Pupillary Light Reflexes are Associated with Autonomic Dysfunction in Bolivian Diabetics But Not Chagas Disease Patients.

Autonomic dysfunction is common in Chagas disease and diabetes. Patients with either condition complicated by cardiac autonomic dysfunction face increased mortality, but no clinical predictors of autonomic dysfunction exist. Pupillary light reflexes (PLRs) may identify such patients early, allowing for intensified treatment. To evaluate the significance of PLRs, adults were recruited from the outpatient endocrine, cardiology, and surgical clinics at a Bolivian teaching hospital. After testing for Chagas disease and diabetes, participants completed conventional autonomic testing (CAT) evaluating their cardiovascular responses to Valsalva, deep breathing, and orthostatic changes. PLRs were measured using specially designed goggles, then CAT and PLRs were compared as measures of autonomic dysfunction. This study analyzed 163 adults, including 96 with Chagas disease, 35 patients with diabetes, and 32 controls. PLRs were not significantly different between Chagas disease patients and controls. Patients with diabetes had longer latency to onset of pupil constriction, slower maximum constriction velocities, and smaller orthostatic ratios than nonpatients with diabetes. PLRs correlated poorly with CAT results. A PLR-based clinical risk score demonstrated a 2.27-fold increased likelihood of diabetes complicated by autonomic dysfunction compared with the combination of blood tests, CAT, and PLRs (sensitivity 87.9%, specificity 61.3%). PLRs represent a promising tool for evaluating subclinical neuropathy in patients with diabetes without symptomatic autonomic dysfunction. Pupillometry does not have a role in the evaluation of Chagas disease patients.

Trypanosoma cruzi-infected pregnant women without vector exposure have higher parasitemia levels: implications for congenital transmission risk.

BACKGROUND: Congenital transmission is a major source of new Trypanosoma cruzi infections, and as vector and blood bank control continue to improve, the proportion due to congenital infection will grow. A major unanswered question is why reported transmission rates from T. cruzi-infected mothers vary so widely among study populations. Women with high parasite loads during pregnancy are more likely to transmit to their infants, but the factors that govern maternal parasite load are largely unknown. Better understanding of these factors could enable prioritization of screening programs to target women most at risk of transmission to their infants. METHODOLOGY/PRINCIPAL FINDINGS: We screened pregnant women presenting for delivery in a large urban hospital in Bolivia and followed infants of infected women for congenital Chagas disease. Of 596 women screened, 128 (21.5%) had confirmed T. cruzi infection; transmission occurred from 15 (11.7%) infected women to their infants. Parasite loads were significantly higher among women who transmitted compared to those who did not. Congenital transmission occurred from 31.3% (9/29), 15.4% (4/26) and 0% (0/62) of women with high, moderate and low parasite load, respectively (χx2 for trend 18.2; p<0.0001). Twin births were associated with higher transmission risk and higher maternal parasite loads. Infected women without reported vector exposure had significantly higher parasite loads than those who had lived in an infested house (median 26.4 vs 0 parasites/mL; p<0.001) with an inverse relationship between years of living in an infested house and parasite load. CONCLUSIONS/SIGNIFICANCE: We hypothesize that sustained vector-borne parasite exposure and repeated superinfection by T. cruzi may act as an immune booster, allowing women to maintain effective control of the parasite despite the down-regulation of late pregnancy.

Use of a novel chagas urine nanoparticle test (chunap) for diagnosis of congenital chagas disease.

BACKGROUND: Detection of congenital T. cruzi transmission is considered one of the pillars of control programs of Chagas disease. Congenital transmission accounts for 25% of new infections with an estimated 15,000 infected infants per year. Current programs to detect congenital Chagas disease in Latin America utilize microscopy early in life and serology after 6 months. These programs suffer from low sensitivity by microscopy and high loss to follow-up later in infancy. We developed a Chagas urine nanoparticle test (Chunap) to concentrate, preserve and detect T. cruzi antigens in urine for early, non-invasive diagnosis of congenital Chagas disease. METHODOLOGY/PRINCIPAL FINDINGS: This is a proof-of-concept study of Chunap for the early diagnosis of congenital Chagas disease. Poly N-isopropylacrylamide nano-particles functionalized with trypan blue were synthesized by precipitation polymerization and characterized with photon correlation spectroscopy. We evaluated the ability of the nanoparticles to capture, concentrate and preserve T. cruzi antigens. Urine samples from congenitally infected and uninfected infants were then concentrated using these nanoparticles. The antigens were eluted and detected by Western Blot using a monoclonal antibody against T. cruzi lipophosphoglycan. The nanoparticles concentrate T. cruzi antigens by 100 fold (western blot detection limit decreased from 50 ng/ml to 0.5 ng/ml). The sensitivity of Chunap in a single specimen at one month of age was 91.3% (21/23, 95% CI: 71.92%-98.68%), comparable to PCR in two specimens at 0 and 1 month (91.3%) and significantly higher than microscopy in two specimens (34.8%, 95% CI: 16.42%-57.26%). Chunap specificity was 96.5% (71/74 endemic, 12/12 non-endemic specimens). Particle-sequestered T. cruzi antigens were protected from trypsin digestion. CONCLUSION/SIGNIFICANCE: Chunap has the potential to be developed into a simple and sensitive test for the early diagnosis of congenital Chagas disease.

Field evaluation of the InBios Chagas detect plus rapid test in serum and whole-blood specimens in Bolivia.

Trypanosoma cruzi causes Chagas disease, which affects an estimated 7 million to 8 million people. Chagas disease is endemic throughout Latin America, with the highest prevalence in Bolivia. Conventional diagnosis requires a well-equipped laboratory with experienced personnel. We evaluated the Chagas Detect Plus (CDP) (InBios, Seattle, WA), a rapid immunochromatographic assay for IgG antibodies to T. cruzi. CDP performance was compared to infection status based on results obtained by indirect hemagglutination assay, immunofluorescent-antibody test, and enzyme-linked immunosorbent assay. Confirmed infection required positive results by at least 2 conventional assays. We used specimens from adults of both sexes in a general hospital in the city of Santa Cruz and from pregnant women in a hospital and children in villages in the Bolivian Chaco, an area of hyperendemicity. CDP was performed in paired whole-blood and serum specimens from 385 individuals in the two hospital studies and in 200 serum specimens from the community study. CDP showed sensitivities/specificities of 96.2% (95% confidence interval, 92.7 to 98.4)/98.8% (95.9 to 99.9) in whole blood and 99.3% (97.5 to 99.9)/96.9% (94.2 to 98.6) in serum, with no differences by sex, age group, or study site. CDP showed excellent sensitivity and specificity in our study population, comparable to those of conventional serology. The test is reliable for field surveys, requires no laboratory equipment, and performed well in serum and whole blood. The CDP could also be used for accurate maternal screening to identify neonates at risk of congenital transmission. CDP performance data in diverse geographic areas are needed to strengthen the evidence base for its use.

Regional variation in the correlation of antibody and T-cell responses to Trypanosoma cruzi.

Chagas disease, caused by the protozoan parasite Trypanosoma cruzi, is a major cause of morbidity and mortality in Central and South America. Geographic variations in the sensitivity of serologic diagnostic assays to T. cruzi may reflect differences in T. cruzi exposure. We measured parasite-specific T-cell responses among seropositive individuals in two populations from South America with widely varying antibody titers against T. cruzi. Antibody titers among seropositive individuals were significantly lower in Arequipa, Peru compared with Santa Cruz, Bolivia. Similarly, the proportion of seropositive individuals with positive T-cell responses was lower in Peru than Bolivia, resulting in overall lower frequencies of interferon-γ (IFNγ)-secreting cells from Peruvian samples. However, the magnitude of the IFNγ response was similar among the IFNγ responders in both locations. These data indicate that immunological discrepancies based on geographic region are reflected in T-cell responses as well as antibody responses.

Epidemiology of and impact of insecticide spraying on Chagas disease in communities in the Bolivian Chaco.

BACKGROUND: Chagas disease control campaigns relying upon residual insecticide spraying have been successful in many Southern American countries. However, in some areas, rapid reinfestation and recrudescence of transmission have occurred. METHODOLOGY/PRINCIPAL FINDINGS: We conducted a cross-sectional survey in the Bolivian Chaco to evaluate prevalence of and risk factors for T. cruzi infection 11 years after two rounds of blanket insecticide application. We used a cubic B-spline model to estimate change in force of infection over time based on age-specific seroprevalence data. Overall T. cruzi seroprevalence was 51.7%. The prevalence was 19.8% among children 2-15, 72.7% among those 15-30 and 97.1% among participants older than 30 years. Based on the model, the estimated annual force of infection was 4.3% over the two years before the first blanket spray in 2000 and fell to 0.4% for 2001-2002. The estimated annual force of infection for 2004-2005, the 2 year period following the second blanket spray, was 4.6%. However, the 95% bootstrap confidence intervals overlap for all of these estimates. In a multivariable model, only sleeping in a structure with cracks in the walls (aOR = 2.35; 95% CI = 1.15-4.78), age and village of residence were associated with infection. CONCLUSIONS/SIGNIFICANCE: As in other areas in the Chaco, we found an extremely high prevalence of Chagas disease. Despite evidence that blanket insecticide application in 2000 may have decreased the force of infection, active transmission is ongoing. Continued spraying vigilance, infestation surveillance, and systematic household improvements are necessary to disrupt and sustain interruption of infection transmission.

Chagas cardiomyopathy in the context of the chronic disease transition.

BACKGROUND: Patients with Chagas disease have migrated to cities, where obesity, hypertension and other cardiac risk factors are common. METHODOLOGY/PRINCIPAL FINDINGS: The study included adult patients evaluated by the cardiology service in a public hospital in Santa Cruz, Bolivia. Data included risk factors for T. cruzi infection, medical history, physical examination, electrocardiogram, echocardiogram, and contact 9 months after initial data collection to ascertain mortality. Serology and PCR for Trypanosoma cruzi were performed. Of 394 participants, 251 (64%) had confirmed T. cruzi infection by serology. Among seropositive participants, 109 (43%) had positive results by conventional PCR; of these, 89 (82%) also had positive results by real time PCR. There was a high prevalence of hypertension (64%) and overweight (body mass index [BMI] >25; 67%), with no difference by T. cruzi infection status. Nearly 60% of symptomatic congestive heart failure was attributed to Chagas cardiomyopathy; mortality was also higher for seropositive than seronegative patients (p = 0.05). In multivariable models, longer residence in an endemic province, residence in a rural area and poor housing conditions were associated with T. cruzi infection. Male sex, increasing age and poor housing were independent predictors of Chagas cardiomyopathy severity. Males and participants with BMI

Congenital Trypanosoma cruzi transmission in Santa Cruz, Bolivia.

BACKGROUND: We conducted a study of congenital Trypanosoma cruzi infection in Santa Cruz, Bolivia. Our objective was to apply new tools to identify weak points in current screening algorithms, and find ways to improve them. METHODS: Women presenting for delivery were screened by rapid and conventional serological tests. For infants of infected mothers, blood specimens obtained on days 0, 7, 21, 30, 90, 180, and 270 were concentrated and examined microscopically; serological tests were performed for the day 90, 180, and 270 specimens. Maternal and infant specimens, including umbilical tissue, were tested by polymerase chain reaction (PCR) targeting the kinetoplast minicircle and by quantitative PCR. RESULTS: Of 530 women, 154 (29%) were seropositive. Ten infants had congenital T. cruzi infection. Only 4 infants had positive results of microscopy evaluation in the first month, and none had positive cord blood microscopy results. PCR results were positive for 6 (67%) of 9 cord blood and 7 (87.5%) of 8 umbilical tissue specimens. PCR-positive women were more likely to transmit T. cruzi than were seropositive women with negative PCR results (P < .05). Parasite loads determined by quantitative PCR were higher for mothers of infected infants than for seropositive mothers of uninfected infants P < .01). Despite intensive efforts, only 58% of at-risk infants had a month 9 specimen collected. CONCLUSIONS: On the basis of the low sensitivity of microscopy in cord blood and high rate of loss to follow-up, we estimate that current screening programs miss one-half of all infected infants. Molecular techniques may improve early detection.

Geographic variation in the sensitivity of recombinant antigen-based rapid tests for chronic Trypanosoma cruzi infection.

Chagas disease affects 8-11 million people throughout the Americas. Early detection is crucial for timely treatment and to prevent non-vectorial transmission. Recombinant antigen-based rapid tests had high sensitivity and specificity in laboratory evaluations, but no Peruvian specimens were included in previous studies. We evaluated Stat-Pak and Trypanosoma Detect rapid tests in specimens from Bolivia and Peru. Specimens positive by three conventional assays were confirmed positives; specimens negative by two or more assays were confirmed negatives. In Bolivian specimens, Stat-Pak and Trypanosoma Detect tests were 87.5% and 90.7% sensitive, respectively; both showed 100% specificity. Sensitivity in Peruvian specimens was much lower: 26.6-33.0% (Stat-Pak) and 54.3-55.2% (Trypanosoma Detect); both had specificities > 98%. Even in Bolivian specimens, these sensitivities are inadequate for stand-alone screening. The low sensitivity in Peru may be related to parasite strain differences. Chagas disease rapid tests should be field tested in each geographic site before widespread implementation for screening.

Polymerase chain reaction for chronic Trypanosoma cruzi infection yields higher sensitivity in blood clot than buffy coat or whole blood specimens.

Trypanosoma cruzi polymerase chain reaction (PCR) is widely used, but sensitivity varies widely. We compared PCR using 121/122 primers targeting kinetoplast minicircle DNA in whole blood, buffy coat, and clot from Bolivian women. Sensitivity was significantly higher in clot (60.1%) than buffy coat (46.5%) or whole blood (40%). The use of clot could simplify specimen collection while improving sensitivity.