Rapid diagnostic tests duo as alternative to conventional serological assays for conclusive Chagas disease diagnosis

Chagas disease is caused by the parasite Trypanosoma cruzi. It affects several million people, mainly in Latin America, and severe cardiac and/or digestive complications occur in ~30% of the chronically infected patients. Disease acute stage is mostly asymptomatic and infection goes undiagnosed. In the chronic phase direct parasite detection is hampered due to its concealed presence and diagnosis is achieved by serological methods, like ELISA or indirect hemagglutination assays. Agreement in at least two tests must be obtained due to parasite wide antigenic variability. These techniques require equipped labs and trained personnel and are not available in distant regions. As a result, many infected people often remain undiagnosed until it is too late, as the two available chemotherapies show diminished efficacy in the advanced chronic stage. Easy-to-use rapid diagnostic tests have been developed to be implemented in remote areas as an alternative to conventional tests. They do not need electricity, nor cold chain, they can return results within an hour and some even work with whole blood as sample, like Chagas Stat-Pak (ChemBio Inc.) and Chagas Detect Plus (InBIOS Inc.). Nonetheless, in order to qualify a rapidly diagnosed positive patient for treatment, conventional serological confirmation is obligatory, which might risk its start. In this study two rapid tests based on distinct antigen sets were used in parallel as a way to obtain a fast and conclusive Chagas disease diagnosis using whole blood samples. Chagas Stat-Pak and Chagas Detect Plus were validated by comparison with three conventional tests yielding 100% sensitivity and 99.3% specificity over 342 patients seeking Chagas disease diagnosis in a reference centre in Sucre (Bolivia). Combined used of RDTs in distant regions could substitute laborious conventional serology, allowing immediate treatment and favouring better adhesion to it

Use of rapid diagnostic tests (RDTs) for conclusive diagnosis of chronic Chagas disease – field implementation in the Bolivian Chaco region

Chagas disease, caused by the parasite Trypanosoma cruzi, is the neglected tropical disease with a highest burden in Latin America. Its acute stage is mostly asymptomatic and goes unnoticed. Symptoms appear at the chronic stage, which is when diagnosis is usually made. This is based on the agreement of two conventional serological tests such as Enzyme-Linked Immunosorbent Assays (ELISAs). There are commercial kits with good sensitivity and specificity but their use is impractical in many highly endemic regions with poorly equipped laboratories. Luckily, several rapid diagnostic tests (RDTs) are available for the detection of anti-T. cruzi immunoglobulins. They are easy to operate, require no cold storage, provide fast turnaround of results, and some can work with a tiny volume of whole blood as sample. With the aim to field validate their use we compared an alternative algorithm based on a combination of RDTs with the standard based on ELISAs. In both cases a third test was available in case of discordance. RDTs were implemented by mobile teams in field campaigns to detect chronic T. cruzi-infections in the Chaco region of Bolivia. ELISAs were made in the reference laboratories located in the main hospitals of Yacuiba and Villa Montes, two major cities of the region. We enrolled 685 subjects who voluntarily participated in the study and had not been treated against the disease before. The agreement between the two main RDTs was 93.1% (638/685) (kappa index = 0.86; CI 95% 0.83-0.90). In comparison to the ELISAs algorithm, the combined use of the RDTs provided a sensitivity of 97.7% and a specificity of 96.1%. These results support the use of RDTs for the diagnosis of chronic Chagas disease in the studied region, and encourage their evaluation in other regions of Bolivia and other endemic countries

Characterization of digestive disorders of patients with chronic Chagas disease in Cochabamba, Bolivia

Background: Chagas disease (CD) is endemic in Latin America and particularly common in Bolivia, but there is little information on the characteristics of chronic digestive involvement. Objectives: To determine the prevalence and characterize digestive manifestations in chronic CD patients in Cochabamba, Bolivia. Methods: Eighty-five T. cruzi-seropositive individuals with or without digestive symptoms (G1 group), and fifteen T. cruzi-seronegative patients with similar digestive symptoms to those seen in CD (G2 group) were included in the study. All patients underwent a detailed history including past medical history, epidemiological information, hygiene and dietary habits, a complete physical examination, two serological tests for T. cruzi, video endoscopy, barium swallow, and barium enema. Findings: We observed digestive manifestations in T. cruzi seropositive and seronegative patients. Colonic manifestations were detected in both groups, highlighting the relevance of other confounder factors in the region. Constipation was present in 52.9% of G1 patients, 62.4% presented two or more upper digestive tract symptoms, and 5.9% of them presented esophageal manifestations. Helicobacter pylori infection was detected in 58.8% of G1 patients, and all patients presented gastritis on endoscopy. Conclusions: Prevalence of digestive involvement in CD patients is higher than expected. However, digestive symptoms are not always caused by T. cruzi infection and require differential diagnoses

A strategy for scaling up access to comprehensive care in adults with Chagas disease in endemic countries: The Bolivian Chagas Platform

BACKGROUND: Bolivia has the highest prevalence of Chagas disease (CD) in the world (6.1%), with more than 607,186 people with Trypanosoma cruzi infection, most of them adults. In Bolivia CD has been declared a national priority. In 2009, the Chagas National Program (ChNP) had neither a protocol nor a clear directive for diagnosis and treatment of adults. Although programs had been implemented for congenital transmission and for acute cases, adults remained uncovered. Moreover, health professionals were not aware of treatment recommendations aimed at this population, and research on CD was limited; it was difficult to increase awareness of the disease, understand the challenges it presented, and adapt strategies to cope with it. Simultaneously, migratory flows that led Bolivian patients with CD to Spain and other European countries forced medical staff to look for solutions to an emerging problem. INTERVENTION: In this context, thanks to a Spanish international cooperation collaboration, the Bolivian platform for the comprehensive care of adults with CD was created in 2009. Based on the establishment of a vertical care system under the umbrella of ChNP general guidelines, six centres specialized in CD management were established in different epidemiological contexts. A common database, standardized clinical forms, a and a protocolized attention to adults patients, together with training activities for health professionals were essential for the model success. With the collaboration and knowledge transfer activities between endemic and non-endemic countries, the platform aims to provide care, train health professionals, and create the basis for a future expansion to the National Health System of a proven model of care for adults with CD. RESULTS: From 2010 to 2015, a total of 26,227 patients were attended by the Platform, 69% (18,316) were diagnosed with T. cruzi, 8,567 initiated anti-parasitic treatment, more than 1,616 health professionals were trained, and more than ten research projects developed. The project helped to increase the number of adults with CD diagnosed and treated, produce evidence-based clinical practice guidelines, and bring about changes in policy that will increase access to comprehensive care among adults with CD. The ChNP is now studying the Platform's health care model to adapt and implement it nationwide. CONCLUSIONS: This strategy provides a solution to unmet demands in the care of patients with CD, improving access to diagnosis and treatment. Further scaling up of diagnosis and treatment will be based on the expansion of the model of care to the NHS structures. Its sustainability will be ensured as it will build on existing local resources in Bolivia. Still human trained resources are scarce and the high staff turnover in Bolivia is a limitation of the model. Nevertheless, in a preliminary two-years-experience of scaling up this model, this limitations have been locally solved together with the health local authorities

Molecular detection and parasite load of Trypanosoma cruzi in digestive tract tissue of Chagas disease patients affected by megacolon

Chagas disease, caused by the Trypanosoma cruzi parasite in the Americas affects similar to 7 million people, 30% with cardiac tissue damage and 10-15% with digestive disorders. In this study, we have developed a protocol to detect the presence of the parasite and estimate its load in resected dysfunctional tissue segments of chronically infected patients with digestive megacolon. We have included samples from 43 individuals, 38/5 with positive/negative serology for Chagas disease and digestive syndromes. Samples of 1.5 to 2.0 cm(2) were taken from different points of the dysfunctional digestive tract in specialized centres in Cochabamba, Bolivia. T. cruzi cultures were performed by inoculation with NNN-LIT culture medium, and genomic material was obtained from the samples for multiplex qPCR with TaqMan probes targeting satellite nuclear DNA. Cultures failed to isolate T. cruzi but qPCR reached a sensitivity of 42.1% (16/38) with all three spots and in triplicate. A new quantification methodology using synthetic satellite DNA as quantitation standard revealed parasite loads ranging from 2.2 x 10(2) to 1.0 x 10(6) satellite DNA copies/mu l. Positive samples from the distal end showed a higher parasite load. The results of the present study strengthen and add further evidence to previous findings in an experimental mouse model of chronic T. cruzi infection, providing a valuable tool to improve scientific knowledge on the relevance of the digestive tract in parasite persistence, and underline the need of a better understanding of host-pathogen interaction in digestive tissues, considering pathophysiology, disease immunology and response to treatment

Intradomiciliary and peridomiciliary captures of sand flies (Diptera: Psychodidae) in the leishmaniasis endemic area of Chapare province, Tropic of Cochabamba, Bolivia

In South America, cutaneous leishmaniasis is the most frequent clinical form of leishmaniasis. Bolivia is one of the countries with higher incidence, with 33 cases per 100,000 individuals, and the disease is endemic in 70% of the territory. In the last decade, the number of cases has increased, the age range has expanded, affecting children under 5 years old, and a similar frequency between men and women is found. An entomological study with CDC light traps was conducted in three localities (Chipiriri, Santa Elena and Pedro Domingo Murillo) of the municipality of Villa Tunari, one of the main towns in the Chapare province (Department of Cochabamba, Bolivia). A total of 16 specimens belonging to 6 species of the genus Lutzomyia were captured: Lu. aragaoi, Lu. andersoni, Lu. antunesi, Lu. shawi, Lu. yuilli yuilli and Lu. auraensis. Our results showed the presence of two incriminated vectors of leishmaniasis in an urbanized area and in the intradomicile. More entomological studies are required in the Chapare province to confirm the role of vector sand flies, the intradomiciliary transmission of the disease and the presence of autochthonous cases of cutaneous leishmaniasis

Rapid diagnostic tests duo as alternative to conventional serological assays for conclusive Chagas disease diagnosis

Chagas disease is caused by the parasite Trypanosoma cruzi. It affects several million people, mainly in Latin America, and severe cardiac and/or digestive complications occur in ~30% of the chronically infected patients. Disease acute stage is mostly asymptomatic and infection goes undiagnosed. In the chronic phase direct parasite detection is hampered due to its concealed presence and diagnosis is achieved by serological methods, like ELISA or indirect hemagglutination assays. Agreement in at least two tests must be obtained due to parasite wide antigenic variability. These techniques require equipped labs and trained personnel and are not available in distant regions. As a result, many infected people often remain undiagnosed until it is too late, as the two available chemotherapies show diminished efficacy in the advanced chronic stage. Easy-to-use rapid diagnostic tests have been developed to be implemented in remote areas as an alternative to conventional tests. They do not need electricity, nor cold chain, they can return results within an hour and some even work with whole blood as sample, like Chagas Stat-Pak (ChemBio Inc.) and Chagas Detect Plus (InBIOS Inc.). Nonetheless, in order to qualify a rapidly diagnosed positive patient for treatment, conventional serological confirmation is obligatory, which might risk its start. In this study two rapid tests based on distinct antigen sets were used in parallel as a way to obtain a fast and conclusive Chagas disease diagnosis using whole blood samples. Chagas Stat-Pak and Chagas Detect Plus were validated by comparison with three conventional tests yielding 100% sensitivity and 99.3% specificity over 342 patients seeking Chagas disease diagnosis in a reference centre in Sucre (Bolivia). Combined used of RDTs in distant regions could substitute laborious conventional serology, allowing immediate treatment and favouring better adhesion to it

Tegumentary leishmaniasis by Leishmania braziliensis complex in Cochabamba, Bolivia including the presence of L. braziliensis outlier

Leishmaniasis is caused by protozoans of the Leishmania genus, which includes more than 20 species capable of infecting humans worldwide. In the Americas, the most widespread specie is L. braziliensis, present in 18 countries including Bolivia. The taxonomic position of the L. braziliensis complex has been a subject of controversy, complicated further by the recent identification of a particular subpopulation named L. braziliensis atypical or outlier. The aim of this study was to carry out a systematic analysis of the L. braziliensis complex in Bolivia and to describe the associated clinical characteristics. Forty-one strains were analyzed by sequencing an amplified 1245 bp fragment of the hsp70 gene, which allowed its identification as: 24 (59%) L. braziliensis, 16 (39%) L. braziliensis outlier, and one (2%) L. peruviana. In a dendrogram constructed, L. braziliensis and L. peruviana are grouped in the same cluster, whilst L. braziliensis outlier appears in a separate branch. Sequence alignment allowed the identification of five non-polymorphic nucleotide positions (288, 297, 642, 993, and 1213) that discriminate L. braziliensis and L. peruviana from L. braziliensis outlier. Moreover, nucleotide positions 51 and 561 enable L. peruviana to be discriminated from the other two taxa. A greater diversity was observed in L. braziliensis outlier than in L. braziliensis-L. peruviana. The 41 strains came from 32 patients with tegumentary leishmaniasis, among which 22 patients (69%) presented cutaneous lesions (11 caused by L. braziliensis and 11 by L. braziliensis outlier) and 10 patients (31%) mucocutaneous lesions (eight caused by L. braziliensis, one by L. braziliensis outlier, and one by L. peruviana). Nine patients (28%) simultaneously provided two isolates, each from a separate lesion, and in each case the same genotype was identified in both. Treatment failure was observed in six patients infected with L. braziliensis and one patient with L. peruviana