More productive in vitro culture of Cryptosporidium parvum for better study of the intra- and extracellular phases

The great difficulties in treating people and animals suffering from cryptosporidiosis have prompted the development of in vitro experimental models. Due to the models of in vitro culture, new extracellular stages of Cryptosporidium have been demonstrated. The development of these extracellular phases depends on the technique of in vitro culture and on the species and genotype of Cryptosporidium used. Here, we undertake the molecular characterization by polymerase chain reaction-restriction fragment lenght polymorphism of different Cryptosporidium isolates from calves, concluding that all are C. parvum of cattle genotype, although differing in the nucleotide at positions 472 and 498. Using these parasites, modified the in vitro culture technique for HCT-8 cells achieving greater multiplication of parasites. The HCT-8 cell cultures, for which the culture had not been renewed in seven days, were infected with C. parvum sporozoites in RPMI-1640 medium with 10% IFBS, CaCl2 and MgCl2 1 mM at pH 7.2. Percentages of cell parasitism were increased with respect to control cultures (71% at 48 h vs 14.5%), even after two weeks (47% vs 1.9%). Also, the percentage of extracellular stages augmented (25.3% vs 1.1% at 96 h). This new model of in vitro culture of C. parvum will enable easier study of the developmental phases of C. parvum in performing new chemotherapeutic assays

Sensitivity and representativeness of One-Health surveillance for diseases of zoonotic potential at health facilities relative to household visits in rural Guatemala

Most human and animal disease notification systems are unintegrated and passive, resulting in underreporting. Active surveillance can complement passive efforts, but because they are resource-intensive, their attributes must be evaluated. We assessed the sensitivity and representativeness of One-Health surveillance conducted at health facilities compared to health facilities plus monthly household visits in three rural communities of Guatemala. From September 2017 to November 2018, we screened humans for acute diarrheal, febrile and respiratory infectious syndromes and canines, swine, equines and bovines for syndromic events or deaths. We estimated the relative sensitivity as the incidence rate ratio of detecting an event in health facility surveillance compared to household surveillance from Poisson models. We used interaction terms between the surveillance method and sociodemographic factors or time trends to assess effect modification as a measure of relative representativeness. We used generalized additive models with smoothing splines to model incidence over time by surveillance method. We randomized 216 households to health facility surveillance and 198 to health facility surveillance plus monthly household visits. Health facility surveillance alone was less sensitive than when combined with household surveillance by 0.42 (95% CI: 0.34, 0.53), 0.56 (95% CI: 0.39, 0.79), 0.02 (95% CI: 0.00, 0.10), 0.28 (95% CI: 0.15, 0.50) and 0.22 (95% CI: 0.03, 0.92) times for human acute infections, human severe acute infections, and deaths in canines, swine and equines, respectively. Health facility surveillance alone underrepresented Spanish speakers (interaction p-value = 0.0003) and persons in higher economic assets (interaction p-values = 0.0008). The trend in incidence over time was different between the two study groups, with a larger decrease in the group with household surveillance (all interaction p-values <0.10). Surveillance at health facilities under ascertains syndromes in humans and animals which leads to underestimation of the burden of zoonotic disease. The magnitude of under ascertainment was differentially by sociodemographic factors, yielding an unrepresentative sample of health events. However, it is less time-intensive, thus might be sustained over time longer than household surveillance. The choice between methodologies should be evaluated against surveillance goals and available resources

Community-based sampling methods for surveillance of the Chagas disease vector, Triatoma dimidiata (Hemiptera: Reduviidae: Triatominae).

In Guatemala, the most widespread vector of Trypanosoma cruzi (Kinetoplastida: Trypanosomatidae), the causative agent of Chagas disease, is Triatoma dimidiata (Latreille) (Hemiptera: Reduviidae: Triatominae). T. dimidiata is native to Guatemala and is present in both domestic and sylvatic habitats. Consequently, control of T. dimidiata is difficult because after successful elimination from homes, individual insects can recolonize homes from the surrounding environment. Therefore, intensive long-term surveillance of this species is essential to ensure adequate control is achieved. Manual inspection for signs of infestation, the current method used to monitor Triatominae throughout Central and South America, is labor and time-consuming, so cost-effective alternatives are needed. The current study compared the effectiveness of the current method of surveillance of T. dimidiata with community-based techniques of G6mez-Nuñez sensor boxes, collection and observation of bugs by householders, and presence of triatomine-like feces on walls. Although manual inspection was the most sensitive method when used alone, collection by householders also was sensitive and specific and involved less effort. Sensor boxes were not sensitive indicators of T. dimidiata infestation when used alone. Two recorded variables, visual inspection for feces and the sighting of bugs by householders, were sensitive and specific indicators of infestation, and in combination with collection by householders and sensor boxes these methods were significantly more likely to detect infestations than manual inspection alone. A surveillance program that combines multiple community-based techniques should have low cost and involve minimal effort from the government and at the same time promote sustainable community involvement in disease prevention

Risk factors for domestic infestation by the Chagas disease vector, Triatoma dimidiata in Chiquimula, Guatemala.

In Guatemala prior to control initiatives, the main vectors of Trypanosoma cruzi, the causative agent of Chagas disease, were Rhodnius prolixus and Triatoma dimidiata. This study conducted in 2006 in the department of Chiquimula recorded a high level of T. dimidiata infestation and an absence of R. prolixus in all surveyed communities. In Guatemala, the presence of T. dimidiata as domestic, peridomestic and sylvatic populations results in control difficulties as houses are re-infested from the surrounding environment. Entomological surveys, the current method used to select houses in need of control efforts, are labour intensive and time consuming. A time- and cost-effective way to prioritize houses for evaluation and subsequent treatment is the stratification of houses based on the risk of triatomine infestation. In the present study, 17 anthropogenic risk factors were evaluated for associations with house infestation of T. dimidiata including: wall, floor and roof type. There was an increased likelihood of domestic infestation with T. dimidiata associated with the presence of dirt floors (18/29; OR 8.075, 95% CI 2.13-30.6), uncoated bajareque walls (12/17; OR 4.80, 95% CI 1.35-17.1) and triatomine-like faeces on walls (16/26; OR 3.89, 95% CI 1.19-12.7). These factors could be used to target control of T. dimidiata to communities with an increased risk of being infested

Impact of residual spraying on Rhodnius prolixus and Triatoma dimidiata in the department of Zacapa in Guatemala

As a vector control program to control Chagas disease in Guatemala, residual spraying of Rhodnius prolixus and Triatoma dimidiata was performed, and its impact was measured in the department of Zacapa. In order to identify infested villages and determine the degree of infestation, a baseline entomological survey to identify municipalities infested with vectors followed by an additional vector survey in areas known to be infested was conducted. Residual spraying using pyrethroid insecticides was performed at all the villages identified as being infested with the vectors. The residual spraying was shown to be highly effective against both vectors by the decrease in infestation indices after spraying. Analysis of the cost-effectiveness of the spraying showed that the average cost of insecticides per house is high when compared with that in Southern Cone countries