the external carotid artery (ECA) and branches of the middle cerebral artery (MCA) are presented. Very often these aneurysms are unclippable because they are fusiform or have a large neck. Occlusion of the internal carotid artery (ICA) is the treatment of choice in many cases. This procedure has however a high risk of brain infarction. Revascularization of the brain by extra-intracranial anastomosis between the superficial temporal artery (STA) and branches of the MCA is frequently performed. This procedure provides however a low flow bypass and brain infarction may occur. We report two cases of giant cavernous sinus aneurysms treated by high flow bypass and endovascular balloon occlusion of the ICA. Immediate high flow revascularization of MCA branches was achieved and the patients showed no ischemic events. Follow-up of 8 and 14 months after operation shows patency of the venous graft and no neurological deficits. Angiographic control examination showed complete aneurysm occlusion in both cases.

Prevalence of mutans streptococci isolated from complete dentures and their susceptibility to mouthrinses

The aims of this study were to evaluate the incidence of mutans streptococci (MS - sessile form) on complete maxillary dentures after use of a specific denture paste, and to determine the minimum inhibitory concentration (MIC) and maximum inhibitory dilution (MID) of 3 oral mouthrinses: Cepacol, Plax and Periogard. Seventy-seven complete denture wearers were randomly assigned into 2 groups, according to the product used for denture cleaning: Control group - conventional dentifrice (Kolynos-Super White); and Test group: experimental denture cleaning paste. Denture biofilm was collected at baseline and after 90 and 180 days after treatment by brushing the dentures with saline solution. After decimal serial dilution, samples were seeded onto agar sucrose bacitracin to count colonies with morphological characteristics of MS. MS identification was performed by the sugar fermentation tests. After this procedure, brain heart infusion broth (BHI) was added to oral mouthrinses (Plax, Cepacol e Periogard) and seeded on Petri dishes. The colonies were seeded using the Steers multiplier and, after the incubation, the MIC and MID of the mouthrinses were calculated. The results showed an incidence of 74.0% (n=57) of MS in the 77 complete dentures examined in the study, being 76.3% (n=29) of the Control group (conventional dentifrice) and 71.8% (28) of the Test group (experimental denture cleaning paste). In both groups, the number of positive cases for MS decreased from day 0 to day 180. In the Test group there was a slight decrease in the incidence of Streptococcus mutans 90 days after use of the experimental denture cleaning paste, which was not observed in the Control group. As regards to mouthrinses, for both groups, Periogard showed antimicrobial action with the highest dilution, followed by Cepacol and Plax. In conclusion, the incidence of MS in complete dentures was high and Periogard was the mouthrinse with the strongest antimicrobial action against MS. The experimental denture cleaning paste showed a slight action against S. mutans after 90 days of treatment.

Antimicrobial susceptibility of coagulase-negative Staphylococci isolated from mastitic cattle in Brazil

Um total de 109 cepas de Staphylococci coagulase-negativa foi isolado de leite de vacas com mastite clínica e subclínica, em 35 fazendas, situadas em nove estados brasileiros, no período de fevereiro a maio de 2005. Os isolados foram investigados em relação a susceptibilidade in vitro a diversos agentes antimicrobianos. A resistência à penicilina foi a observação mais freqüente (93,5%), seguida por sulfonamida (88,9%), novobiocina (88,6%) e ampicilina (85,3%). Todas as cepas examinadas mostraram resistência a pelo menos uma das drogas antimicrobianas testadas. Cepas apresentando resistência múltipla foram extremamente comuns, com 10,0% dos microrganismos isolados apresentando resistência a todas as drogas antimicrobianas. Os resultados obtidos indicaram que as cepas de Staphylococci coagulase-negativas, isoladas no Brasil, apresentaram um alto grau de resistência a antimicrobianos. Estes resultados são, provavelmente, uma conseqüência da pressão devida ao uso intensivo de drogas antimicrobianas.

Distribution of biotypes and leukotoxic activity of Aggregatibacter actinomycetemcomitans isolated from Brazilian patients with chronic periodontitis

Aggregatibacter actinomycetemcomitans is an important etiologic agent of the periodontitis and is associated with extra-oral infections. In this study, the detection of the ltxA gene as well as the ltx promoter region from leukotoxic A. actinomycetemcomitans isolated from 50 Brazilian patients with periodontitis and 50 healthy subjects was performed. The leukotoxic activity on HL-60 cells was also evaluated. Leukotoxic activity was determined using a trypan blue exclusion method. The 530 bp deletion in the promoter region was evaluated by PCR using a PRO primer pair. A. actinomycetemcomitans was detected by culture and directly from crude subgingival biofilm by PCR using specific primers. By culture, A. actinomycetemcomitans was detected in nine (18%) of the periodontal patients and one (2%) healthy subject. However, by PCR, this organism was detected in 44% of the periodontal patients and in 16% of the healthy subjects. It was verified a great discrepancy between PCR detection of the ltx operon promoter directly from crude subgingival biofilm and from bacterial DNA. Only one periodontal sample harbored highly leukotoxic A. actinomycetemcomitans. Moreover, biotype II was the most prevalent and no correlation between biotypes and leukotoxic activity was observed. The diversity of leukotoxin expression by A. actinomycetemcomitans suggests a role of this toxin in the pathogenesis of periodontal disease and other infectious diseases.

Risk indicators for aggressive periodontitis in an untreated isolated young population from Brazil

This study aimed to assess the prevalence of aggressive periodontitis (AgP), and to investigate the association between demographic, socioeconomic and behavioral risk indicators with AgP in an untreated and isolated young population in Southeastern Brazil. For this cross-sectional survey, 134 subjects aged 12-29 years were selected by a census. Of those eligible, 101 subjects received a full-mouth clinical examination, and were interviewed using a structured written questionnaire. Cases were defined as individuals with 4 or more teeth with attachment loss > 4 mm or > 5 mm in the age groups 12-19 and 20-29, respectively. Overall, 9.9% of the subjects presented AgP (10.3% of the 12-19-year-olds and 9.7% of the 20-29-year-olds). The only risk indicator significantly associated with AgP in this isolated population was a high proportion of sites (> 30%) presenting supragingival calculus [OR = 23.2]. Having experienced an urgency dental treatment was a protective factor for AgP [OR = 0.1]. The authors concluded that this isolated and untreated population from Brazil presented a high prevalence of AgP. Local plaque-retaining factors played a major role in the prevalence of AgP in this isolated population, and should be included in further studies evaluating this destructive periodontal disease form.

Should reproductively isolated populations of Lutzomyia longipalpis sensu lato receive taxonomically valid names?

A group of 18 research workers involved in different aspects of the biology of Lutzomyia longipalpis discussed whether or not it is important to give taxonomically valid names to populations that have been defined by biological, biochemical and molecular methods to be reproductively isolated. The type material of this medically important species has been lost and because of this it was recommended that a colony should be established from insects captured in the region of the type area and that their description should serve as the basis for future descriptions. It was pointed out that there is a lack of uniformity in the naming of closely related American sand flies and that some of the differences between populations of Lu. longipalpis are greater than those between accepted species. The majority of the participants agreed that the populations that have been defined in the literature as sibling species should be named.

Characterization of Vibrio Parahaemolyticus isolated from oysters and mussels in São Paulo, Brazil

Vibrio parahaemolyticus is a marine bacterium, responsible for gastroenteritis in humans. Most of the clinical isolates produce thermostable direct hemolysin (TDH) and TDH-related hemolysin (TRH) encoded by tdh and trh genes respectively. In this study, twenty-three V. parahaemolyticus, previously isolated from oysters and mussels were analyzed by PCR using specific primers for the 16S rRNA and virulence genes (tdh, trh and tlh) and for resistance to different classes of antibiotics and PFGE. Nineteen isolates were confirmed by PCR as V. parahaemolyticus. The tlh gene was present in 100% of isolates, the tdh gene was identified in two (10.5%) isolates, whereas the gene trh was not detected. Each isolate was resistant to at least one of the nine antimicrobials tested. Additionally, all isolates possessed the blaTEM-116 gene. The presence of this gene in V. parahaemolyticus indicates the possibility of spreading this gene in the environment. Atypical strains of V. parahaemolyticus were also detected in this study.

Antimicrobial susceptibility of coagulase-negative staphylococci isolated from meat-producing ewes with mastitis

Avaliou-se a sensibilidade antimicrobiana in vitro de 121 cepas de estafilococos coagulase-negativa isolada de leite de ovelhas Santa Inês, aos fármacos: penicilina, amoxicilina, ampicilina, estreptomicina, oxaciclina, neomicina, cefalotina, gentamicina e sulfonamida. A resistência à sulfonamida foi a mais frequente (27,3%), seguida pela estreptomicina (14,0%) e pela oxaciclina (14,0%), enquanto da gentamicina (1,6%) foi a menos frequente. Todas as cepas foram sensíveis a pelo menos um antimicrobiano, e 20,3% das cepas apresentaram resistência múltipla. Os resultados mostram a importância de Staphylococci coagulase-negativas como agentes causadores de mastite em ovinos, e o perfil de resistência múltipla indica a importância da determinação da resistência à oxaciclina como indicador da presença de ilhas de patogenicidade que contêm fatores de virulência e resistência a outros antimicrobianos que contribuem para a sobrevivência da bactéria ao tratamento.

Occurrence and molecular characterization of Cryptosporidium spp. isolated from domestic animals in a rural area surrounding Atlantic dry forest fragments in Teodoro Sampaio municipality, State of São Paulo, Brazil

The aim of this study was to assess the occurrence of Cryptosporidium in domestic animals in rural properties surrounding rain forest fragments within the municipality of Teodoro Sampaio, southeastern Brazil. Conventional sucrose flotation method followed by molecular characterization of the parasites by sequencing PCR products amplified from SSU rRNA gene were used. Stool samples were collected from domestic animals raised as pets and livestock in all rural properties surrounding three forest fragments. Samples from cattle (197), equine (63), pigs (25), sheep (11), and dogs (28) were collected from 98 rural properties. The frequency of occurrence of Cryptosporidium within each animal species was 3.0% (6/197) among cattle and 10.7% (3/28) among dogs. Cryptosporidium was not detected in stool samples from equine, sheep, and pigs. All sequences obtained from the six samples of calves showed molecular identity with Cryptosporidium andersoni while all sequences from dog samples were similar to C. canis. The frequency of occurrence of Cryptosporidium in these domestic animal species was low. The absence of C. parvum in the present study suggests that the zoonotic cycle of cryptosporidiosis may not be relevant in the region studied. The presence of Cryptosporidium species seldom described in humans may be, otherwise, important for the wild fauna as these animals are a source of infection and dissemination of this protozoan to other animal species. The impact and magnitude of infection by C. andersoni in wild ruminants and C. canis in wild canids have to be assessed in future studies to better understand the actual importance of these species in this region.

Hepatotrophic factors reduce hepatic fibrosis in rats

CONTEXT: Hepatic fibrosis occurs in response to several aggressive agents and is a predisposing factor in cirrhosis. Hepatotrophic factors were shown to stimulate liver growth and to restore the histological architecture of the liver. They also cause an improvement in liver function and accelerate the reversion of fibrosis before it progresses to cirrhosis. OBJECTIVE: To test the effects of hepatic fibrosis solution composed by amino acids, vitamins, glucose, insulin, glucagon and triiodothyronine on hepatic fibrosis in rats. METHODS: Fibrosis was induced in rats by gastric administration of dimethylnitrosamine (10 mg/kg) for 5 weeks. After liver biopsy, the rats received either hepatotrophic factors solution (40 mg/kg/day) or saline solution for 10 days by intraperitoneal injection. Blood samples and liver fragments were collected for hepatic function analysis, standard histopathology evaluation, and morphometric collagen quantification. RESULTS: Rats in the hepatotrophic factors group showed a decrease of the histopathological components of fibrosis and an increase of their hepatic mass (12.2%). There was no development of neoplasic lesions in both groups. Compared with the saline group, the hepatotrophic factors group also had a decrease of blood levels of hepatic-lesion markers (AST, ALT) and a decrease of collagen content in the portal spaces (31.6%) and perisinusoidal spaces (42.3%), as well as around the hepatic terminal vein (57.7%). Thus, hepatotrophic factors administration in the portal blood promoted a regenerative hepatic response, with an overall reduction of the volumetric density of collagen, improved hepatic function, and a general improvement in the histopathological aspects of fibrosis. CONCLUSION: Taken together, these results suggest the potential therapeutic use of this hepatotrophic factors solution to treat chronic liver diseases.

Clonal relationship among atypical enteropathogenic Escherichia coli strains isolated from different animal species and humans

Forty-nine typical and atypical enteropathogenic Escherichia coli (EPEC) strains belonging to different serotypes and isolated from humans, pets (cats and dogs), farm animals (bovines, sheep, and rabbits), and wild animals (monkeys) were investigated for virulence markers and clonal similarity by pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST). The virulence markers analyzed revealed that atypical EPEC strains isolated from animals have the potential to cause diarrhea in humans. A close clonal relationship between human and animal isolates was found by MLST and PFGE. These results indicate that these animals act as atypical EPEC reservoirs and may represent sources of infection for humans. Since humans also act as a reservoir of atypical EPEC strains, the cycle of mutual infection of atypical EPEC between animals and humans, mainly pets and their owners, cannot be ruled out since the transmission dynamics between the reservoirs are not yet clearly understood.