Evaluation of serological diagnostic tests of human brucellosis for prevention and control in Mexico.

Brucellosis is a zoonosis mainly present in developing countries. The WHO reports 500,000 new cases every year. From 2012 to 2016, 13,677 cases were reported in Mexico, with 2.00 to 2.64 rate per 100,000 inhabitants. To analyze the diagnostic algorithm of brucellosis in Mexico, we compared the commercial laboratory tests ELISA, Brucellacapt®, and lateral flow test (LFT) in a study of 473 individuals from two endemic Mexican populations. All patients were treated in first-level medical units for presenting brucellosis compatible symptoms and without a history of the disease. Clinical-epidemiological information was gathered and initial serum samples were obtained to react with anti-Brucella antibodies; subsequent samples were collected at follow-up treatment visits. Using the Rose Bengal screening, we found 165 negative samples and 308 positive reactive samples, of which 222 cases were confirmed and 234 were positive on at least one marker (IgG or IgM) or LFT. When Brucellacapt® was used, similar results to those observed with the conventional algorithm were found as judged by the Cohen's kappa coefficient (κ) (0.813, 95% CI 0.7788-0.8472). Similar κ indices between conventional algorithm and ELISA pair were found, 0.7038 (95% CI 0.6555-0.7521), representing high similarity between both groups of diagnosis. We conclude that conventional serodiagnoses, Brucellacapt® and LFT, presented inconclusive results and poor correlation between them. By contrast, ELISA test pair (IgG + IgM) presented high correlation with the conventional algorithm and greater capacity for correct positive and negative classification.

From ISET to InDRE. V. Institute of Epidemiological Diagnosis and Reference. Global strategic position, 2012-2019.

This document describes the changes at the Institute of Epidemiological Diagnosis and Reference (InDRE) from 2012 to 2019, the administrative and equipment modifications, the new headquarters and the National System of Epidemiological Surveillance legal modifications. The process of relocation is mentioned, especially the careful transfer of the biological material protected by the Institute, and the new way of studying epidemic outbreaks, endemic diseases and the negative network is analyzed. At the international level, the promotion of links with global networks of the Pan American Health Organization, the World Health Organization (WHO) and other international organizations is described. The assignation to InDRE of four WHO collaborating centres is also mentioned. The Global Health Security Initiative Laboratory Network acknowledged InDRE's leadership, which co-chaired the working group during the study period.

En este documento se describen los cambios en el Instituto de Diagnóstico y Referencia Epidemiológicos (InDRE) de 2012 a 2019, las modificaciones administrativas y de equipamiento, la nueva sede y las modificaciones jurídicas al Sistema Nacional de Vigilancia Epidemiológica. Se menciona el proceso de mudanza, en especial el cuidadoso traslado del material biológico que resguarda el Instituto y se analiza la nueva forma de estudiar los brotes epidémicos, los padecimientos endémicos y la red negativa. Respecto al ámbito internacional, se describe el fomento de la vinculación con redes globales de la Organización Panamericana de la Salud, la Organización Mundial de la Salud (OMS) y otros organismos internacionales. También se menciona la designación en el InDRE de cuatro centros colaboradores de la OMS. La Red de Laboratorios de la Iniciativa Global para la Seguridad en Salud reconoció el liderazgo del InDRE, cuyo director ocupó la copresidencia del grupo de trabajo en el periodo de estudio.

Comparative evaluation of 11 commercialized rapid diagnostic tests for detecting Trypanosoma cruzi antibodies in serum banks in areas of endemicity and nonendemicity.

Chagas disease is one of the main public health issues in Latin America. Increasingly during the past few decades, Trypanosoma cruzi infection has been detected in North America, Europe, and the Western Pacific, mainly as a result of population movement. The limited availability of rapid serological diagnostic tests hinders rapid diagnosis and early treatment in areas of endemicity and nonendemicity. In collaboration with 11 national reference laboratories (NRLs) from different geographical areas, we evaluated the performances of commercialized serological rapid diagnostic tests (RDT) for T. cruzi infection. Eleven commercialized T. cruzi infection RDTs were evaluated on a total of 474 samples extensively tested with at least three different techniques for Chagas disease, maintained at controlled low temperatures, and stored in the serum banks of the 11 NRLs. We measured the sensitivity, specificity, and concordance of each RDT and provided an additional questionnaire to evaluate its ease of use. The selected RDTs in this study were performed under controlled laboratory conditions. Out of the 11 RDTs, we found 8 of them to be useful, with the cassette format favored over the strip. We did not observe significant differences in RDT performances in the different regions. Overall, the performance results were lower than those disclosed by the manufacturers. The results of this evaluation validate the possibility of using RDTs to diagnose Chagas disease, thereby decreasing the time to treatment at a primary health care facility for patients who are willing to be treated. Further studies should be conducted in the laboratory and in the field to confirm these data, expressly to evaluate reproducibility in resource-limited settings, or using whole blood in clinical settings in areas of endemicity and nonendemicity.

Linker for activation of T cells is displaced from lipid rafts and decreases in lupus T cells after activation via the TCR/CD3 pathway.

Systemic lupus erythematosus (SLE) is characterized by abnormal signal transduction mechanisms in T lymphocytes. Linker for activation of T cells (LAT) couples TCR/CD3 activation with downstream signaling pathways. We reported diminished ERK 1/2 kinase activity in TCR/CD3 stimulated lupus T cells. In this study we evaluated the expression, phosphorylation, lipid raft and immunological synapse (IS) localization and colocalization of LAT with key signalosome molecules. We observed a diminished expression and an abnormal localization of LAT in lipid rafts and at the IS in activated lupus T cells. LAT phosphorylation, capture by GST-Grb2 fusion protein, and coupling to Grb2 and PLCγ1, was similar in healthy control and lupus T cells. Our results suggest that an abnormal localization of LAT within lipid rafts and its accelerated degradation after TCR/CD3 activation may compromise the assembly of the LAT signalosome and downstream signaling pathways required for full MAPK activation in lupus T cells.

Presence of Acute Chagas Disease Among Febrile Patients in the Western Coast of Mexico.

Chagas disease (ChD) is a parasitosis caused by the protozoan Trypanosoma cruzi (Tc). It is endemic to almost all Latin American countries, including the southern United States. The acute form of ChD and its actual incidence have rarely been described in Mexico, despite the extensive presence of favorable niches for its transmission. The objective of this study was to estimate the frequency of acute ChD in febrile patients at the central Pacific coast of Mexico. For this, we surveyed patients with persistent fever (5 to 10 days) in five hospitals at the Mexican states of Jalisco, Colima, and Nayarit in 2012. Samples were taken from a total of 485 patients to detect Tc in blood using the polymerase chain reaction (PCR) test and direct microscopic examination. Of these subjects, 10 were positive for PCR and none for microscopic examination (2% in 12 months). We adjusted this rate by the total people at risk in the area and obtained an incidence of 7.4/100,000 habs./year. The positive cases showed no association with sex, rural settlement, or pet ownership, only with the contact with Triatominae insects (odds ratio = 9.22 and confidence interval: 1.93-44.06). The clinical picture of positive patients showed an association with the diagnosis of lower respiratory tract infections. Meanwhile, only one fatal case showed the typical picture of acute fatal cardiomyopathy. The pulmonary manifestations of our patients suggest possible lung pathogenicity of Tc, which merits further investigation. Our findings differ markedly from the official reports for ChD. This difference suggests an underestimation of the disease. These findings urge the Mexican health authorities to implement more vigorous actions aimed at improving medical skills in the timely diagnosis of ChD, as well as to apply efficient preventive programs.

Prevalence of anemia and deficiencies of iron, folic acid and vitamin B12 in an indigenous community from the Venezuelan Amazon with a high incidence of malaria.

The objective of this work was to determine the prevalence of anemia and deficiencies of iron, folic acid and vitamin B12 in Betania del Topocho, a Piaroa community from Estado Amazonas, Venezuela, a zone with a high incidence of malaria. The group studied included 184 subjects of all ages that assisted to the local health center for malaria diagnosis. Analysis performed included hematology by coulter counter, ferritin quantification by ELISA with monoclonal antibodies and folic acid and vitamin B12 determinations by an immunoradiometric assay. It was found that the prevalence of anemia was 89.6% and deficiencies of iron, folic acid and vitamin B12 affected 37.1,70.3 and 12.4% of the population studied, respectively. Plasmodium infection was detected by molecular diagnosis in 53.2% of the cases, and 86% of them were anemic. The highest incidence of anemia was found in children, with a prevalence of 100% in infants of both sexes. The high prevalence of anemia, iron and folic acid deficiencies found, indicates an important health and nutrition problem that should be immediately and properly addressed. The number of cases of anemia due to iron deficiency could be underestimated, since ferritin concentration increased as a acute phase protein, although prevalence data was also analyzed with a cutoff point of 30 microg/L for ferritin concentration.

Dengue Virus Coinfection in Human Immunodeficiency Virus-1-Infected Patients on the West Coast of Mexico.

Dengue virus infection in human immunodeficiency virus (HIV)-positive patients is not well studied. Previous reports suggest a transitory inhibition of the HIV-1 viral load, as well as a benign clinical progression of dengue. The follow-up of six HIV-1-infected patients, diagnosed and hospitalized with dengue virus infection in the State of Colima, Mexico, was carried out to analyze the progression of this viral coinfection. The presence of dengue virus serotype 1 was confirmed through molecular tests. No severe complications were observed in any of the patients during dengue virus infection. Significant alteration of the HIV-1 viral loads was not observed during dengue virus infection and 6 months after coinfection. Further studies are required to understand the pathology, as well as the clinical course, of these viral coinfections.

Del ISET al InDRE. V. Instituto de Diagnóstico y Referencia Epidemiológicos. Posición estratégica global, 2012-2019 / From ISET to InDRE. V. Institute of Epidemiological Diagnosis and Reference. Global strategic position, 2012-2019

Resumen En este documento se describen los cambios en el Instituto de Diagnóstico y Referencia Epidemiológicos (InDRE) de 2012 a 2019, las modificaciones administrativas y de equipamiento, la nueva sede y las modificaciones jurídicas al Sistema Nacional de Vigilancia Epidemiológica. Se menciona el proceso de mudanza, en especial el cuidadoso traslado del material biológico que resguarda el Instituto y se analiza la nueva forma de estudiar los brotes epidémicos, los padecimientos endémicos y la red negativa. Respecto al ámbito internacional, se describe el fomento de la vinculación con redes globales de la Organización Panamericana de la Salud, la Organización Mundial de la Salud (OMS) y otros organismos internacionales. También se menciona la designación en el InDRE de cuatro centros colaboradores de la OMS. La Red de Laboratorios de la Iniciativa Global para la Seguridad en Salud reconoció el liderazgo del InDRE, cuyo director ocupó la copresidencia del grupo de trabajo en el periodo de estudio.

Abstract This document describes the changes at the Institute of Epidemiological Diagnosis and Reference (InDRE) from 2012 to 2019, the administrative and equipment modifications, the new headquarters and the National System of Epidemiological Surveillance legal modifications. The process of relocation is mentioned, especially the careful transfer of the biological material protected by the Institute, and the new way of studying epidemic outbreaks, endemic diseases and the negative network is analyzed. At the international level, the promotion of links with global networks of the Pan American Health Organization, the World Health Organization (WHO) and other international organizations is described. The designation of four WHO collaborating centres granted to InDRE is also mentioned. The Global Health Security Initiative Laboratory Network acknowledged InDRE's leadership, which co-chaired the working group during the study period.

Association of leishmaniasis with TNF alpha promoter and SLC11A1 gene polymorphisms in patients of two endemic areas in Mexico.

Some Single Nucleotide Polymorphisms (SNPs) of interleukins and other modulatory molecules of the immune response play an important role in susceptibility to infectious diseases, particularly those involving intracellular parasites. In this study, we evaluated allele, genotype and haplotype associations of two SNPs of the TNF-α promoter and seven of the SLC11A1 gene in 79 patients with localized cutaneous leishmaniasis (CL) and 15 with visceral leishmaniasis (VL), compared with 127 and 89 locality paired controls, respectively, from two endemic areas of Chiapas State, Mexico. None of the TNF-α alleles and genotypes was associated either to CL or to VL. Alleles rs2276631-C (P = 0.02; OR [95%CI] = 2.11 [1.16-3.86]) and rs2279015-G (P = 0.005; OR [95%CI] = 2.42 [1.33-4.41]) of SLC11A1, were associated with susceptibility to VL, whereas genotypes rs2276631 C/C (P = 0.003; OR [95%CI] = 2.65 [1.41-5.00]) and rs2279015 G/G (P = 0.018; OR [95%CI] = 2.05 [1.15-3.64]) were significantly increased in CL and VL patients, respectively. Complete haplotypes involved in susceptibility were CGCCGDins with VL and CGCCADins with CL. CGCCA was the minimal susceptibility haplotype for CL and CCG for VL. Our data suggest that SLC11A1 gene polymorphisms might have a relevant role in the pathology of leishmaniasis, directing towards susceptibility outcome of this disease in residents of an endemic area.

Infectious diseases in Mexico. A survey from 1995-2000.

Data obtained at a central laboratory for emerging, re-emerging, and other infectious diseases in Mexico from 1995-2000 are presented. An outstanding increase of DEN-3 circulation was identified. Aedes aegypti, the dengue vector, is widely distributed. Leptospirosis has become the most important differential diagnosis for dengue. Identification of rabies virus variants allowed cataloging of new transmitters of rabies. Rotavirus showed a clear seasonal distribution, while different proportions of pathogenic classes of Escherichia coli under endemic and outbreak conditions were seen. Serotypes of several bacteria are reported as well as the sources of isolation and frequency of Shigella, Salmonella, and Vibrio cholerae. Rise and disappearance of cholera could be followed along the past decade. Influenza strains were identified, as were several pathogens causing sexually transmitted infections. Laboratory support was important for surveillance after Hurricane Mitch. Multidrug-resistant strains of Mycobacterium tuberculosis are emerging and primary resistance is very high. It is now mandatory to search for antibodies to Trypanosoma cruzi in blood banks. Triatoma barberi, a peridomestic bug, is the main vector of Chagas disease. Localized cutaneous leishmaniosis increased in regions having a guerrilla element in Chiapas. Modern immunodiagnostic techniques are used for control studies of cysticercosis and similar techniques were recently standardized for Trichinella spiralis detection. Low iodine values in children's urine were found in several Mexican states; therefore, use of iodized salt should be encouraged.

Prevalence of Chagas' disease in Mulungu do Morro northeastern Brazil.

OBJECTIVE: The aim of this paper is to describe the prevalence of T. Cruzi infection in patients of from Mulungu do Morro, a rural tropical region of Northeastern Brazil. METHODS: A cross-sectional study was performed. After randomly selecting samples of the population, and obtaining their consents, patients completed pretested epidemiological and clinical questionnaires. Serum samples from all patients were collected and screened for the presence of T. cruzi antibodies. RESULTS: Of 694 patients examined, 174 patients (25.1%) tested had a positive serology for Chagas' disease. Of the study population, 341 patients were male with 27% Chagas' disease prevalence, without a statistical difference. Illiteracy was the only variable related to T. cruzi infection in our population. CONCLUSION: In conclusion, our study points to the high prevalence of Chagas' disease among patients in Mulungu do Morro, suggesting that this region has a high frequency of infection and probably active vectorial transmission.

ITS1 PCR-RFLP Diagnosis and Characterization of Leishmania in Clinical Samples and Strains from Cases of Human Cutaneous Leishmaniasis in States of the Mexican Southeast.

American cutaneous leishmaniasis includes a spectrum of clinical forms localized cutaneous, diffuse cutaneous, and mucocutaneous leishmaniasis which can be caused by different strains of Leishmania belonging to the L. mexicana or L. braziliensis complexes which may coexist in the same endemic area. We evaluated the PCR-RFLP assay of the ITS1 genes for direct identification of Leishmania species in 163 clinical samples and 21 Mexican isolates of Leishmania. In relation to the Mexican isolates of Leishmania 52% displayed a pattern similar to the L. (L.) mexicana, 5% showed a mixed pattern compatible with L. (L.) mexicana and L. (V.) braziliensis, eight with L. (L.) amazonensis and L. (L.) mexicana, and one to L. (V.) braziliensis. Most of the clinical samples, 109/116 (94%), gave a pattern similar to that of the L. mexicana, two clinical samples gave similar patterns to that of Leishmania braziliensis, and 5 samples gave patterns that suggest a coinfection of L. (L.) mexicana and L. (V.) braziliensis or L. (L.) mexicana and L. (L.) amazonensis. The ITS1 PCR-RFLP assay is a multipurpose tool for diagnosis of Leishmania from clinical samples and enables determination of the infecting species of New World Leishmania in the field in relatively short time and low cost.

American visceral leishmaniasis in Chiapas, Mexico.

We report the results of a study conducted during 1990-2006 with 89 cases of American visceral leishmaniasis in Chiapas State in southeastern Mexico and a seroprevalence study performed with 726 persons and 224 dogs that lived near cases of American visceral leishmaniasis. Clinical aspects, epidemiologic profiles, and risk factors are described. Most cases were in children ≤ 5 years of age, the prevalence of seropositive persons was 77%. The main risk factors associated with this disease were having 1-3 rooms in a house compared with ≥ 4 rooms, having a roof that was not made of cement, and having domestic animals. In contrast, only 19% of dogs were seropositive, suggesting that this species is not important in the transmission cycle of Leishmania. These data indicate that active transmission is taking place in the central valley of Chiapas State, Mexico, in communities located < 1,000 meters above sea level near the Grijalva River.

Prevalence of Trypanosoma cruzi in dogs (Canis familiaris) and triatomines during 2008 in a sanitary region of the State of Mexico, Mexico.

American trypanosomiasis is a public health problem in Latin America and southern parts of the United States. Infection in triatomines (vector) and domestic dogs (reservoir host) is a good indicator of Trypanosoma cruzi circulation and human risk of infection. The State of Mexico, Mexico, has been considered free of T. cruzi, and no detailed epidemiologic study has been conducted to assess the intricacies of the transmission cycle of the parasite in the region. Such studies would enhance our understanding of the epidemiology of T. cruzi infection in this geographic region and provide regional sanitary authorities with stronger fundamental knowledge for making decisions and allocating funds for Chagas disease control programs in the State of Mexico. The objective of this study was to determine the prevalence of T. cruzi infection in dogs (seroprevalence) and triatomines (fecal parasites) in a previously identified, discrete endemic region of parasite circulation and to widen our studies in the Tejupilco Sanitary Region located in the southern part of the State of Mexico. Dog blood samples (n=102) were analyzed for the presence of anti-T. cruzi antibodies by two assays, namely indirect hemagglutination assay and enzyme-linked immunosorbent assay. Triatomines (n=88) were collected and fecal aliquots were analyzed for the presence of parasites by light microscopy. Average seroprevalence in dogs in the Tejupilco Sanitary region was 24.5%, and the overall triatomine infection rate was 34.01%. Triatoma pallidipennis was the only triatomine species found in this region. Our data demonstrate that T. cruzi is actively circulating in the Tejupilco Sanitary Region and emphasize the requirement for epidemiologic surveillance programs throughout the putative endemic areas of the State of Mexico.

Trypanosoma cruzi circulating in the southern region of the State of Mexico (Zumpahuacan) are pathogenic: a dog model.

Here we describe clinical and pathologic evidence of Chagas disease caused in dogs by circulating Trypanosoma cruzi from a newly recognized endemic area in Mexico. We show that the Zumpahuacan isolate, although less virulent than the Sylvio-X10 reference strain that caused acute myocarditis and death, was pathogenic in dogs. Dogs infected with the Zumpahuacan isolate exhibited electrocardiographic alterations, left- and right-ventricle dilation, and hydropericardium. Histologically, diffused perimysial and endomysial lymphoplasmacytic cell infiltration, cardiomyocyte necrosis, and amastigote nests were noted in Zumpahuacan-infected dogs. These findings suggest that the risk of T. cruzi infection and Chagas disease is present in the State of Mexico, and further research is needed to identify the T. cruzi bio-types circulating in southern State of Mexico.

Caveolins and flotillin-2 are present in the blood stages of Plasmodium vivax.

Blood stages of Plasmodium vivax induce the development of caveolae and caveola-vesicle complexes (CVC) in the membrane of their host erythrocyte. Caveolae are found in almost all types of cells and are involved in endogenous processes as calcium and cholesterol homeostasis, cell signalling, transporting, ligand internalization and transcytosis of serum components. Major structural components of caveolae are the proteins caveolins and flotillins. The functional role of caveolae in the P. vivax-infected erythrocyte is not properly understood. As these organelles have been shown to contain malaria antigens, it has been suggested that they are involved in the transport and release of specific parasite antigens from the infected erythrocyte and in the uptake of plasma proteins. Using specific antibodies to classical caveolae proteins and an immunolocalization approach, we found caveolin-2, caveolin-3, and flotillin-2 in the vesicle profiles and some CVC of P. vivax-infected erythrocytes. Caveolin-1-3 were not found in uninfected erythrocytes. This is the first report of identification and localization of caveolins in the CVC present in erythrocytes infected with P. vivax, thereby providing evidence of the role of this particular organelle in the protein-trafficking pathway that connect parasite-encoded proteins with the erythrocyte cytoplasm and the cell surface throughout the asexual blood cycle of vivax malaria parasite.

Human Trypanosoma cruzi infection and seropositivity in dogs, Mexico.

We used 5 diagnostic tests in a cross-sectional investigation of the prevalence of Trypanosoma cruzi in Tejupilco municipality, State of Mexico, Mexico. Our findings showed a substantial prevalence of immunoglobulin G (IgG) and IgM antibodies to T. cruzi in human (n = 293, IgG 2.05%, IgM 5.5%, both 7.1%) and dog (n = 114, IgG 15.8%, IgM 11.4%, both 21%) populations. We also found antibodies to T. cruzi (n = 80, IgG 10%, IgM 15%, both 17.5%) in dogs from Toluca, an area previously considered free of T. cruzi. Our data demonstrate the need for active epidemiologic surveillance programs in these regions. A direct correlation (r2 = 0.955) of seropositivity between humans and dogs suggests that seroanalysis in dogs may help identify the human prevalence of T. cruzi infection in these areas.

Traffic pathways of Plasmodium vivax antigens during intraerythrocytic parasite development.

We investigated the secretory traffic of a Plasmodium vivax antigen (Pv-148) synthesised by the parasite during the blood cycle, exported into the host cell cytosol and then transported to the surface membrane of the infected erythrocyte. Studies of the ultrastructure of erythrocytes infected with P. vivax showed that intracellular schizogony is accompanied by the generation of parasite-induced membrane profiles in the erythrocyte cytoplasm. These structures are detectable soon after the parasite invades the erythrocyte and develop an elaborate organisation, leading to a tubovesicular membrane (TVM) network, in erythrocytes infected with mature trophozoites. Interestingly, the clefts formed stacked, flattened cisternae resembling a classical Golgi apparatus. The TVM network stained with the fluorescent Golgi marker Bodipy-ceramide. Specific immunolabelling showed that Pv-148 was transferred from the parasite to the erythrocyte surface membrane via the clefts and the TVM network. These findings suggest that the TVM network is part of the secretory pathways involved in parasite protein transport across the Plasmodium-infected erythrocyte and that Pv- 148 may represent a marker that links the parasite with the host cell cytoplasm and, in turn, with the extracellular milieu.

[Biotic and abiotic determinants of seroprevalence of antibodies against Trypanosoma cruzi in Palmar de Bravo, Puebla, Mexico]. / Factores bióticos y abióticos que determinan la seroprevalencia de anticuerpos contra Trypanosoma cruzi en el municipio de Palmar de Bravo, Puebla, México.

OBJECTIVE: To establish the relationship between seroprevalence for antibodies against Trypanosoma cruzi and its relationship with biotic and abiotic factors. MATERIAL AND METHODS: A cross-sectional study was conducted between August 2000 and September 2001. The study population consisted of a simple random sample of 390 volunteers residing in Palmar de Bravo, Puebla, Mexico. Sample and data collection procedures included assaying antibodies against T. cruzi with validated assays, and searching for domestic reservoirs and triatomine bugs. The relationship between biotic and abiotic factors with seropositivity was assessed. Statistical analysis was conducted using Kappa values for diagnostic tests; statistical significance was assessed with 2 x 2 tables, chi-squared test with Yates' correction, Fisher exact test, and odds ratios. RESULTS: The seroprevalence of T. cruzi infection in humans was 4%; in domestic reservoirs (horses, pigs, and dogs) only 10% of canine reservoirs were positive. Vector species recognized were T. borberi and T. pallidipennis, with a Dispersion Area Index and a Colonization Index of 55% and 40%, respectively. The most important risk factors associated with positive serology were altitude (>2,150 and <2,180 meters above sea level), presence of triatomines, age, time of residence, and participation in a social assistance program. CONCLUSIONS: T. cruzi infection was identified in human beings, vectors, and possibly in domestic reservoirs, in communities located over 2,000 meters above sea level.