Central nervous system involvement in experimental trypanosomiasis cruzi

A review of the available literature on central nervous system involvement in experimental trypanosomiasis cruzi is undertaken. From a critical analysis of 26 works on experimental infections with Trypanosoma cruzi (23 on the acute phase, 2 on the chronic phase, and one describing sequentially both phases), all supported by neuropathologic studies, it can be concluded that: 1) central nervous system involvement during the acute phase, in the form of encephalitis in multiple foci, with variable intensity of the parasitism and inflamatory changes, is frequent and well documented; 2) in animals with more severe central nervous system involvement death occurs as a result of the brain lesions or acute chagasic myocarditis, the latter being always present; 3) in animals with more discrete brain involviment death during the acute phase is due to complications not related to the nervous system, among which congestive heart failure second to acute chagasic myocarditis, a condition that is always present, regardless of whether or not the central nervous system is infected; 4) it is possible that in surviving animals that had mild encephalitis the inflammatory changes from the acute phase usually regress as the infection progress to the chronic phase.

Force-induced globule-coil transition in laminin binding protein and its role for viral-cell membrane fusion.

The specific interactions of the pairs laminin binding protein (LBP)-purified tick-borne encephalitis viral surface protein E and certain recombinant fragments of this protein, as well as West Nile viral surface protein E and certain recombinant fragments of that protein, are studied by combined methods of single-molecule dynamic force spectroscopy (SMDFS), enzyme immunoassay and optical surface waves-based biosensor measurements. The experiments were performed at neutral pH (7.4) and acid pH (5.3) conditions. The data obtained confirm the role of LBP as a cell receptor for two typical viral species of the Flavivirus genus. A comparison of these data with similar data obtained for another cell receptor of this family, namely human αVβ3 integrin, reveals that both these receptors are very important. Studying the specific interaction between the cell receptors in question and specially prepared monoclonal antibodies against them, we could show that both interaction sites involved in the process of virus-cell interaction remain intact at pH 5.3. At the same time, for these acid conditions characteristic for an endosome during flavivirus-cell membrane fusion, SMDFS data reveal the existence of a force-induced (effective already for forces as small as 30-70 pN) sharp globule-coil transition for LBP and LBP-fragments of protein E complexes. We argue that this conformational transformation, being an analog of abrupt first-order phase transition and having similarity with the famous Rayleigh hydrodynamic instability, might be indispensable for the flavivirus-cell membrane fusion process. Copyright © 2014 John Wiley & Sons, Ltd.

Biomphalaria prona (Gastropoda: Planorbidae): a morphological and biochemical study

Two samples of Biomphalaria prona (Martens, 1873) from Lake Valencia (type locality) and seven from other Venezuelan localities were studied morphologically (shell and reproductive system) and biochemically (allozyme electrophoresis). In spite of marked differences in shell characters, all of them proved indistinguishable under the anatomic and biochemical criteria. So far B. prona has been considered an endemic species, restricted to Lake Valencia. It is now demonstrated that the extralacustrine populations refered to Biomphalaria havanensis (Pfeiffer, 1839) by several authors correspond in shell characters to an extreme variant of B. prona from the Lake and really belong to the last*mentioned species. They may be regarded as the result of a process of directional selection favoring a shell phenotype other than those making up the modal class in the Lake.

Real-time PCR for type-specific identification of herpes simplex in clinical samples: evaluation of type-specific results in the context of CNS diseases.

BACKGROUND: HSV-1 and HSV-2 cause CNS infections of dissimilar clinico-pathological characteristics with prognostic and therapeutic implications. OBJECTIVES: To validate a type-specific real-time PCR that uses MGB/LNA Taqman probes and to review the virologico-clinical data of 25 eligible patients with non-neonatal CNS infections. RESULTS: This real-time PCR was evaluated against conventional PCR (26 CSF and 20 quality controls), and LightCycler assay (51 mucocutaneous, 8 CSF and 32 quality controls) and culture/immunofluorescence (75 mucocutaneous) to assess typing with independent methods. Taqman real-time PCR detected 240 HSV genomes per ml CSF, a level appropriate for the management of patients, and provided unambiguous typing for the 104 positive (62 HSV-1 and 42 HSV-2) out the 160 independent clinical samples tested. HSV type diagnosed by Taqman real-time PCR predicted final diagnosis (meningitis versus encephalitis/meningoencephalitis, p<0.001) in 24/25 patients at time of presentation, in contrast to clinical evaluation. CONCLUSIONS: Our real-time PCR, as a sensitive and specific means for type-specific HSV diagnosis, provided rapid prognostic information for patient management.

New approaches for the control and eradication of schistosomiasis in Venezuela

Schistosomiasis in Americawith the exception of Brazil, behaves as a chronic mild disease with few clinical manifestations due to low parasite burden. These features restrict the clinical and parasitological diagnosis. The most commonly used stool examination method, Kato-Katz, becomes intensitive when the majority of individuals excrete less than 100 eggs/g of feces. In view that antigen-detecting techniques have not been able to reveal light infections, the antibody detecting assays remain as a very valuable diagnostic tool for epidemiological surveillance. The Venezuelan Schistosomiasis Research group (CECOICE) has designed a mass chemotherapy strategy based on sero-diagnosis. Since blood sampling is one of the important limitating factors for large seroepidemiological trials we developed a simple capillary technique that sucessfully overcomed most of the limitations of blood drawing. In this sense, ELISA seems to be the most adecuate test for epidemiological studies. Soluble egg Schistosoma mansoni antigen (SEA) has been largely used in Venezuela. The sensitivity ELISA-SEA in our hands is 90% moreover its specific reach 92% when populations from non-endemic areas but heavily infected with other intestinal parasites are analyzed. The Schistosomiasis Control Program is currently carrying out the surveillance of endemic areas using ELISA-SEA as the first screening method, followed by the Circumoval Precipitin test for validation assay. The results with these two serological techniques allowed us to defined the criteria of chemotherapy in populations of the endemic areas. On the search of better diagnostic technique, Alkaline Phosphatase Immunoenzyme Assay (APIA) is being evaluated in field surveys.

Antibody-mediated status epilepticus: a retrospective multicenter survey.

Background: In recent years, an increasing number of auto-antibodies (AB) have been detected in the CSF and serum of patients with new onset epilepsy. Some of these patients develop convulsive or nonconvulsive status epilepticus (AB-SE), necessitating intensive medical care and administration of multiple antiepileptic and immunomodulatory treatments of uncertain effectiveness. Objectives: In this retrospective multicenter survey we aimed to determine the spectrum of gravity, the duration and the prognosis of the disorder. In addition, we sought to identify the antibodies associated with this condition, as well as determine whether there is a most effective treatment regime. Methods: 12 European Neurology University Clinics, with extensive experience in the treatment of SE patients, were sent a detailed questionnaire regarding symptoms and treatment of AB-SE patients. Seven centers responded positively, providing a total of 13 patients above the age of 16. Results: AB-SE affects mainly women (12/13, 92%) with a variable age at onset (17-69 years, median: 25 years). The duration of the disease is also variable (10 days to 12 years, median: 2 months). Only the 3 oldest patients died (55-69 years). Most patients were diagnosed with anti NMDAR encephalitis (8/13) and had oligoclonal bands in the CSF (9/13). No specific treatment regimen (antiepileptic, immunomodulatory) was found to be clearly superior. Most of the surviving 10 patients (77%) recovered completely or nearly so within 2 years of index poststatus. Conclusion: AB-SE is a severe but potentially reversible condition. Long duration does not seem to imply fatal outcome; however, age older than 50 years at time of onset appears to be a risk factor for death. There was no evidence for an optimal antiepileptic or immunomodulatory treatment. A prospective multicenter study is warranted in order to stratify the optimal treatment algorithm, determine clear risk factors of unfavorable outcome and long-term prognosis.

Sensitivity of Leishmania spp. to Glibenclamide and 4-Aminopiridine: A Tool for the Study of Drug Resistance Development

We have demonstrated that Leishmania spp. grown as promastigotes, are sensitive to the K+ channel inhibitors 4-aminopyridine and glibenclamide. Their host cells, the macrophages, are not affected by similar concentrations of the drugs. We have also initiated the molecular characterization of the mechanisms involved in the development of drug resistance to glibenclamide by the parasite. Therefore, we have selected experimentally and begun to characterize the Venezuelan Leishmania (Leishmania) strain, NR resistant to glibenclamide [NR(Gr)]. The analysis of genomic DNA evidenced the existence of a fragment which apparently is amplified in NR(Gr). The fragment recognized by the pgpA probe, related to the Leishmania P-glycoprotein family and which was originally isolated from L. tarentolae, showed a size polymorfism between the sensitive and the resistant strain. These results suggest that the development of resistance to glibenclamide in the strain NR(Gr) might be associated with the amplification of the ltpgpA or related gene(s)

Infections by protozoa in immunocompromised hosts

Protozoa are among the most important pathogens that can cause infection in immunocompromised patients. They infect particularly individuals with impaired cell immunity, such as those with hematologic neoplasias, those submitted to transplant of solid organs, those under high-dose corticosteroid therapy, and carriers of the human immunodeficiency virus. Among the protozoa that most commonly cause disease in immunocompromised individuals are Toxoplasma gondii, Trypanosoma cruzi, Cryptosporidium parvum, Isospora belli, Cyclospora cayetanensis and microsporidia; the former two cause severe encephalitis and myocarditis, and the others cause gastrointestinal infections. Early diagnosis and prompt institution of specific therapy for each of these organisms are basic measures to decrease morbidity and mortality associated with these infections.

The yellow fever 17D vaccine virus as a vector for the expression of foreign proteins: development of new live flavivirus vaccines

The Flaviviridae is a family of about 70 mostly arthropod-borne viruses many of which are major public health problems with members being present in most continents. Among the most important are yellow fever (YF), dengue with its four serotypes and Japanese encephalitis virus. A live attenuated virus is used as a cost effective, safe and efficacious vaccine against YF but no other live flavivirus vaccines have been licensed. The rise of recombinant DNA technology and its application to study flavivirus genome structure and expression has opened new possibilities for flavivirus vaccine development. One new approach is the use of cDNAs encopassing the whole viral genome to generate infectious RNA after in vitro transcription. This methodology allows the genetic mapping of specific viral functions and the design of viral mutants with considerable potential as new live attenuated viruses. The use of infectious cDNA as a carrier for heterologous antigens is gaining importance as chimeric viruses are shown to be viable, immunogenic and less virulent as compared to the parental viruses. The use of DNA to overcome mutation rates intrinsic of RNA virus populations in conjunction with vaccine production in cell culture should improve the reliability and lower the cost for production of live attenuated vaccines. The YF virus despite a long period ignored by researchers probably due to the effectiveness of the vaccine has made a come back, both in nature as human populations grow and reach endemic areas as well as in the laboratory being a suitable model to understand the biology of flaviviruses in general and providing new alternatives for vaccine development through the use of the 17D vaccine strain.

Effect of a second bloodmeal on the oesophagus colonization by Leishmania mexicana complex in Lutzomyia evansi (Diptera: Psychodidae)

Migration and colonization of the oesophagus by Leishmania mexicana parasites were enhanced after digestion of a second bloodmeal intake in Lutzomyia evansi. This event has epidemiological significance since it affects the infection susceptibility of this sand fly species, which is a proven vector of L. chagasi in Colombian and Venezuelan visceral leishmaniasis foci. Also, it may explain the host seeking behaviour displayed by some partially bloodfed flies found inside houses.

Schistosomiasis mansoni in low transmission areas: abdominal ultrasound

In endemic areas with low prevalence and low intensity of infection, the diagnosis of hepatic pathology due to the Schistosoma mansoni infection is very difficult. In order to establish the hepatic morbidity, a double-blind study was achieved in Venezuelan endemic areas, with one group of patients with schistosomiasis and the other one of non-infected people, that were evaluated clinically and by abdominal ultrasound using the Cairo classification. Schistosomiasis diagnosis was established based on parasitologic and serological tests. The increase of the hepatic size at midclavicular and midsternal lines (in hepatometry) and the hard liver consistency were the clinical parameters able to differentiate infected persons from non infected ones, as well as the presence of left lobe hepatomegaly detected by abdominal ultrasound. The periportal thickening, especially the mild form, was frequent in all age groups in both infected and uninfected patients. There was not correlation between the intensity of infection and ultrasound under the current circumstances. Our data suggest that in Venezuela, a low endemic area of transmission of schistosomiasis, the hepatic morbidity is mild and uncommon. The Cairo classification seems to overestimate the prevalence of periportal pathology. The specificity of the method must be improved, especially for the recognition of precocious pathology. Other causes of hepatopathies must be investigated.

Trypanosoma cruzi-elicited CD8+ T cell-mediated myocarditis: chemokine receptors and adhesion molecules as potential therapeutic targets to control chronic inflammation?

In Chagas disease, during the acute phase, the establishment of inflammatory processes is crucial for Trypanosoma cruzi control in target tissues and for the establishment of host/parasite equilibrium. However, in about 30% of the patients, inflammation becomes progressive, resulting in chronic disease, mainly characterized by myocarditis. Although several hypothesis have been raised to explain the pathogenesis of chagasic myocardiopathy, including the persistence of the parasite and/or participation of autoimmune processes, the molecular mechanisms underlying the establishment of the inflammatory process leading to parasitism control but also contributing to the maintenance of T. cruzi-elicited chronic myocarditis remain unsolved. Trying to shed light on these questions, we have for several years been working with murine models for Chagas disease that reproduce the acute self-resolving meningoencephalitis, the encephalitis resulting of reactivation described in immunodeficient individuals, and several aspects of the acute and chronic myocarditis. In the present review, our results are summarized and discussed under the light of the current literature. Furthermore, rational therapeutic intervention strategies based on integrin-mediated adhesion and chemokine receptor-driven recruitment of leukocytes are proposed to control T. cruzi-elicited unbalanced inflammation.

Brazilian Flavivirus phylogeny based on NS5

In this work, a comprehensive phylogenetic study based on 600 base pair nucleotide and on putative 200 amino acid sequences of NS5 was carried out in order to establish genetic relationships among 15 strains of 10 Brazilian flaviviruses: Bussuquara, Cacipacore, dengue type 1, 2 and 4, Iguape, Ilheus, Rocio, Saint Louis encephalitis (SLE), and yellow fever. Phylogenetic trees were created by neighbor-joining and maximum parsimony methods. These trees showed Brazilian flaviviruses grouped into three main branches: yellow fever branch, dengue branch subdivided in types 1, 2 and 4 branches, and Japanese encephalitis virus (JEV) complex branch including SLE virus strains, Cacipacore, Iguape, Rocio, Ilheus and Bussuquara. Viruses transmitted by Aedes mosquitoes, such as dengue and urban yellow fever, that are also the only Flavivirus causing hemorrhagic fevers in Brazil, were grouped in the same cluster. Encephalitis associated viruses, transmitted by Culex mosquitoes such as JEV complex branch including SLE virus strains, Cacipacore, Iguape, Rocio, Ilheus and Bussuquara were also grouped in the same clade.

Surgical glues: are they really adhesive?

OBJECTIVE: The aim of this study is to create a standard test to approve the efficacy of a surgical sealant. An industrial test, the bulge-and-blister test, which is very convenient for measuring adhesion energy, is applied to the surgical field to quantify adhesion of bioadhesives. METHODS: Samples were composed of two circular layers of equine pericardium glued by the surgical sealant studied. The sample was fixed to a support with an industrial glue. The support and the bottom layer were perforated in the centre to allow injection of pressurised water. Water was progressively introduced through the hole in the support and the bottom layer to create a blister with constant radius, increasing height and internal pressure during this first step. At a critical pressure, delamination started, the radius and height of the blister increased and the pressure decreased. At this point, the adhesion energy could be determined. The experimental parameters were measured with a pressure sensor and an optical profilometry device for deflection. RESULTS: Adhesion testing was carried out in eight paired equine pericardium samples bonded with a Dermabond cyanoacrylate glue. The average value of the practical adhesion energy is 2.3 Jm(-2) with a standard deviation of 1.5 Jm(-2). CONCLUSION: Application of the bulge-and-blister test to the surgical field was achieved and allowed a quantification of adhesion of a surgical glue. Such information is essential to compare the different surgical glues presently available. The study of the impact of bonding conditions such as pressure, hygrometry or setting conditions will provide a better understanding of the characteristics of adhesion in the surgical field.

Cross-sectional study defines difference in malaria morbidity in two Yanomami communities on Amazonian boundary between Brazil and Venezuela

It is well established that immunity to malaria is short-lived and is maintained by the continuous contact with the parasite. We now show that the stable transmission of malaria in Yanomami Amerindian communities maintains a degree of immunity in the exposed population capable to reduce prevalence and morbidity of malaria. We examined 508 Yanomami Amerindians living along Orinoco (407) and Mucajaí (101) rivers, on the Venezuelan and Brazilian Amazon region, respectively. At Orinoco villages, malaria was hyperendemic and presented stable transmission, while at Mucajaí villages it was mesoendemic and showed unstable transmission. The frequency of Plasmodium vivax and P. falciparum was roughly comparable in Venezuelan and Brazilian communities. Malaria presented different profiles at Orinoco and Mucajaí villages. In the former communities, malaria showed a lower prevalence (16% x 40.6%), particularly among those over 10 years old (5.2% x 34.8%), a higher frequency of asymptomatic cases (38.5% x 4.9%), and a lower frequency of cases of severe malaria (9.2% x 36.5%). Orinoco villagers also showed a higher reactivity of the immune system, measured by the frequency of splenomegaly (72.4% x 29.7%) and by the splenic index (71.4% over level 1 x 28.6), and higher prevalence (91.1% x 72.1%) and mean titer (1243 x 62) of antiplasmodial IgG antibodies, as well as a higher prevalence (77.4% x 24.7%) and mean titer (120 x 35) of antiplasmodial IgM antibodies. Our findings show that in isolated Yanomami communities the stability of malaria transmission, and the consequent continuous activation of the immune system of the exposed population, leads to the reduction of malaria prevalence and morbidity.