A selective medium for the isolation of the acinetobacter genus bacteria

A selective and differencial medium was developed for the isolation of Acinetobacter genus bacteria. This Acinobacter Agar Medium (p.H + 7.4) contains in grams per litre: thiotone, 10; yeast extract, 3; naC1, 5; saccharose, 10; mannitol, 10; sodium citrate, 0.5; sodium desoxycholate, 0.1; crystal violet, 0.00025; phenol red, 0.04 and agar-agar 15. This medium has the advantage of inhibiting the growth of cocci and Gram-positive bacilli, by the use of sodium citrate and sodium desoxycholate associated with the crystal violet; and of differentiating the Gram-negative bacilli from the Enterobacteriaceae, through the fermentative activity upon the saccharose and/or mannitol, contrasting with the complete inactivity of the Acinetobacter genus bacteria over those substances.

Chagas' disease: selective affinity and cytotoxicity of Trypanosoma cruzi-immune lymphocytes to parasympathetic ganglion cells

The megaesophagus and megacolon endemic in South America are related , to Chagas' disease. These mega conditions are found in patients with chronic Chagas's infection, when the parasite is not demonstrable in the lesions. These are characterized by depopulation of parasympathetic ganglion cells, dilation and hypertrophy of the viscera. In the experiments described here we deminstrate a selective affinity and adherence of Trypanosoma cruzi-immune lymphocytes to myenteric, parasympathetic ganglion cells, leading to neuronolysis. None of these features are observed when non-immune lymphocytes from control rabbits are used, or when the immune lymphocytes are allowed to react with CNS neurons. This demonstration is an indication of the high degree of specificity of the destruction of parasympathetic neurons in Chagas' disease. We postulate that the T. cruzi-immune lymphocyte rejection of parasympathetic neurons, but not of CNS neurons, might be related to recognition of a cross-reacting antigenic determinant secreted only by the target neurons. In favor of this interpretation is the observation of lymphocytic infiltrates and parasympathetic ganglion cell destruction in chronic Chagas' infection in the absence of encephalitis.

Use of a selective medium with potassium tellurite to follow intestinal colonization of hospitalized patients by drug-resistant Enterobacteriaceae

Nosocomial infections are a relevant factor in complicating the recovery of patients interned for even minor causes. In attempt to determine their origin it is crucial to consider that their origin is of an endogenous nature. Looking for an acessible expression of intestinal colonization we analyzed fecal samples from 3 separate groups of hospital patients collected after different lenghts of time. For practical reasons one group was studied prospectively and two other groups (patients hospitalized for up to 7 days and patients hospitalized for more than 7 days) were compared to one another. We looked for the emergence of tellurite resistance among Enterobacteriaceae using a selective medium. MacConkey potassium tellurite (MCPT). The frequence of prospectively studied patients with tellurite resistant strains was significantly greater after 7 days of hospitalization. For the two other groups, patients with more than 7 days of hospitalization showed a significant increase of bacterial species and of strains with new antimicrobial resistance markers. High molecular weigth plasmids were detected in some of these strains. These data show that the MCPT medium is a useful tool for the investigation of bowel colonization in hospitalized patients by drug-resistant Enterobacteriaceae.

The role of the eosinophil-selective chemokine, eotaxin, in allergic and non-allergic airways inflammation

Blood eosinophilia and tissue infiltration by eosinophils are frequently observed in allergic inflammation and parasitic infections. This selective accumulation of eosinophils suggested the existence of endogenous eosinophil-selective chemoattractants. We have recently discovered a novel eosinophil-selective chemoattractant which we called eotaxin in an animal model of allergic airways disease. Eotaxin is generated in both allergic and non-allergic bronchopulmonary inflammation. The early increase in eotaxin paralled eosinophil infiltration in the lung tissue in both models. An antibody to IL-5 suppressed lung eosinophilia, correlating with an inhibition of eosinophil release from bone marrow, without affecting eotaxin generation. This suggests that endogenous IL-5 is important for eosinophil migration but does not appear to be a stimulus for eotaxin production. Constitutive levels of eotaxin observed in guinea-pig lung may be responsible for the basal lung eosinophilia observed in this species. Allergen-induced eotaxin was present mainly in the epithelium and alveolar macrophages, as detected by immunostaining. In contrast there was no upregulation of eotaxin by the epithelial cells following the injection of Sephadex beads and the alveolar macrophage and mononuclear cells surrounding the granuloma were the predominant positive staining cells. Eotaxin and related chemokines acting through the CCR3 receptor may play a major role in eosinophil recruitment in allergic inflammation and parasitic diseases and thus offer an attractive target for therapeutic intervention.

A comparison of the inhibitory activity of selective PDE4 inhibitors on eosinophil recruitment in guinea pig skin

The elevation of intracellular cyclic AMP by phosphodiesterase (PDE)4 inhibitors in eosinophils is associated with inhibition of the activation and recruitment of these cells. We have previously shown that systemic treatment with the PDE4 inhibitor rolipram effectively inhibt eosinophil migration in guinea pig skin. In the present study we compare the oral potency and efficacy of the PDE4 inhibitors rolipram, RP 73401 and CDP 840 on allergic and PAF-induced eosinophil recruitment. Rolipram and RP 73401 were equally effective and potent when given by the oral route and much more active than the PDE4 inhibitor CDP 840. We suggest that this guinea pig model of allergic and mediator-induced eosinophil recruitment is both a sensitive and simple tool to test the efficacy and potency of PDE4 inhibitors in vivo.

Effect of selective phosphodiesterase inhibitors on the rat eosinophil chemotactic response in vitro

In the present study, we have performed a comparative analysis of the effect of selective inhibitors of phosphodiesterase (PDE) type III, IV and V on eosinophil chemotaxis triggered by platelet activating factor (PAF) and leukotriene B4 (LTB4) in vitro. The effect of the analogues N6-2'-O-dibutyryladenosine 3':5' cyclic monophosphate (Bt2 cyclic AMP) and N2-2'-O- dibutyrylguanosine 3':5' cyclic monophosphate (Bt2 cyclic GMP) has also been determined. The eosinophils were obtained from the peritoneal cavity of naive Wistar rats and purified in discontinuous Percoll gradients to 85-95% purity. We observed that pre-incubation of eosinophils with the PDE type IV inhibitor rolipram suppressed the chemotactic response triggered by PAF and LTB4, in association with an increase in the intracellular levels of cyclic AMP. In contrast, neither zaprinast (type V inhibitor) nor type III inhibitors milrinone and SK&F 94836 affected the eosinophil migration. Only at the highest concentration tested did the analogue Bt2 cyclic AMP suppress the eosinophil chemotaxis, under conditions where Bt2 cyclic GMP was ineffective. We have concluded that inhibition of PDE IV, but not PDE III or V, was able to block the eosinophil chemotaxis in vitro, suggesting that the suppressive activity of selective PDE IV inhibitors on tissue eosinophil accumulation may, at least, be partially dependent on their ability to directly inhibit the eosinophil migration.

Morphological study of adult male worms of Schistosoma mansoni Sambon, 1907 by confocal laser scanning microscopy

Aiming to detail data obtained through brightfield microscopy (BM) on reproductive, excretory and digestive system, specimens of Schistosoma mansoni eight weeks old, were recovered from SW mice, stained with Langeron's carmine and analyzed under a confocal laser scanning microscope CLSM 410 (Carl Zeiss). The reproductive system presented a single and lobate testis, with intercommunications between the lobes without efferent duct. Supernumerary testicular lobe was amorphous and isolated from the normal ones. Collecting tubules (excretory ducts), followed by the excretory bladder, opening to the external media through the excretory pore, were observed at the posterior extremity of the body. In the digestive tract, a cecal swelling was noted at the junction that originates the single cecum. It was concluded that through confocal laser scanning microscopy, new interpretations of morphological structures of S. mansoni worms could be achieved, modifying adopted and current descriptions. The gonad consists of a single lobed testis, similar to that observed in some trematode species. Moreover, the same specimens can be observed either by BM or CLSM, considering that the latter causes only focal and limited damage in tissue structures.

Morphological aspects of Schistosoma mansoni adult worms isolated from nourished and undernourished mice: a comparative analysis by confocal laser scanning microscopy

Malnutrition hampers the course of schistosomiasis mansoni infection just as normal growth of adult worms. A comparative morphometric study on adult specimens (male and female) recovered from undernourished (fed with a low protein diet - regional basic diet) and nourished (rodent commercial laboratory food, NUVILAB) white mice was performed. Tomographic images and morphometric analysis of the oral and ventral suckers, reproductive system and tegument were obtained by means of confocal laser scanning microscopy. Undernourished male specimens presented smaller morphometric values (length and width) of the reproductive system (first, third and last testicular lobes) and thickness of the tegument than controls. Besides that, it was demonstrated that the dorsal surface of the male worms bears large tubercles unevenly distributed, but kept grouped and flat. At the subtegumental region, vacuolated areas were detected. It was concluded that the inadequate nutritional status of the vertebrate host has a negative influence mainly in the reproductive system and topographical somatic development of male adult Schistosoma mansoni, inducing some alterations on the structure of the parasite.

Reproductive system abnormalities in Schistosoma mansoni adult worms isolated from Nectomys squamipes (Muridae: Sigmodontinae): brightfield and confocal laser scanning microscopy analysis

Schistosoma mansoni adult worms with genital anomalies isolated from Nectomys squamipes (Muridae: Sigmodontinae) were studied by confocal laser scanning microscopy under the reflected mode. One male without testicular lobes (testicular agenesia/anorchism) and two females, one with an atrophied ovary and another with 17 uterine eggs, were identified. The absence of testicular lobes occurred in a worm presenting otherwise normal male adult characteristics: tegument, tubercles and a gynaecophoric canal with spines. In both female specimens the digestive tube showed a vacuolated appearance, and the specimen with supernumerary uterine eggs exhibited a developing miracidium and an egg with a formed shell. The area of the ventral sucker was similar in both specimens however the tegument thickness, ovary and vitelline glands of the specimen with the atrophied ovary were smaller than those of the one with supernumerary eggs. These reported anomalies in the reproductive system call attention to the need to improve our understanding of genetic regulation and the possible role of environmental influences upon trematode development.

Phenotypic plasticity in adult worms of Schistosoma mansoni (Trematoda:Schistosomatidae) evidenced by brightfield and confocal laser scanning microscopies

A comparative morphometric study was performed to identify host-induced morphological alterations in Schistosoma mansoni adult worms. A wild parasite population was obtained from a naturally infected rodent (Nectomys squamipes)and then recovered from laboratory infected C3H/He mice. Furthermore, allopatric worm populations maintained for long-term under laboratory conditions in Swiss Webster mice were passed on to N. squamipes. Suckers and genital system (testicular lobes, uterine egg, and egg spine) were analyzed by a digital system for image analysis. Confocal laser scanning microscopy (CLSM) showed details of the genital system (testicular lobes, vitelline glands, and ovary) and the tegument just below the ventral sucker. Significant morphological changes (p < 0.05) were detected in male worms in all experimental conditions, with no significant variability as assessed by CLSM. Significant changes (p < 0.05) were evident in females from the wild population related to their ovaries and vitelline glands, whereas allopatric females presented differences only in this last character. We conclude that S. mansoni worms present the phenotypic plasticity induced by modifications in the parasite's microenvironment, mainly during the first passage under laboratory conditions.

Selective PDE4 inhibitors as potent anti-inflammatory drugs for the treatment of airway diseases

Phosphodiesterases (PDEs) are responsible for the breakdown of intracellular cyclic nucleotides, from which PDE4 are the major cyclic AMP metabolizing isoenzymes found in inflammatory and immune cells. This generated greatest interest on PDE4 as a potential target to treat lung inflammatory diseases. For example, cigarette smoke-induced neutrophilia in BAL was dose and time dependently reduced by cilomilast. Beside the undesired side effects associated with the first generation of PDE4 inhibitors, the second generation of selective inhibitors such as cilomilast and roflumilast showed clinical efficacy in asthma and chronic obstrutive pulmonary diseases trials, thus re-enhancing the interest on these classes of compounds. However, the ability of PDE4 inhibitors to prevent or modulate the airway remodelling remains relatively unexplored. We demonstrated that selective PDE4 inhibitor RP 73-401 reduced matrix metalloproteinase (MMP)-9 activity and TGF-beta1 release during LPS-induced lung injury in mice and that CI-1044 inhibited the production of MMP-1 and MMP-2 from human lung fibroblasts stimulated by pro-inflammatory cytokines. Since inflammatory diseases of the bronchial airways are associated with destruction of normal tissue structure, our data suggest a therapeutic benefit for PDE4 inhibitors in tissue remodelling associated with chronic lung diseases.

Selective suppression of leukocyte recruitment in allergic inflammation

Allergic diseases result in a considerable socioeconomic burden. The incidence of allergic diseases, notably allergic asthma, has risen to high levels for reasons that are not entirely understood. With an increasing knowledge of underlying mechanisms, there is now more potential to target the inflammatory process rather than the overt symptoms. This focuses attention on the role of leukocytes especially Th2 lymphocytes that regulate allergic inflammation and effector cells where eosinophils have received much attention. Eosinophils are thought to be important based on the high numbers that are recruited to sites of allergic inflammation and the potential of these cells to effect both tissue injury and remodelling. It is hoped that future therapy will be directed towards specific leukocyte types, without overtly compromising essential host defence responses. One obvious target is leukocyte recruitment. This necessitates a detailed understanding of underlying mechanisms, particularly those involving soluble che-moattractants signals and cell-cell adhesion molecules.

A RT-PCR method for selective amplification and phenotypic characterization of all three serotypes of Sabin-related polioviruses from viral mixtures

Outbreaks caused by vaccine-derived polioviruses are challenging the final eradication of paralytic poliomyelitis. Therefore, the surveillance of the acute flaccid paralysis cases based on poliovirus isolation and characterization remains an essential activity. Due to the use of trivalent oral poliovirus vaccine (OPV), mixtures containing more than one serotype of Sabin-related polioviruses are frequently isolated from clinical samples. Because each poliovirus isolate needs to be individually analyzed, we designed polymerase chain reaction primers that can selectively distinguish and amplify a genomic segment of the three Sabin-related poliovirus serotypes present in mixtures, thus, optimizing the diagnosis and providing prompt information to support epidemiologic actions.

Plasmid-based controls to detect rpoB mutations in Mycobacterium tuberculosis by quantitative polymerase chain reaction-high-resolution melting

Quantitative polymerase chain reaction-high-resolution melting (qPCR-HRM) analysis was used to screen for mutations related to drug resistance in Mycobacterium tuberculosis. We detected the C526T and C531T mutations in the rifampicin resistance-determining region (RRDR) of the rpoB gene with qPCR-HRM using plasmid-based controls. A segment of the RRDR region from M. tuberculosis H37Rv and from strains carrying C531T or C526T mutations in the rpoB were cloned into pGEM-T vector and these vectors were used as controls in the qPCR-HRM analysis of 54 M. tuberculosis strains. The results were confirmed by DNA sequencing and showed that recombinant plasmids can replace genomic DNA as controls in the qPCR-HRM assay. Plasmids can be handled outside of biosafety level 3 facilities, reducing the risk of contamination and the cost of the assay. Plasmids have a high stability, are normally maintained in Escherichia coli and can be extracted in large amounts.

Laser therapy in pressure ulcers: evaluation by the Pressure Ulcer Scale for Healing and Nursing Outcomes Classification

AbstractOBJECTIVETo describe the pressure ulcer healing process in critically ill patients treated with conventional dressing therapy plus low-intensity laser therapy evaluated by the Pressure Ulcer Scale for Healing (PUSH) and the result of Wound Healing: Secondary Intention, according to the Nursing Outcomes Classification (NOC).METHODCase report study according to nursing process conducted with an Intensive Care Unit patient. Data were collected with an instrument containing the PUSH and the result of the NOC. In the analysis we used descriptive statistics, considering the scores obtained on the instrument.RESULTSA reduction in the size of lesions of 7cm to 1.5cm of length and 6cm to 1.1cm width, in addition to the increase of epithelial tissue and granulation, decreased secretion and odor.CONCLUSIONThere was improvement in the healing process of the lesion treated with adjuvant therapy and the use of NOC allowed a more detailed and accurate assessment than the PUSH.