Biomedical subjects
Publications and source records attributed to I Aguilar.
The susceptibility to Crotalus venom of 14 yeast and 10 mould fungal isolates was assessed. This venom was tested in a standardized well diffusion test, using 400 microg/20 microl well. The percentage of susceptibility to yeast isolates was 78.6% (> 8 mm); that for filamentous isolates was 50% (> 8 mm).
Acute renal insufficiency related to acute tubular necrosis is the most important complication caused by crotalid bite. For structural and ultrastructural studies of renal tissue, mice injected with crude venom or C. vegrandis haemorrhagic fraction, and controls were tested. Light microscopy analysis of kidneys at 24 h after injection of crude venom showed only moderate alterations such as tubular epithelia microvacuolisation. After 120 h marked glomerular and tubular capillaries congestion and interstitial oedema were observed. At 24 h after Uracoina-1 i.p. injection, intense glomerular and peritubular capillaries congestion was observed. Electron microscopic analysis of kidneys 24 h after i.p. injection of crude venom showed, capillary endothelial cell debris and pleomorphic mitochondria. Loss of interdigitations regularity, abundant dense bodies and light widening of the basal membrane were observed. Autophagic vacuoles were present as well as endothelia unfolding to the lumen and altered forms of podocytes. At 48 h, augmented endothelia without fenestrae formation with sequestration of low optical density debris inside the protrusions were noticed. At 120 h, capillary residues with loss of the endothelium were present and the basal membrane was widened. At 15 days, the number of vesicles and vacuoles in the tubules was increased and only few interdigitations were noticed. Autophagic vacuoles and mitochondrial matrix low electron density were observed. At 120 h after injection of crude venom, vascular damage with loss of capillary cell structures and collagen fibres were observed. At 24 h of haemorrhagic fraction injection, presence of autophagic vacuoles and myelinic figures were noticed.
Uracoan rattlesnake (Crotalus vegrandis) venom was subjected to chromatographic, electrophoretic, biochemical and in vivo haemorrhagic analysis. A haemorrhagic toxin (Uracoina-1) active on skin at the site of inoculation in mice was purified by Mono Q2 anion-exchange chromatography and size exclusion (SE) high-performance liquid chromatography. The purified preparation was a protein of M(r) 58,000 as revealed by sodium dodecyl sulphate--polyacrylamide gel electrophoresis under denatured conditions and with silver staining. The use of EDTA, EGTA and 1,10-phenanthroline inhibited haemorrhagic and proteolytic activities. Inhibitors of serine proteinases such as PMSF and TCLK had no effect on the haemorrhagic fraction. Uracoina-1 hydrolyses casein, hide powder azure and fibrinogen have an optimal pH of 8.2. It rapidly digests the A alpha-chain of fibrinogen. Thermal denaturation of Uracoina-1 after exposure at 60 degrees C for 15 min led to inactivation of the haemorrhagic activity. In addition, Uracoina-1 is myotoxic, lacking haemolytic, defibrinating and lethal effects. The N-terminal amino acid sequence (20 residues) was determined.
The main aim of this work was the development of a primary hepatocyte culture from Didelphis marsupialis, to determine the possible use of culture medium supernatants as a source of inhibitors of the Bothrops lanceolatus venom hemorrhagic activity. The cellular culture was carried out from isolated hepatocytes by the double perfusion technique, and digestion of the liver with collagenase and culturing the hepatocytes in a liquid media under continuous agitation at 37 degrees C in 5% CO2. The hemorrhagic activity inhibition assays were performed inoculating intradermically, a mixture of Bothrops lanceolatus venom plus a pool of liver spheroids culture supernatants, in mice. These liver Didelphis marsupialis spheroid cultures were adequate to obtain large supernatant volumes with inhibitors of hemorrhagic activity.
We have investigated the role of bacterial resistance to oxidative stress in pathogenesis. The oxyR gene from the pathogenic bacterium Erwinia chrysanthemi has been characterized. It is closely related to that found in Escherichia coli (88% overall amino acid identity). An E. chrysanthemi oxyR mutant strain was constructed by marker exchange. After induction with a sublethal dose of H2O2, this mutant was more sensitive to H2O2 and showed reduced levels of catalase and glutathione reductase activities, compared with the wild type. The oxyR mutant was unable to form individual colonies on agar plates unless catalase was added exogenously. However, it retained full virulence in potato tubers and tobacco leaves. These results suggest that the host-produced H2O2 has no direct antimicrobial effect on the interaction of E. chrysanthemi with the two plant species.
Xanthorrhizol, xanthorrhizol glycoside, and trachylobanoic acid, compounds isolated from medicinal plants that are grouped in the complex known as Cachani have been shown to inhibit the tonic contraction of rat uterus induced by: (a) depolarizing K+ solution (60 mM), (b) CaCl2 (1 mM), and (c) BAY K 8644 (0.3 microM) in a concentration-dependent manner (1-30 micrograms/mL). The inhibitory potency was displayed as follows: xanthorrhizol > xanthorrhizol glycoside > trachylobanoic acid. These results suggest that the assayed compounds might block voltage operated calcium influx in myometrial cells as they displayed a calcium-antagonistic activity. This effect is not due to peripheral receptor activation (beta 2-adrenergic, H2-histaminergic) as neither propranolol nor cimetidine modified the inhibitory effect of the compounds assayed. This is the first report showing that plants belonging to the Cachani complex may contain uterine smooth muscle bioactive substances.
Ethnotheory is suggested as a way to study the roles Mexican American female injecting drug users play. It is suggested that ethnotheory extends present theoretical models used to explain and modify female IDUs' behavior; provide insights on female IDUs' roles useful for the development of effective intervention strategies; and suggests a type of ethnographic data collection that uncovers the subject's view of reality. Three detailed ethnographic excerpts from Mexican American female IDUs illustrate how female IDUs manage the array of roles they play and how information gained through ethnotheory is useful in the design of culturally relevant interventions.
We describe the separation of an active glutamate dehydrogenase [GDH (NADP+)] enzyme from the plasma of patients with P. falciparum infection using columns of sepharose anti-GDH (NADP+) of Proteus spp. The activity of this enzyme was also detected in P. falciparum culture supernatant. The parasitic origin of this enzyme was suggested by western blot analysis using anti-P. falciparum culture supernatant and anti-whole parasite antibodies. The differential inhibition of the P. falciparum GDH (NADP+) indicates that some epitopes recognised by the antibodies in both preparations may be different. The determination of P. falciparum GDH (NADP+) activity could be developed into a specific technique for the diagnosis of falciparum malaria.
In South America rattlesnake venom activities have not been entirely characterised. Some studies have shown haemorrhagic, myotoxic, and neurotoxic effects as manifestations of envenoming in experimental animals and humans. Biopsy specimens were obtained from liver and immediately fixed in situ and observed in Hitachi H-500 and H-7100 electron microscopes. In this work the ultrastructural analysis of experimental mice liver showed hepatocytes with increased lipid droplets content and significant vacuolation in areas of their cytoplasm limiting with the Disse space. Lysosomes and altered peroxisomes exhibiting a very dense electron content were also evident. Mitochondrial pleomorphism including cup-shaped and ring-shaped mitochondria were frequently found. The cristae were scarce or absent in the majority of mitochondria observed. The rough endoplasmic reticulum showed a preferentially disposition lining the outer mitochondrial membranes. In some section glycogen particles were scarce and lipofuchsin granules could be observed. Red blood cells showed endothelial cell adherence and, in many instances, the liver sinusoids were observed plugged with aggregated red blood cells. In conclusion, using an animal model that probably correlates well with the pathological effects found in envenomed humans, we have shown the severe hepatocellular alterations caused by this venom.
In this ultrastructural study we examined skeletal muscle fibres from mice intraperitoneally inoculated with a sublethal dose of Crotalus vegrandis (rattlesnake) venom. The group of mice inoculated presented neurological symptoms characterised by respiratory failure and hind limbs paralysis. Skeletal muscle fibres showed different degrees of alterations. Most of them presented the characteristic pattern of necrosis in progress. Atrophied and hypercontracted fibres were frequently seen. Some atrophied and necrotic fibres showed several nucleoli-like bodies in the nucleus. In the atrophic and hypercontracted fibres, sarcoplasmic vacuolation and abnormal mitochondria with stacked cristae were observed. Areas of segmental necrosis were also frequently found. In connection with these altered muscle fibres, capillary abnormalities were detected. This study suggests that in envenomed mice respiratory failure symptoms may be related with muscle damage caused by Crotalus vegrandis venom components.
In this work we have demonstrated for the first time in any Venezuelan Crotalus, haemorrhagic activities that are present in the Neotropical Uracoan rattlesnake (Crotalus vegrandis) venom. This venom has been little studied, perhaps because the snake is restricted to a small habitat located in the dry savannahs of northeastern Venezuela. In our experiments Crotalus vegrandis venom caused a very evident haemorrhagic area consisting of approximately 2/3 diameter of the area caused by a positive control Bothrops lanceolatus venom. Crotalus vegrandis venom affects blood coagulation and causes intense haemorrhages. It does not clot fibrinogen, therefore it has neither thrombin-like activity which transforms fibrinogen to fibrin nor procoagulant enzymatic function which produces thrombin. On the other hand, it degrades fibrinogen making it incoagulable to thrombin. The venom, when injected in the animals, resulted in a high increase of the Partial Time of Thromboplastin (PTT) tests. It is interesting to observe that the haemorrhagic capacity in the Crotalus genus (widely distributed in the American continent) increases from south to the north, being present in North American Crotalus, a venom with wide haemorrhagic activities, and almost non-existent in most of the South American Crotalus species and subspecies.
Full-length cDNA clones of turnip mosaic virus were assembled under the control of T7 or 35S promoter. The 35S or nopaline synthase terminator signals were introduced downstream the full length cDNA controlled by 35S promoter. Both the capped in vitro transcripts from T7 controlled template, and the cDNAs from 35S controlled plasmids were infectious on Arabidopis thaliana plants according to systemically induced symptoms and to ELISA assays. The cDNAs from 35S controlled plasmids induced local lesions in Chenopodium amaranticolor and Chenopodium quinoa plants. A spontaneous silent C/T transition, giving rise to an additional SpeI restriction site, not present in the original viral RNA template, was used as a marker of the origin of infection.
The major aim of this study was to characterize a soluble Plasmodium falciparum antigen from the plasma of malaria-infected humans and Plasmodium falciparum culture supernatants, using immunoabsorbent techniques and Western blotting. An Mr 60-kDa protein was isolated from the plasma of patients with Plasmodium falciparum malaria by affinity chromatography using rabbit anti-Proteus spp GDH(NADP+) serum as ligand. This protein, present in plasma of patients with acute Plasmodium falciparum infection, in Plasmodium falciparum culture supernatants, and in immune complexes, was tested with Plasmodium falciparum malaria hyperimmune serum from patients living in hyperendemic areas and rabbit anti-Proteus spp GDH(NADP+) serum prepared in the laboratory. In this report, we describe the results of a study showing that parasite GDH(NADP+) can be used to detect the presence of Plasmodium falciparum. It appears that this technique permits the chromatographic detection of a Plasmodium falciparum excretion antigen that may be used in the production of monoclonal antibodies to improve immunodiagnostic assays for the detection of antigenemia, and opens the possibility of its use as a non-microscopic screening method.
As liver can be an important target organ in malaria, we performed an ultrastructural study of hepatic alterations in the final stage of Plasmodium berghei infection in mice. Significant hepatocyte abnormalities were found. An elevated number of cells showed mitochondria with a high electron-dense matrix and multiple changes in shape and size, alterations in the structure of Golgi complex, swelling and disorganisation of both rough and smooth-surfaced endoplasmic reticulum, differently shaped peroxisome nucleoids, and disappearance of glycogen granules. In other areas the hepatocytes were significantly altered with diminished microvilli and exhibited myelin-like figures, autophagic vacuoles, abundant lipid droplets, and swollen mitochondria in their cytoplasm. Necrotic and atrophied hepatocytes with scarce microvilli in the Disse space and biliary canaliculi could be seen. Parasitised red blood cells and parasite debris were found inside degenerated hepatocytes. Alterations were also noticed in microvasculature, including thickened endothelial cells with swollen mitochondria, lysosomes and autophagic vacuoles in their cytoplasm. Our results demonstrate that hepatocyte damage is an important finding associated with the advanced stages of P. berghei malarial infection, which may lead to liver dysfunction in this disease.
The ultrastructural study of adrenal cortex from Plasmodium berghei infected mice showed different degrees of capillary wall alterations including disruption and widening of the fenestrae, capillaries packed with parasitized erythrocytes, necrosis of cortical cells, parasitized erythrocytes outside capillaries and in some instances inside cortical cell cytoplasm. Lymphocytes were also observed in degenerated cortical cells. Our results suggest that adrenal cortex lesions may be relevant in the etiopathogenesis of severe malaria.
The complete nucleotide sequence of Chinese rape mosaic virus has been determined. The virus is a member of the tobamovirus genus of plant virus and is able to infect Arabidopsis thaliana (L.) Heynh systemically. The analysis of the sequence shows a gene array that seems to be characteristic of crucifer tobamoviruses and which is slightly different from the one most frequently found in tobamoviruses. Based on gene organization and on comparisons of sequence homologies between members of the tobamoviruses, a clustering of crucifer tobamoviruses is proposed that groups the presently known crucifer tobamovirus into two viruses with two strains each. A name change of Chinese rape mosaic virus to oilseed rape mosaic virus is proposed.
We have found an opossum serum fraction of approximately 97,000 mol. wt to be highly proficient in inactivating the haemorrhagic and proteolytic fractions of Bothrops lanceolatus venom. This antivenom substance, isolated from opossum serum or a synthetic peptide based on the aforementioned protein, would probably be useful in the medical management of Bothrops accidents.
Following a two-year cohort study of diarrhoeal diseases in children aged 0-35 months from a shanty town in Lima, 62 families were chosen for detailed observation of hygiene practices. All handwashing and defecation episodes which occurred during a 10-hour observation period (8am-6pm) were recorded. The youngest child in each family was selected as the index child for observation. Handwashing was seen on 483 occasions with 71% of the index children and 80% of mothers observed at least once. The use of clean water, soap and the thoroughness of the handwashing varied according to the purpose, with "better" behaviour observed more frequently when the person was preparing to go out. Forty-five index children (72%) were observed to defecate at least once. Infants defecated in their diapers or clothes; toddlers defecated more indiscriminately around the home area. Handwashing after defecation was rare (11% of occasions) and usually without soap. Faeces were often left accessible to children and animals (42% of occasions), especially when defecation occurred around the home/yard, and the data suggested this occurred more frequently in "higher" diarrhoea households. Stools deposited on the floor were usually just swept aside, covered with earth or eaten by dogs. Those deposited outside the home were frequently left untouched during the observation period or similarly cleared. Soiled clothes were usually left or washed separately, and stools in potties were thrown in latrines. These results suggest hygiene interventions might focus on clearance of stools from home surroundings, increased utilisation of potties and separate washing of soiled clothes.