Deadly and venomous Lonomia caterpillars are more than the two usual suspects.

Caterpillars of the Neotropical genus Lonomia (Lepidoptera: Saturniidae) are responsible for some fatal envenomation of humans in South America inducing hemostatic disturbances in patients upon skin contact with the caterpillars' spines. Currently, only two species have been reported to cause hemorrhagic syndromes in humans: Lonomia achelous and Lonomia obliqua. However, species identifications have remained largely unchallenged despite improved knowledge of venom diversity and growing evidence that the taxonomy used over past decades misrepresents and underestimates species diversity. Here, we revisit the taxonomic diversity and distribution of Lonomia species using the most extensive dataset assembled to date, combining DNA barcodes, morphological comparisons, and geographical information. Considering new evidence for seven undescribed species as well as three newly proposed nomenclatural changes, our integrative approach leads to the recognition of 60 species, of which seven are known or strongly suspected to cause severe envenomation in humans. From a newly compiled synthesis of epidemiological data, we also examine the consequences of our results for understanding Lonomia envenomation risks and call for further investigations of other species' venom activities. This is required and necessary to improve alertness in areas at risk, and to define adequate treatment strategies for envenomed patients, including performing species identification and assessing the efficacy of anti-Lonomia serums against a broader diversity of species.

Pathogens, reservoirs, and vectors involved in the transmission of vector-borne and zoonotic diseases in a Colombian region.

The ecology of vector-borne diseases (VBDs) is an important system of great complexity, which involves the knowledge about the pathogens and animal species entailed in maintaining transmission cycles in a given locality, including those that act as vectors and reservoirs for the transmitted pathogens. To understand the ecology of some VBDs, we studied vectors, reservoirs, and pathogens of different VBDs, including dengue, leishmaniasis, Chagas disease, malaria, Zika, and chikungunya in the municipality of La Mesa, Cundinamarca, Colombia, a locality close to the capital, Bogotá. Vectors and mammals were sampled in urban and rural areas between May and August 2019. Molecular analyses were performed for the detection of pathogens in mammals and vectors, and of blood-meal sources in insects. Several vectors and mammals collected in this study have been involved in pathogen transmission cycles or may have a potential role in them. The findings of this study suggest that in the municipality of La Mesa, there are both vector and potential reservoir species, which are or could be implicated in the maintenance of the cycles of vector-borne diseases such as leishmaniasis and Chagas disease. Although arbovirus infections, such as dengue, are reported in the municipality, arbovirus presence was not detected. These findings highlight the importance of ongoing surveillance of vectors and associated control operations in La Mesa, of relevance to other locations where vectors and animal hosts also occur.

Morphological and molecular characterization of the parasite Dipylidium caninum infecting an infant in Colombia: a case report.

BACKGROUND: Dipylidium caninum is the causal agent of dipylidiasis affecting mainly cats and dogs worldwide. Human cases of dipylidiasis are rare, and the diagnosis is prevalently based on morphological features of the parasite. Here we report the diagnosis of dipylidiasis through morphological and molecular characterization of D. caninum infecting an 11-month-old boy in Cajicá, Colombia. METHODS: Fresh faecal samples were obtained from the infant, and morphological identification of the parasite was performed through faecal smears. DNA was extracted from proglottids and used in PCR analyses for amplification of a 653-bp fragment of the nuclear ribosomal RNA (rRNA) encoding the 28S rRNA gene. A phylogeny study to better characterize the obtained DNA sequence was inferred using the maximum likelihood method and the Tamura-Nei model. RESULTS: After morphological and molecular analyses, D. caninum was identified as the etiological agent causing the infection in the infant. Results of phylogenetical analyses showed that the obtained sequence clusters within the feline genotype clade. After the diagnosis of the parasite, effective treatment with praziquantel was administered to the infant. CONCLUSIONS: This is the third human case of dipylidiasis reported in Colombia, and the first study in South America to provide a molecular identification of D. caninum.

Sequencing of hsp70 for discernment of species from the Leishmania (Viannia) guyanensis complex from endemic areas in Colombia.

BACKGROUND: Colombia is ranked very high among countries with the highest numbers of endemic Leishmania species (n = 9) causing human disease. Although much effort has been devoted to generating simple and specific tools for Leishmania species identification, challenges remain in the discrimination of species belonging to the Leishmania (Viannia) guyanensis complex: L. (V.) guyanensis and L. (V.) panamensis. METHODS: A set of seven reference strains of species belonging to the L. (Leishmania) and L. (Viannia) subgenera, clinical strains from human cases of cutaneous leishmaniasis (CL; n = 26) and samples collected from sylvatic mammals and sand flies (n = 7) from endemic areas in Colombia were analyzed in this study. The heat-shock protein 70 gene (hsp70) was amplified by PCR from DNA extracted from logarithmic-phase promastigotes or tissue samples, and the PCR products were sequenced. Sequence alignment was performed against a set of previously published and curated sequences, and phylogenetic analysis based on the maximum-likelihood and Bayesian inference approaches was conducted. Haplotype diversity among strains and species of the L. (V.) guyanensis complex was explored using a median-joining network. RESULTS: Sequencing of the hsp70 gene for L. (Viannia) spp. typing was comparable to species identification using isoenzyme electrophoresis or monoclonal antibodies. Complete species matching was found, except for one sylvatic sample with an identity yet unsolved. Among the L. (V.) panamensis clinical strains, two distinctive phylogenetic clusters were found to correlate with two different zymodemes: L. (V.) panamensis Z2.2 and Z2.3. Analysis of samples from sylvatic environments identified novel records of naturally infected wild mammal and sand fly species. CONCLUSIONS: Our results support the adequacy of hsp70 gene sequencing as a single-locus approach for discrimination of L. (Viannia) spp., as well as for exploring the genetic diversity within the L. (V.) guyanensis complex.

Multicenter international assessment of a SARS-CoV-2 RT-LAMP test for point of care clinical application.

Continued waves, new variants, and limited vaccine deployment mean that SARS-CoV-2 tests remain vital to constrain the coronavirus disease 2019 (COVID-19) pandemic. Affordable, point-of-care (PoC) tests allow rapid screening in non-medical settings. Reverse-transcription loop-mediated isothermal amplification (RT-LAMP) is an appealing approach. A crucial step is to optimize testing in low/medium resource settings. Here, we optimized RT-LAMP for SARS-CoV-2 and human ß-actin, and tested clinical samples in multiple countries. "TTTT" linker primers did not improve performance, and while guanidine hydrochloride, betaine and/or Igepal-CA-630 enhanced detection of synthetic RNA, only the latter two improved direct assays on nasopharygeal samples. With extracted clinical RNA, a 20 min RT-LAMP assay was essentially as sensitive as RT-PCR. With raw Canadian nasopharygeal samples, sensitivity was 100% (95% CI: 67.6% - 100%) for those with RT-qPCR Ct values ≤ 25, and 80% (95% CI: 58.4% - 91.9%) for those with 25 < Ct ≤ 27.2. Highly infectious, high titer cases were also detected in Colombian and Ecuadorian labs. We further demonstrate the utility of replacing thermocyclers with a portable PoC device (FluoroPLUM). These combined PoC molecular and hardware tools may help to limit community transmission of SARS-CoV-2.

Entomological characterization of Aedes mosquitoes and arbovirus detection in Ibagué, a Colombian city with co-circulation of Zika, dengue and chikungunya viruses.

BACKGROUND: Dengue, Zika and chikungunya are arboviruses of significant public health importance that are transmitted by Aedes aegypti and Aedes albopictus mosquitoes. In Colombia, where dengue is hyperendemic, and where chikungunya and Zika were introduced in the last decade, more than half of the population lives in areas at risk. The objective of this study was to characterize Aedes spp. vectors and study their natural infection with dengue, Zika and chikungunya in Ibagué, a Colombian city and capital of the department of Tolima, with case reports of simultaneous circulation of these three arboviruses. METHODS: Mosquito collections were carried out monthly between June 2018 and May 2019 in neighborhoods with different levels of socioeconomic status. We used the non-parametric Friedman, Mann-Whitney and Kruskal-Wallis tests to compare mosquito density distributions. We applied logistic regression analyses to identify associations between mosquito density and absence/presence of breeding sites, and the Spearman correlation coefficient to analyze the possible relationship between climatic variables and mosquito density. RESULTS: We collected Ae. aegypti in all sampled neighborhoods and found for the first time Ae. albopictus in the city of Ibagué. A greater abundance of mosquitoes was collected in neighborhoods displaying low compared to high socioeconomic status as well as in the intradomicile compared to the peridomestic space. Female mosquitoes predominated over males, and most of the test females had fed on human blood. In total, four Ae. aegypti pools (3%) were positive for dengue virus (serotype 1) and one pool for chikungunya virus (0.8%). Interestingly, infected females were only collected in neighborhoods of low socioeconomic status, and mostly in the intradomicile space. CONCLUSIONS: We confirmed the co-circulation of dengue (serotype 1) and chikungunya viruses in the Ae. aegypti population in Ibagué. However, Zika virus was not detected in any mosquito sample, 3 years after its introduction into the country. The positivity for dengue and chikungunya viruses, predominance of mosquitoes in the intradomicile space and the high proportion of females fed on humans highlight the high risk for arbovirus transmission in Ibagué, but may also provide an opportunity for establishing effective control strategies.

Host selection pattern and flavivirus screening of mosquitoes in a disturbed Colombian rainforest.

Studies on the feeding behavior of hematophagous insects, particularly those of medical importance, are relevant for tracking possible pathogen transmission routes and identifying biases in the choice of vertebrates. We evaluated host selection of blood-feeding mosquitoes in a disturbed forest in the Magdalena Medio valley in Colombia from March 2017 to April 2018, after the introduction of Zika virus to the Americas from the 2015-2016 outbreak. We estimated vertebrate diversity and collected blood-engorged female mosquitoes. Genomic DNA/RNA was extracted from the mosquito's abdomen for vertebrate host identification and pathogen detection. We performed conventional PCR and sequencing, using universal primers targeting vertebrate regions of the eukaryotic mitochondrial genome to determine bloodmeal host. Additionally, we tested for the presence of flaviviruses in all mosquito samples with RT-PCR. Based on the identity and quantity of detected bloodmeals, we performed mosquito-vertebrate interaction network analysis and estimated topology metrics. In total, we collected 292 engorged female mosquitoes representing 20 different species. Bloodmeal analyses identified 26 vertebrate species, the majority of which were mammals (N = 16; 61.5%). No flaviviruses of medical importance were detected from the samples. Although feeding patterns varied, network analyses showed a high degree of specialization by mosquitoes and revealed ecological and phylogenetic relationships among the host community. We conclude that host selection or preference by mosquitoes is species specific.

Measuring spatial co-occurrences of species potentially involved in Leishmania transmission cycles through a predictive and fieldwork approach.

The Leishmaniases are a group of neglected tropical diseases caused by different species of the protozoan parasite Leishmania, transmitted to its mammalian hosts by the bites of several species of female Phlebotominae sand flies. Many factors have contributed to shifts in the disease distribution and eco epidemiological outcomes, resulting in the emergence of Cutaneous Leishmaniasis outbreaks and the incrimination of vectors in unreported regions. New research development is vital for establishing the new paradigms of the present transmission cycles, hoping to facilitate new control strategies to reduce parasite transmission. Hereafter, this work aims to model and infer the current transmission cycles of Cutaneous Leishmaniasis in Colombia defined by vector and mammal species distributed and interacting in the different regions and validate them by performing sand fly and mammal collections. Vector-host co-occurrences were computed considering five ecoregions of the Colombian territory defined by the World Wide Fund for Nature (WWF) and downloaded from The Nature Conservancy TNC Maps website. Four validation sites were selected based on Cutaneous Leishmaniasis prevalence reports. Sand flies and mammals captured in the field were processed, and species were defined using conventional taxonomic guidelines. Detection of infection by Leishmania was performed to identify transmission cycles in the selected areas. This study uses predictive models based on available information from international gazetteers and fieldwork to confirm sand fly and mammalian species' sustaining Leishmania transmission cycles. Our results show an uneven distribution of mammal samples in Colombia, possibly due to sampling bias, since only two departments contributed 50% of the available samples. Bats were the vertebrates with the highest score values, suggesting substantial spatial overlap with sand flies than the rest of the vertebrates evaluated. Fieldwork allowed identifying three circulating Leishmania species, isolated from three sand fly species. In the Montane Forest ecosystem, one small marsupial, Gracilinanus marica, was found infected with Leishmania panamensis, constituting the first record of this species infected with Leishmania. In the same locality, an infected sand fly, Pintomyia pia, was found. The overall results could support the understanding of the current transmission cycles of Leishmaniasis in Colombia.

How microclimatic variables and blood meal sources influence Rhodnius prolixus abundance and Trypanosoma cruzi infection in Attalea butyracea and Elaeis guineensis palms?

Chagas disease is a zoonosis that affects several million people and is caused by the parasite Trypanosoma cruzi, which is mainly transmitted through the feces of triatomine bugs. Within triatomines, several Rhodnius species have been found inhabiting palms, and certain factors such as palm species and location have been related to the abundance and T. cruzi infection of those insects in palms. In this study, the main goal was to determine if R. prolixus abundances and infection rates in Attalea butyracea and Elaeis guineensis palms are related to ecological factors such as palm species, crown microclimate, and available blood meal sources. Triatomine sampling was performed in two municipalities of Casanare, Colombia, specifically in the intersection of riparian forests and oil palm plantations. For R. prolixus abundance per palm, the predictors showing more relationship were palm species and blood meal species identified in the palm, and for T. cruzi infection per triatomine, they were palm species and nymphal stage. Palm microclimate was very similar in both palm species and did not show a relationship with triatomine abundance. Comparing palm species, A. butyracea showed more blood meal species, including more refractory host species, than E. guineensis, but lower T. cruzi infection rate and parasitaemia. Interestingly, non-arboreal blood meal species were frequently found in the analyzed nymphs, indicating that the blood source for R. prolixus in palms corresponded to all the fauna located in the surrounded landscape and not only in the palm. These results could expose a new ecological scenario to interpret the T. cruzi transmission in sylvatic environments.

Ecology of Sand Flies (Psychodidae: Phlebotominae) and Natural Infection of Pintomyia townsendi With Leishmania amazonensis in a Cutaneous Leishmaniasis Focus in Colombia.

Humans have influenced the epidemiological patterns of American cutaneous leishmaniasis by habitat disturbance, which has led to the emergence of new transmission foci. In these transmission areas, detecting natural infection of sand fly species with Leishmania parasites is of prime importance in epidemiological studies. In this study, we examined the species composition, spatial distribution, seasonality, and natural infection with Leishmania of the sand fly fauna in an emergent leishmaniasis focus located in Colombia. Sand flies were collected from September 2014 to June 2015 using CDC light traps located in indoor, peridomestic, and outdoor habitats within areas with confirmed clinical patients. In total, 13,488 sand flies of 12 species and seven genera were collected. Among these, Pintomyia townsendi (Ortiz) was the most abundant species comprising 76.3% of total flies collected. The sand fly richness and abundance were influenced by habitat and weather conditions. Outdoor areas harbored the greatest diversity of sand flies. Rainfall negatively affected abundance, whereas increased temperature showed a low positive effect. We detected Leishmania amazonensis parasites in Pi. townsendi for the first time.

Detection of Trypanosoma cruzi strains circulating in Córdoba department (Colombia) isolated from triatomines (Hemiptera: Reduviidae) collected by the community. / Detección y caracterización molecular de cepas de Trypanosoma cruzi aisladas de triatominos recolectados por la comunidad en el departamento de Córdoba, Colombia

INTRODUCTION: From 2011 to 2016, 24 cases of Chagas disease were reported in Córdoba according to the national public health surveillance system (Sistema Nacional de Vigilancia en Salud Pública, Sivigila), but the information regarding Trypanosoma cruzi circulating strains and infection rates are unknown. OBJECTIVES: To establish the triatomine species with which people come in contact and recognize as Chagas disease vectors, as well as to assess the infection with trypanosomes and make an exploratory approach to host feeding preferences with the participation of the local community. MATERIALS AND METHODS: Triatomines sampling was conducted in 12 municipalities between 2011 and 2016; T. cruzi infection was established by k-PCR, SAT-PCR, while strain genotyping was done by mini-exon and SL-IR (spliced-leader intergenic region) sequence characterization. We also screened for blood sources. RESULTS: Local community members collected the majority of triatomines and we identified three species: Rhodnius pallescens, Panstrongylus geniculatus, and Eratyrus cuspidatus. The overall T. cruzi infection rate in collected triatomines was 66.6% and we detected the TcIDOM and TcI sylvatic strains. Community-based insect collection allowed reporting the presence of P. geniculatus in two new disperse rural settlements, T. cruzi infection of P. geniculatus in Córdoba, and the first report of triatomines infected with T. cruzi in Montería municipality. CONCLUSIONS: These results revealed the presence of triatomines infected with T. cruzi inside dwellings in five municipalities of Córdoba. The dominant circulating T. cruzi strain was TcIDOM, a genotype associated with human Chagas disease and cardiomyopathies in Colombia. Our results highlight the importance of local community participation in entomological surveillance tasks.

Introducción. Entre el 2011 y el 2016, se reportaron 24 casos de enfermedad de Chagas en Córdoba, según el Sistema Nacional de Vigilancia en Salud Pública (Sivigila), pero la información sobre las unidades discretas de tipificación de Trypanosoma cruzi circulantes y las tasas de infección se desconoce. Objetivos. Identificar las especies de triatominos con las cuales las personas entran en contacto y que reconocen como vectores de la enfermedad de Chagas, así como establecer la infección por tripanosomas y explorar posibles fuentes de alimentación de los triatominos con la participación de la comunidad. Materiales y métodos. El muestreo de triatominos se hizo en 12 municipios entre el 2011 y el 2016. T. cruzi se detectó mediante las técnicas de kinetic-polymerase chain reaction (k-PCR) y serial amplification of targets-polymerase chain reaction (SAT-PCR), en tanto que la genotipificación de las cepas se logró mediante la caracterización de secuencias de genes miniexon y de la región intergénica SL-IR (Spliced-Leader Intergenic Region). Se evaluaron, asimismo, las fuentes de alimento. Resultados. La mayoría de los triatominos fue recolectada por miembros de la comunidad y se identificaron tres especies: Rhodnius pallescens, Panstrongylus geniculatus y Eratyrus cuspidatus. La tasa de infección general por T. cruzi fue de 66,6 % y se detectaron las cepas TcIDOM y TcI sylvatic. La participación de la comunidad permitió reportar la presencia de P. geniculatus en dos nuevas localidades, la infección con T. cruzi de P. geniculatus en Córdoba y reportar por primera vez triatominos infectados con T. cruzi en Montería. Conclusiones. Se demostró la presencia de triatominos infectados con T. cruzi dentro de las viviendas en cinco municipalidades. La cepa circulante dominante fue T. cruzi TcIDOM, asociada con la enfermedad de Chagas y con cardiomiopatías en Colombia. Los resultados resaltan la importancia de vincular a miembros de la comunidad en la vigilancia entomológica.

Prevalence of Plasmodium parasites in non-human primates and mosquitoes in areas with different degrees of fragmentation in Colombia.

BACKGROUND: Parasites from the genus Plasmodium, the aetiological agent of malaria in humans, can also infect non-human primates (NHP), increasing the potential risk of zoonotic transmission with its associated global public health concerns. In Colombia, there are no recent studies on Plasmodium spp. infecting free-ranging NHP. Thus, this study aimed to determine the diversity of Plasmodium species circulating in fragmented forests in central Colombia, both in Anopheles mosquitoes and in the four sympatric NHP in the region (Ateles hybridus, Cebus versicolor, Alouatta seniculus and Aotus griseimembra), in order to evaluate the risk of infection to humans associated with the presence of sylvatic hosts and vectors infected with Plasmodium spp. METHODS: Overall, there were collected 166 fecal samples and 25 blood samples from NHP, and 442 individuals of Anopheles spp. DNA extraction, nested PCR using mitochondrial (cox3 gene) and ribosomal (18S rDNA) primers, electrophoresis and sequencing were conducted in order to identify Plasmodium spp. from the samples. RESULTS: Plasmodium falciparum was detected in two fecal samples of Alouatta seniculus, while Plasmodium vivax/simium infected Ateles hybridus, Cebus versicolor and Alouatta seniculus. Co-infections with P. vivax/simium and Plasmodium malariae/brasilianum were found in three individuals. The highest prevalence from blood samples was found for Plasmodium malariae/brasilianum in two Alouatta seniculus while Plasmodium vivax/simium was most prevalent in fecal samples, infecting four individuals of Alouatta seniculus. Seven Anopheles species were identified in the study site: Anopheles (Anopheles) punctimacula, Anopheles (An.) malefactor, Anopheles (Nyssorhynchus) oswaldoi, Anopheles (Nys.) triannulatus, Anopheles (An.) neomaculipalpus, Anopheles (Nys.) braziliensis and Anopheles (Nys.) nuneztovari. Infection with P. vivax/simium was found in An. nuneztovari, An. neomaculipalpus, and An. triannulatus. Furthermore, An. oswaldoi and An. triannulatus were found infected with P. malariae/brasilianum. The effect of fragmentation and distance to the nearest town measured in five forests with different degrees of fragmentation was not statistically significant on the prevalence of Plasmodium in NHP, but forest fragmentation did have an effect on the Minimum Infection Rate (MIR) in Anopheles mosquitoes. CONCLUSIONS: The presence of Plasmodium spp. in NHP and Anopheles spp. in fragmented forests in Colombia has important epidemiological implications in the human-NHP interface and the associated risk of malaria transmission.

Analytical Performance of a Loop-Mediated Isothermal Amplification Assay for Leishmania DNA Detection in Sandflies and Direct Smears of Patients with Cutaneous Leishmaniasis.

Loop-mediated isothermal amplification (LAMP) is ideal for the detection of Leishmania DNA as it is a quick and easy-to-perform test that does not require complex or sophisticated equipment or infrastructure. However, the application of this technique in the detection of Leishmania DNA has not been comprehensively analyzed to date (analytical validation). Our objective was to evaluate the sensitivity and analytical specificity (anticipated reportable range [ARR], the limit of detection [LoD], and accuracy) of LAMP targeting the 18S rRNA gene in the diagnosis of six New World Leishmania species. We then applied the validated LAMP assay across 50 samples of sandflies and 50 direct smears from a recent outbreak of cutaneous leishmaniasis in Colombia to determine its diagnostic performance. The LAMP assay exclusively amplified the DNA of Leishmania spp., and an ARR of between 1 × 104 and 1 × 10-2 equivalent parasites/mL was determined. An LoD of 1 × 10-2 equivalent parasites/mL was established and there was no statistically significant variation in terms of accuracy. Finally, a sensitivity of 100% in direct smears and sandflies samples was calculated and a specificity of 90.9% for direct smears using microscopy as reference and 96.8% for sandflies using real-time polymerase chain reaction as reference were determined. To our knowledge, this is the first attempt to analytically validate a LAMP test to detect Leishmania DNA, which showed good diagnostic potential from sandflies and direct smear samples.

Diversity patterns, Leishmania DNA detection, and bloodmeal identification of Phlebotominae sand flies in villages in northern Colombia.

Leishmaniases are neglected tropical diseases exhibiting complex transmission cycles due to the number of parasite species circulating, sand fly species acting as vectors and infected mammals, including humans, which are defined in the New World as accidental hosts. However, current transmission scenarios are changing, and the disease is no longer exclusively related to forested areas but urban transmission foci occur, involving some species of domestic animals as suspected reservoirs. The aim of this study was to determine the transmission cycles in urban environments by evaluating sand fly diversity, detection of Leishmania DNA, and bloodmeal sources through intra and peridomestic collections. The study was carried out in Colombia, in 13 municipalities of Cordoba department, implementing a methodology that could be further used for the evaluation of vector-borne diseases in villages or towns. Our sampling design included 24 houses randomly selected in each of 15 villages distributed in 13 municipalities, which were sampled in two seasons in 2015 and 2016. Sand flies were collected using CDC light traps placed in intra and peridomestic habitats. In addition to the morphological identification, molecular identification through DNA barcodes was also performed. A total of 19,743 sand flies were collected and 13,848 of them (10,268 females and 3,580 males) were used in molecular procedures. Circulation of two known parasite species-Leishmania infantum and Leishmania panamensis was confirmed. Blood source analyses showed that sand flies fed on humans, particularly in the case of the known L. infantum vector, P. evansi; further analyses are advised to evaluate the reservoirs involved in parasite transmission. Our sampling design allowed us to evaluate potential transmission cycles on a department scale, by defining suspected vector species, parasite species present in different municipalities and feeding habits.

First Colombian consensus on congenital Chagas and clinical approach for women of child-bearing age diagnosed with Chagas / Primer consenso colombiano sobre Chagas congénito y orientación clínica a mujeres en edad fértil con diagnóstico de Chagas

Congenital transmission of Chagas disease has not been extensively studied in Colombia, and there are no standardized processes in the health system regarding the specific diagnosis, treatment and follow-up of this disease. To generate recommendations on congenital Chagas disease and Chagas in women of childbearing age in Colombia, a consensus of experts was developed. An extensive literature search through the Medline database was carried out using the MeSH terms: «Chagas disease/congenital¼, «prevention and control¼, «diagnosis¼, «therapeutics¼ and «pregnancy¼. Appropriate abstracts were selected and the full texts were analyzed. The relevant information was synthesized, classified, and organized into tables and figures and was presented to a panel of experts, which was composed of 30 professionals from various fields. Based on the Delphi methodology, three rounds of consultation were conducted. The first and second rounds were based on electronic questionnaires that measured the level of consensus of each question among the participants. The third round was based on a face-to-face discussion focusing on those questions without consensus in the previous consultations. The evidence was adapted to national circumstances on a case-by-case basis, and the content the final document was approved. These recommendations are proposed for use in routine medical practice by health professionals in Colombia.

La transmisión congénita de la enfermedad de Chagas ha sido poco estudiada en Colombia y existen pocos procedimientos rutinarios en el sistema de salud para el manejo de esta enfermedad. Por ello se desarrolló un consenso de expertos dirigido a generar recomendaciones de diagnóstico y tratamiento de Chagas con- génito y orientación a mujeres en edad fértil. Con ese propósito se realizó una búsqueda extensiva de la literatura, empleando una combinación de términos Mes (Chagas, Chagas congénito, prevención, control, diagnóstico, tratamiento y embarazo) para reflejar el estado del arte en cada tema de interés. Después de ello, se leyeron los resúmenes y aquellos seleccionados para análisis del texto completo. La literatura relevante se sintetizo, clasifico y organizo en tablas y se presentó al panel de expertos, el cual estaba constituido por 30 profesionales en diferentes áreas. Mediante la metodología Delphi se realizaron 2 rondas de cuestionarios virtuales y una reunión presencial en los cuales se evaluaron los niveles de acuerdo entre los participantes. Los puntos con falta de consenso durante las 2 rondas virtuales se expusieron durante las mesas de discusión en la ronda presencial. La evidencia utilizada se adaptó a las particularidades nacionales según el caso y se aprobó el contenido del documento final. Se propone que estas recomendaciones sean usadas por profesionales de la salud en Colombia.

Analytical Performance of Four Polymerase Chain Reaction (PCR) and Real Time PCR (qPCR) Assays for the Detection of Six Leishmania Species DNA in Colombia.

Leishmaniasis comprises a spectrum of parasitic diseases caused by protozoans of the genus Leishmania. Molecular tools have been widely employed for the detection of Leishmania due to its high sensitivity and specificity. However, the analytical performance of molecular platforms as PCR and real time PCR (qPCR) including a wide variety of molecular markers has never been evaluated. Herein, the aim was to evaluate the analytical performance of 4 PCR-based assays (designed on four different targets) and applied on conventional and real-time PCR platforms. We evaluated the analytical performance of conventional PCR and real time PCR, determining exclusivity and inclusivity, Anticipated Reportable Range (ARR), limit of detection (LoD) and accuracy using primers directed to kDNA, HSP70, 18S and ITS-1 targets. We observed that the kDNA was the most sensitive but does not meet the criterion of exclusivity. The HSP70 presented a higher LoD in conventional PCR and qPCR in comparison with the other markers (1 × 101 and 1 × 10-1 equivalent parasites/mL respectively) and had a higher coefficient of variation in qPCR. No statistically significant differences were found between the days of the test with the four molecular markers. The present study revealed that the 18S marker presented the best performance in terms of analytical sensitivity and specificity for the qPCR in the species tested (species circulating in Colombia). Therefore, we recommend to explore the analytical and diagnostic performance in future studies using a broader number of species across America.

New Scenarios of Chagas Disease Transmission in Northern Colombia.

Chagas disease (CD) is a systemic parasitic infection caused by the flagellated form of Trypanosoma cruzi. Córdoba department, located in the Colombian Caribbean Coast, was not considered as a region at risk of T. cruzi transmission. In this article, we describe the first acute CD case in Salitral village in Sahagún, Córdoba, confirmed by microscopy and serological tests. Our results draw attention to a new scenario of transmission of acute CD in nonendemic areas of Colombia and highlight the need to include CD in the differential diagnosis of febrile syndromes in this region.

Entomological characterization of malaria in northern Colombia through vector and parasite species identification, and analyses of spatial distribution and infection rates.

BACKGROUND: Malaria remains a worldwide public health concern and, in Colombia, despite the efforts to stop malaria transmission, the incidence of cases has increased over the last few years. In this context, it is necessary to evaluate vector diversity, infection rates, and spatial distribution, to better understand disease transmission dynamics. This information may contribute to the planning and development of vector control strategies. RESULTS: A total of 778 Anopheles mosquitoes were collected in fifteen localities of Córdoba from August 2015 to October 2016. Six species were identified and overall, Anopheles albimanus was the most widespread and abundant species (83%). Other species of the Nyssorhynchus subgenus were collected, including Anopheles triannulatus (13%), Anopheles nuneztovari (1%), Anopheles argyritarsis (< 1%) and two species belonging to the Anopheles subgenus: Anopheles pseudopunctipennis (3%) and Anopheles neomaculipalpus (< 1%). Four species were found naturally infected with two Plasmodium species: Anopheles nuneztovari was detected naturally infected with Plasmodium falciparum and Anopheles pseudopunctipennis with Plasmodium vivax, whereas An. albimanus and An. triannulatus were found infected with both parasite species and confirmed by nested PCR. CONCLUSIONS: In general, the obtained results were contrasting with previous studies in terms of the most abundant and widespread collected species, and regarding infection rates, which were higher than those previously reported. A positive relationship between mosquito local abundance at the locality level and human infection at the municipality level was found. Mosquito local abundance and the number of houses with mosquitoes in each village are factors explaining malaria human cases in these villages. The obtained results suggest that other factors related to the apparent variation in malaria eco-epidemiology in northern Colombia, must be identified, to provide health authorities with better decision tools aiming to design control and prevention strategies.

Purification of Trypanosoma cruzi metacyclic trypomastigotes by ion exchange chromatography in sepharose-DEAE, a novel methodology for host-pathogen interaction studies.

Metacyclic trypomastigotes are essential for the understanding of the biology of Trypanosoma cruzi, the agent of Chagas disease. However, obtaining these biological stages in axenic medium is difficult. Techniques based on charge and density of the parasite during different stages have been implemented, without showing a high efficiency in the purification of metacyclic trypomastigotes. So far, there is no protocol implemented where sepharose-DEAE is used as a resin. Therefore, herein we tested its ability to purify metacyclic trypomastigotes in Liver Infusion Triptose (LIT) medium cultures. A simple, easy-to-execute and effective protocol based on ion exchange chromatography on Sepharose-DEAE resin for the purification of T. cruzi trypomastigotes is described. T. cruzi strains from the Discrete Typing Units (DTUs) I and II were used. The strains were harvested in LIT medium at a concentration of 1×107epimastigotes/mL. We calculated the time of trypomastigotes increment (TTI). Based on the data obtained, Ion exchange chromatography was performed with DEAE-sepharose resin. To verify the purity and viability of the trypomastigotes, a culture was carried out in LIT medium with subsequent verification with giemsa staining. To evaluate if the technique affected the infectivity of trypomastigotes, in vitro assays were performed in Vero cells and in vivo in ICR-CD1 mice. The technique allowed the purification of metacyclic trypomastigotes of other stages of T. cruzi in a percentage of 100%, a greater recovery was observed in cultures of 12days. There were differences regarding the recovery of metacyclic trypomastigotes for both DTUs, being DTU TcI the one that recovered a greater amount of these forms. The technique did not affect parasite infectivity in vitro or/and in vivo.

Seasonality, richness and prevalence of intestinal parasites of three neotropical primates (Alouatta seniculus, Ateles hybridus and Cebus versicolor) in a fragmented forest in Colombia.

Studies on parasites infecting non-human primates are essential to better understand the potential threat to humans of zoonoses transmission, particularly under the current processes of pervasive land use change and biodiversity loss. The natural ecosystems in the Middle Magdalena river basin in Colombia have suffered a dramatic reduction and transformation into pastures and agroindustrial monocultures, threatening their biodiversity, and probably affecting the dynamics between parasites and their hosts, as well as altering the disease transmission cycles between wild populations and humans. This study evaluated seasonality, prevalence and richness of intestinal parasites in three species of neotropical primates: Cebus versicolor, Ateles hybridus and Alouatta seniculus, in a fragmented forest in the Middle Magdalena river valley, Colombia. One hundred and eighty five faecal samples were collected between 2010 and 2015. Direct faecal smears were performed with saline solution (0.85%) and iodine solution (1%), in order to identify larvae and eggs based on their morphology. A large proportion of the samples examined (72.9%) was positive for intestinal parasites; seven families of nematodes were identified: Trichuridae, Trichostrongylidae, Oxyuridae, Strongyloididae, Ancylostomatidae, Ascarididae and Gnathostomatidae, two of protozoans: Entamoebidae and Balantiididae, as well as some eggs of trematodes, cestodes and acanthocephalans. Additionally, DNA extraction and sequencing were conducted on 30 faecal samples in order to identify Giardia sp. and Blastocystis hominis, two parasite species also present in humans. Molecular testing for Giardia sp. was negative and Blastocystis hominis was identified in a single sample of Alouatta seniculus. No clear patterns were observed for prevalence of intestinal parasites according to the season; nonetheless, parasite species richness was higher during the dry season. This study builds on our current understanding of intestinal parasites infecting wild neotropical primates and provides novel information on the patterns of intestinal parasites in primate communities exposed to anthropogenic disturbance.