T. cruzi OligoC-TesT: A Simplified and Standardized Polymerase Chain Reaction Format for Diagnosis of Chagas Disease

Chagas disease (American trypanosomiasis) is caused by the protozoan parasite Trypanosoma cruzi and represents a major public health problem in Latin America. Furthermore, growing human population movements extend the disease distribution to regions outside the South American continent. Accurate diagnosis is crucial in patient care and in preventing transmission through blood transfusion, organ transplantation, or vertical transmission from mother to child. Routine diagnosis of Trypanosoma cruzi infection generally is based on detection of the host's antibodies against the parasite. However, antibody detection tests are liable to specificity problems and are of limited use in assessing treatment outcome and congenital infections. The introduction of the polymerase chain reaction (PCR) to amplify specific DNA sequences opened promising diagnostic perspectives. Despite its reported high sensitivity and specificity, broad use of the PCR technique in diagnosis of Chagas disease is hampered by its complexity and the lack of any standardization. We here present the development and evaluation of the T. cruzi OligoC-TesT, a simple and standardized dipstick format for detection of PCR amplified T. cruzi DNA. The new tool is an important step towards simplified and standardized molecular diagnosis of Chagas disease

Prevalence, Clinical Staging and Risk for Blood-Borne Transmission of Chagas Disease among Latin American Migrants in Geneva, Switzerland

Chagas disease, a parasitic disease caused by Trypanosoma cruzi, is a leading cause of cardiac and digestive tract disorders in Mexico, Central and South America. An increasing number of cases have recently been reported in North America and Europe due to international human migration, but data outside Latin America remains scarce. This study showed that Chagas disease is an emerging health problem in Switzerland, affecting a substantial proportion of Latin American migrants (13%). Persons at increased risk of infection were Bolivian, older than 35 years or had a mother infected with T. cruzi. Early signs of cardiac or digestive tract disease were found in one out of six infected patients. The risk of local transmission by blood transfusion or organ transplant was illustrated by the frequent willingness expressed by patients to donate blood or organs in Switzerland. The authors recommend the screening of persons at risk of infection and the diffusion of appropriate information to the medical community to increase awareness of this emerging health problem. Considering that affected persons frequently lack health insurance in Switzerland, a facilitated access to medical care is an important step towards better recognition and management of Chagas disease

Transfusion-transmitted infections

Although the risk of transfusion-transmitted infections today is lower than ever, the supply of safe blood products remains subject to contamination with known and yet to be identified human pathogens. Only continuous improvement and implementation of donor selection, sensitive screening tests and effective inactivation procedures can ensure the elimination, or at least reduction, of the risk of acquiring transfusion transmitted infections. In addition, ongoing education and up-to-date information regarding infectious agents that are potentially transmitted via blood components is necessary to promote the reporting of adverse events, an important component of transfusion transmitted disease surveillance. Thus, the collaboration of all parties involved in transfusion medicine, including national haemovigilance systems, is crucial for protecting a secure blood product supply from known and emerging blood-borne pathogens

Mast Cell-Derived Histamine Mediates Cystitis Pain

Background: Mast cells trigger inflammation that is associated with local pain, but the mechanisms mediating pain are unclear. Interstitial cystitis (IC) is a bladder disease that causes debilitating pelvic pain of unknown origin and without consistent inflammation, but IC symptoms correlate with elevated bladder lamina propria mast cell counts. We hypothesized that mast cells mediate pelvic pain directly and examined pain behavior using a murine model that recapitulates key aspects of IC. Methods and Findings: Infection of mice with pseudorabies virus (PRV) induces a neurogenic cystitis associated with lamina propria mast cell accumulation dependent upon tumor necrosis factor alpha (TNF), TNF-mediated bladder barrier dysfunction, and pelvic pain behavior, but the molecular basis for pelvic pain is unknown. In this study, both PRV-induced pelvic pain and bladder pathophysiology were abrogated in mast cell-deficient mice but were restored by reconstitution with wild type bone marrow. Pelvic pain developed normally in TNF- and TNF receptor-deficient mice, while bladder pathophysiology was abrogated. Conversely, genetic or pharmacologic disruption of histamine receptor H1R or H2R attenuated pelvic pain without altering pathophysiology. Conclusions: These data demonstrate that mast cells promote cystitis pain and bladder pathophysiology through the separable actions of histamine and TNF, respectively. Therefore, pain is independent of pathology and inflammation, an

Combined Treatment of Heterocyclic Analogues and Benznidazole upon Trypanosoma cruzi In Vivo

Chagas disease caused by Trypanosoma cruzi is an important cause of mortality and morbidity in Latin America but no vaccines or safe chemotherapeutic agents are available. Combined therapy is envisioned as an ideal approach since it may enhance efficacy by acting upon different cellular targets, may reduce toxicity and minimize the risk of drug resistance. Therefore, we investigated the activity of benznidazole (Bz) in combination with the diamidine prodrug DB289 and in combination with the arylimidamide DB766 upon T. cruzi infection in vivo. The oral treatment of T.cruzi-infected mice with DB289 and Benznidazole (Bz) alone reduced the number of circulating parasites compared with untreated mice by about 70% and 90%, respectively. However, the combination of these two compounds decreased the parasitemia by 99% and protected against animal mortality by 100%, but without providing a parasitological cure. When Bz (p.o) was combined with DB766 (via ip route), at least a 99.5% decrease in parasitemia levels was observed. DB766+Bz also provided 100% protection against mice mortality while Bz alone provided about 87% protection. This combined therapy also reduced the tissular lesions induced by T. cruzi infection: Bz alone reduced GPT and CK plasma levels by about 12% and 78% compared to untreated mice group, the combination of Bz with DB766 resulted in a reduction of GPT and CK plasma levels of 56% and 91%. Cure assessment through hemocultive and PCR approaches showed that Bz did not provide a parasitological cure, however, DB766 alone or associated with Bz cured ≥13% of surviving animals

The Involvement of IL-17A in the Murine Response to Sub-Lethal Inhalational Infection with Francisella tularensis

Background: Francisella tularensis is an intercellular bacterium often causing fatal disease when inhaled. Previous reports have underlined the role of cell-mediated immunity and IFNc in the host response to Francisella tularensis infection. Methodology/Principal Findings: Here we provide evidence for the involvement of IL-17A in host defense to inhalational tularemia, using a mouse model of intranasal infection with the Live Vaccine Strain (LVS). We demonstrate the kinetics of IL-17A production in lavage fluids of infected lungs and identify the IL-17A-producing lymphocytes as pulmonary cd and Th17 cells. The peak of IL-17A production appears early during sub-lethal infection, it precedes the peak of immune activation and the nadir of the disease, and then subsides subsequently. Exogenous airway administration of IL-17A or of IL-23 had a limited yet consistent effect of delaying the onset of death from a lethal dose of LVS, implying that IL-17A may be involved in restraining the infection. The protective role for IL-17A was directly demonstrated by in vivo neutralization of IL-17A. Administration of anti IL-17A antibodies concomitantly to a sub-lethal airway infection with 0.16LD50 resulted in a fatal disease. Conclusion: In summary, these data characterize the involvement and underline the protective key role of the IL-17A axis in the lungs from inhalational tularemia

Cellular and molecular changes and immune response in the intestinal mucosa during Trichinella spiralis early infection in rats

Background:: The main targets of the host's immune system in Trichinella spiralis infection are the adult worms (AW), at the gut level, and the migrant or newborn larvae (NBL), at systemic and pulmonary levels. Most of the studies carried out in the gut mucosa have been performed on the Payer's patches and/or the mesenteric lymph nodes but not on the lamina propria, therefore, knowledge on the gut immune response against T. spiralis remains incomplete. Methods: This study aimed at characterizing the early mucosal immune response against T. spiralis, particularly, the events taking place between 1 and 13 dpi. For this purpose, Wistar rats were orally infected with muscle larvae of T. spiralis and the humoral and cellular parameters of the gut immunity were analysed, including the evaluation of the ADCC mechanism exerted by lamina propria cells. Results: A marked inflammation and structural alteration of the mucosa was found. The changes involved an increase in goblet cells, eosinophils and mast cells, and B and T lymphocytes, initially displaying a Th1 profile, characterised by the secretion of IFN-γand IL-12, followed by a polarization towards a Th2 profile, with a marked increase in IgE, IgG1, IL-4, IL-5 and IL-13 levels, which occurred once the infection was established. In addition, the helminthotoxic activity of lamina propria cells demonstrated the role of the intestine as a place of migrant larvae destruction, indicating that not all the NBLs released in the gut will be able to reach the muscles. Conclusions: The characterization of the immune response triggered in the gut mucosa during T. spiralis infection showed that not only an effector mechanism is directed toward the AW but also towards the NBL as a cytotoxic activity was observed against NBL exerted by lamina propria cells.Fil: Saracino, María Priscila. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Estudios de la Inmunidad Humoral Prof. Ricardo A. Margni. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Instituto de Estudios de la Inmunidad Humoral Prof. Ricardo A. Margni; ArgentinaFil: Vila, Cecilia Celeste. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Estudios de la Inmunidad Humoral Prof. Ricardo A. Margni. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Instituto de Estudios de la Inmunidad Humoral Prof. Ricardo A. Margni; ArgentinaFil: Cohen, Melina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Estudios de la Inmunidad Humoral Prof. Ricardo A. Margni. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Instituto de Estudios de la Inmunidad Humoral Prof. Ricardo A. Margni; ArgentinaFil: Gentilini, Maria Virginia. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Estudios de la Inmunidad Humoral Prof. Ricardo A. Margni. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Instituto de Estudios de la Inmunidad Humoral Prof. Ricardo A. Margni; ArgentinaFil: Falduto, Guido Hernán. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Estudios de la Inmunidad Humoral Prof. Ricardo A. Margni. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Instituto de Estudios de la Inmunidad Humoral Prof. Ricardo A. Margni; ArgentinaFil: Calcagno, Marcela Adriana. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Estudios de la Inmunidad Humoral Prof. Ricardo A. Margni. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Instituto de Estudios de la Inmunidad Humoral Prof. Ricardo A. Margni; ArgentinaFil: Roux, Estela. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Cardiológicas. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Cardiológicas; ArgentinaFil: Venturiello, Stella Maris. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Estudios de la Inmunidad Humoral Prof. Ricardo A. Margni. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Instituto de Estudios de la Inmunidad Humoral Prof. Ricardo A. Margni; ArgentinaFil: Malchiodi, Emilio Luis. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Estudios de la Inmunidad Humoral Prof. Ricardo A. Margni. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Instituto de Estudios de la Inmunidad Humoral Prof. Ricardo A. Margni; Argentin

ELISA versus PCR for diagnosis of chronic Chagas disease: systematic review and meta-analysis

<p>Abstract</p> <p>Background</p> <p>Most current guidelines recommend two serological tests to diagnose chronic Chagas disease. When serological tests are persistently inconclusive, some guidelines recommend molecular tests. The aim of this investigation was to review chronic Chagas disease diagnosis literature and to summarize results of ELISA and PCR performance.</p> <p>Methods</p> <p>A systematic review was conducted searching remote databases (MEDLINE, LILACS, EMBASE, SCOPUS and ISIWeb) and full texts bibliography for relevant abstracts. In addition, manufacturers of commercial tests were contacted. Original investigations were eligible if they estimated sensitivity and specificity, or reliability -or if their calculation was possible - of ELISA or PCR tests, for chronic Chagas disease.</p> <p>Results</p> <p>Heterogeneity was high within each test (ELISA and PCR) and threshold effect was detected only in a particular subgroup. Reference standard blinding partially explained heterogeneity in ELISA studies, and pooled sensitivity and specificity were 97.7% [96.7%-98.5%] and 96.3% [94.6%-97.6%] respectively. Commercial ELISA with recombinant antigens studied in phase three investigations partially explained heterogeneity, and pooled sensitivity and specificity were 99.3% [97.9%-99.9%] and 97.5% [88.5%-99.5%] respectively. ELISA's reliability was seldom studied but was considered acceptable. PCR heterogeneity was not explained, but a threshold effect was detected in three groups created by using guanidine and boiling the sample before DNA extraction. PCR sensitivity is likely to be between 50% and 90%, while its specificity is close to 100%. PCR reliability was never studied.</p> <p>Conclusions</p> <p>Both conventional and recombinant based ELISA give useful information, however there are commercial tests without technical reports and therefore were not included in this review. Physicians need to have access to technical reports to understand if these serological tests are similar to those included in this review and therefore correctly order and interpret test results. Currently, PCR should not be used in clinical practice for chronic Chagas disease diagnosis and there is no PCR test commercially available for this purpose. Tests limitations and directions for future research are discussed.</p