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Background: Plasmodium vivax is the main causative agent of malaria in Panama. However, the prevalence of asymptomatic infections in the different endemic regions remains unknown. Understanding the epidemiological behavior of asymptomatic infections is essential for the elimination of malaria. This study aimed to determine the prevalence of asymptomatic malarial infections in one of the main endemic regions of Panama using multiplex real-time reverse transcription RT-MqPCR., Methods: A cross-sectional study was conducted in three communities in the Guna Yala Comarca. A total of 551 thick blood smears and their respective samples on filter paper were collected from volunteers of different ages and sexes from June 20 to 25, 2016. Infections by the Plasmodium spp. were diagnosed using microscopy and RT-MqPCR. All statistical analyses were performed using the R software., Results: The average prevalence of asymptomatic infections by P. vivax in the three communities detected by RT-MqPCR was 9.3%, with Ukupa having the highest prevalence (13.4%), followed by Aidirgandi (11.1%) and Irgandi (3.3%). A total of 74 samples were diagnosed as asymptomatic infections using RT-MqPCR. Light microscopy (LM) detected that 17.6% (13/74) of the asymptomatic samples and 82.4% (61/74) were diagnosed as false negatives. A 100% correlation was observed between samples diagnosed using LM and RT-MqPCR. A total of 52.7% (39/74) of the asymptomatic patients were female and 85.1% (63/74) were registered between the ages of 1 and 21 years. Factors associated with asymptomatic infection were community (aOR = 0.38 (95% CI 0.17-0.83), p < 0.001) and age aOR = 0.98 (95% CI 0.97-1.00), p < 0.05); F = 5.38; p < 0.05)., Conclusions: This study provides novel evidence of the considerable prevalence of asymptomatic P. vivax infections in the endemic region of Kuna Yala, representing a new challenge that requires immediate attention from the National Malaria Program. The results of this study provide essential information for the health authorities responsible for developing new policies. Furthermore, it will allow program administrators to reorient and design effective malaria control strategies that consider asymptomatic infections as a fundamental part of malaria control and move towards fulfilling their commitment to eliminate it., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 Cáceres Carrera et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Leishmania ( Viannia ) spp. can harbor a double-stranded RNA virus known as Leishmania RNA virus 1 (LRV-1), whose presence has been reported in nine countries across the Americas and seven Leishmania species. Here, we studied 100 Leishmania ( Viannia ) isolates from patients with cutaneous leishmaniasis collected from different endemic areas in Panama from 2016 to 2022. We identified L. ( V. ) panamensis , L. ( V. ) guyanensis , L. ( V. ) braziliensis/guyanensis hybrid, and L. ( V. ) panamensis sp.1. (genetic variant). LRV-1 was detected by RT-PCR in 9% of L. (Viannia) isolates (eight cases in L. ( V. ) panamensis , and one in L. ( V. ) guyanensis) . Phylogenetic analysis based on sequencing data classified all LRV-1 isolates within genotype A, suggesting that LRV phylogenetic proximity is closely aligned with geographical distribution or to the phylogenetic proximity of the Leishmania host in the case of the L. ( V. ) panamensis and L. ( V. ) guyanensis in Panama.

The impacts of climate change on vector-borne diseases are uneven across human populations. This pattern reflects the effect of changing environments on the biology of transmission, which is also modulated by social and other inequities. These disparities are also linked to research outcomes that could be translated into tools for transmission reduction, but are not necessarily actionable in the communities where transmission occurs. The transmission of vector-borne diseases could be averted by developing research that is both hypothesis-driven and community-serving for populations affected by climate change, where local communities interact as equal partners with scientists, developing and implementing research projects with the aim of improving community health. In this Personal View, we share five principles that have guided our research practice to serve the needs of communities affected by vector-borne diseases., Competing Interests: Declaration of interests Work by LFC was conducted while he was hosted by the National Institute of Allergy and Infectious Diseases (NIAID) as a member of the inaugural class of the National Institutes of Health (NIH) Climate and Health Scholars program, which also supported open access fees for this Personal View. LFC and JEC received additional support from National Science Foundation CNH2-1924200. MDF was supported by the NASA New Investigator Program in Earth Science grant 80NSSC21K1736. MP was supported by NIH grant R01 AI153444-01. JEC is a member of the Sistema Nacional de Investigación, SENACYT–Panamá. SL was supported by NIH NIAID grants R01 AI165481 and R01 AI170506. LRB was supported by the Canada Research Chairs Program. Research presented in figure 2 was funded by Ministerio de Economía y Finanzas de la República de Panamá, grant number 9044-024., (Copyright © 2024 The Author(s). Published by Elsevier Ltd. This is an Open Access article under the CC BY 4.0 license. Published by Elsevier Ltd.. All rights reserved.)

Despite ongoing efforts for elimination, malaria continues to be a major public health problem in the Republic of Panama. For effective elimination, it is key that malaria foci and areas of high transmission are identified in a timely manner. Here, we study malaria transmission records for the 2015-2022 period, a time when cases have increased by a factor of ten. Using several methods to study spatial and spatiotemporal malaria confirmed case clusters at the level of localities, including LISA and scan, we found that cases are clustered across indigenous villages located within the autonomous indigenous regions of Ngäbe-Buglé, Guna Yala, and Embera, with the latter on the eastern border of Panama (with Colombia). We discuss the different factors that might be shaping the marked increase in malaria transmission associated with these clusters, which include an inflow of malaria-exposed migrating populations hoping to reach the USA, insufficient health services, and the lack of culturally sensitive actionable tools to reduce malaria exposure among the ethnically diverse and impoverished indigenous populations of Panama.

Leishmaniasis is a disease caused by parasites of the genus Leishmania and transmitted by sand fly vectors. Tegumentary leishmaniasis is the most prevalent clinical outcome in Latin America, afflicting people from 18 countries. In Panama, the annual incidence rate of leishmaniasis is as high as 3000 cases, representing a major public health problem. In endemic regions, L. panamensis is responsible for almost eighty percent of human cases that present different clinical outcomes. These differences in disease outcomes could be the result of the local interplay between L. panamensis variants and human hosts with different genetic backgrounds. The genetic diversity of L. panamensis in Panama has only been partially explored, and the variability reported for this species is based on few studies restricted to small populations and/or with poor resolutive markers at low taxonomic levels. Accordingly, in this study, we explored the genetic diversity of sixty-nine L. panamensis isolates from different endemic regions of Panama, using an MLST approach based on four housekeeping genes (Aconitase, ALAT , GPI and HSP70 ). Two to seven haplotypes per locus were identified, and regional differences in the genetic diversity of L. panamensis were observed. A genotype analysis evidenced the circulation of thirteen L. panamensis genotypes, a fact that might have important implications for the local control of the disease.

A total of 123 DNA samples from Panamanian patients with cutaneous leishmaniasis (CL) lesions were evaluated. These samples were previously confirmed with CL by a specific KDNA-Viannia PCR but had a negative parasitological diagnosis (Group A). Epidemiological variables, such as age, sex, geographic origin, evolution time, and the number and location of the lesions, were analyzed. No significant differences (p < 0.05) were found when these variables were evaluated against a control panel of 123 CL lesion samples from CL patients with positive parasitological diagnoses (Group B). Of the 123 samples (Group A), 67% (82/123) gave positive results when re-analyzed by PCR-hsp70. An analysis of 69 of these samples via PCR-hsp70-RFLP showed that 59.4% (41/69) of the found restriction patterns corresponded to Leishmania (Viannia) panamensis and 40.6% (28/69) to Leishmania (Viannia) guyanensis. Finally, the sequence and phylogenetic analysis of 32 of the samples confirmed the species in 21 (65.6%, 21/32) samples, originally characterized as L. (V.) panamensis. However, 11 samples (34.4%, 11/32), initially identified via RFLP-Hsp70 as L. (V.) guyanensis, matched the sequence of a genetic variant known as Leishmania sp.1. These results point out the species/genetic variants of Leishmania in the case of CL lesions with an apparently low parasite load.

According to the last leishmaniasis report from the Pan American Health Organization (2021) so far Panama is considered free of visceral leishmaniasis (VL). Although the presence of potential vectors and reservoirs involved in the VL transmission cycle have been described in some rural regions of the country, no cases have been reported in humans and domestic or wild animals. Dogs play an important role in the urban transmission of VL; therefore, it is important to detect possible cases of canine visceral leishmaniasis (CVL) in the country. In this sense,this study reports for the first time the Leishmania (Leishmania) infantum infection in imported dogs in Panama. Eleven dogs with clinical suspicion of CVL were evaluated by parasitological (bone marrow aspirate smear), serological (indirect immunofluorescence and/or reference immunochromatographic rapid test) and molecular tests (conventional PCR). The dogs included in this study were analyzed during the period from 2013 to 2020. All dogs presented clinical manifestations compatible with CVL. The samples were initially evaluated by smears and/or rapid serological tests by private practice veterinarians, and later confirmed by serological and/or molecular tests at the national reference laboratory for Leishmania diagnosis. The diagnosis was confirmed in 5/11 dogs by serological, parasitological and/or conventionals PCR targeting kDNA minicircle and Hsp70 gene. Leishmania (L.) infantum species was identified in 3/5 dogs by PCR-RFLP and by sequencing Hsp70-PCR products. This study evidenced the need to increase awareness of private practitioners as well as public health veterinarians on visceral leishmaniasis (VL), and to consider this parasitosis in the differential diagnosis of dogs with clinical and epidemiological characteristics compatible with the disease., (Copyright © 2022 Elsevier B.V. All rights reserved.)

The objective of this study was to provide information on Trypanosoma cruzi genetic diversity among isolates obtained from different biological sources circulating in endemic areas of Panama. Initial discrete typing units (DTUs) assignment was performed evaluating three single locus molecular markers (mini-exon, heat shock protein 60 and glucose-6-phosphate isomerase genes). Further diversity within TcI lineages was explored using a multi-locus sequence typing approach with six maxicircle genes. Haplotype network analysis and evolutionary divergency estimations were conducted to investigate the genetic relatedness between Panamanian TcI isolates and isolates from different endemic regions in the Americas. Our molecular approach validated that TcI is the predominant DTU circulating in Panama across different hosts and vector species, but also confirmed the presence of TcIII and TcVI circulating in the country. The phylogenetic tree topography for most Panamanian TcI isolates displayed a high level of genetic homogeneity between them. The haplotype network analysis inferred a higher genetic diversity within Panamanian TcI isolates, displaying eight different haplotypes circulating in endemic regions of the country, and revealed geographical structuring among TcI from different endemic regions in the Americas. This study adds novelty on the genetic diversity of T. cruzi circulating in Panama and complements regional phylogeographic studies regarding intra-TcI variations.

Didelphis marsupialis has been reported as a competent reservoir for trypanosomatid parasites infections. The aim of this study was to measure Trypanosoma cruzi, T. rangeli, and Leishmania spp. infection rates and to characterize discrete typing units (DTUs) of T. cruzi in D. marsupialis from two Chagas disease endemic sites in Panama. Blood from 57 wild-caught D. marsupialis were examined from two rural communities, Las Pavas (N = 18) and Trinidad de las Minas (N = 39). Twenty-two (38.60%) opossums were positive for flagellates by general hemoculture. T. cruzi infection was confirmed by positive hemoculture and/or kDNA based PCR performed in 31/57 (54.39%) blood samples from opossums. T. rangeli infection was confirmed by hemoculture and/or TrF/R2-Primer PCR assay applied on 12/57 (21.05%) blood samples. Nine (15.79%) D. marsupialis harbored T. cruzi/T. rangeli coinfections. All opossums tested negative for Leishmania spp. by PCR assays based on kDNA and HSP70 gene amplification. There was a significant association between T. cruzi infection and site (Fisher exact test, p = 0.02), with a higher proportion of T. cruzi infected opossums in Las Pavas (77.78%, n = 14/18) compared to Trinidad de las Minas (43.59%, n = 17/39). A significant association was found between habitat type and T. cruzi infection in opossums across both communities, (X 2  = 6.91, p = 0.01, df = 1), with a higher proportion of T. cruzi infection in opossums captured in forest remnants (76%, 19/25) compared to peridomestic areas (37.5%, 12/32). T. rangeli detection, but not T. cruzi detection, may be improved by culture followed by PCR. TcI was the only DTU detected in 22 T. cruzi samples using conventional and real-time PCR. Eight T. rangeli positive samples were characterized as KP1(-)/lineage C. Trypanosome infection data from this common synanthropic mammal provides important information for improved surveillance and management of Chagas disease in endemic regions of Panama., Competing Interests: The authors declare that there are no conflicts of interest., (© 2021 Published by Elsevier Ltd on behalf of Australian Society for Parasitology.)

Panama and all nations within the Mesoamerican region have committed to eliminate malaria within this decade. With more than 90% of the malaria cases in this region caused by Plasmodium vivax , an efficient national/regional elimination plan must include a comprehensive study of this parasite's genetic diversity. Here, we retrospectively analyzed P. vivax genetic diversity in autochthonous and imported field isolates collected in different endemic regions in Panama from 2007 to 2020, using highly polymorphic markers ( csp , msp -1, and msp -3α). We did the analysis using molecular techniques that are cost-effective for malaria molecular surveillance within Mesoamerica. Thus, we used molecular analyses that are feasible for malaria molecular surveillance within the region, and that can provide useful information for policy and decision making about malaria elimination. We also evaluated if haplotypes established by combining the genotypes found in these genes were associated with relevant epidemiological variables and showed structure across the transmission foci that have been observed in Panama. Ten different haplotypes were identified, some of them strongly associated with geographical origin, age, and collection year. Phylogenetic analysis of csp (central repeat domain) revealed that both major variant types (vk210 and vk247) were circulating in Panama. Variant vk247 was restricted to the eastern endemic regions, while vk210 was predominant (77.3%) and widespread, displaying higher diversity (14 alleles) and geographically biased alleles. The regional implications of these molecular findings for the control of P. vivax malaria to achieve elimination across Mesoamerica are discussed.

Background: More than 85% of the malaria cases in Panama occur in poor, rural and indigenous regions like Darien Province. Vector diversity, infection rate and spatial distribution are important entomological parameters of malaria transmission dynamics. Their understanding is crucial for the development of effective disease control strategies. The objective of this study was to determine the composition of Anopheles species, their natural infection rate and their geographic distribution to better understand the malaria transmission dynamics in Darién, Panama., Methods: Anophelines mosquitoes were captured during the rainy and dry season of 2016. We selected five communities where adult anophelines were collected using CDC light-traps, and through protective human-baited traps. Detection of natural infection and Plasmodium genotype were detected via nested PCR through the amplification of ssrRNA and the circumsporozoite protein gene (csp), respectively., Results: A total of 1,063 mosquitoes were collected mosquitoes were collected for the detection of natural infection with Plasmodium spp. Nine Anophelines species were identified, with the predominant species being: An. (Nys.) darlingi (45.0%) and An. (Nys.) albimanus (42.6%). Natural infection in An. (Nys.) albimanus with P. vivax was detected in one mosquito pool from the community Pueblo Tortuga (0.6%), three from Marraganti (1.7%), two from Bajo Chiquito (1.1%) and three pools from Alto Playona 3 (1.7%). For An. (Nys.) darlingi mosquitoes, we detected seven positive pools from the community Bajo Chiquito (4.0%), two pools from Marraganti (1.1%) and two pools from Alto Playona (1.1%). The P. vivax allelic variant VK210 was detected in infected mosquitoes., Conclusion: The results from this study provide new information on the transmission dynamics associated with anophelines vectors in the Darién region. This is the first report of natural P. vivax infection in An. (Nys.) darlingi and its incrimination as a potential malaria vector in this region of Panama. Additional studies are necessary to expand our knowledge and determine crucial parameters in malaria transmission in Darién, which in turn will aid the National Malaria Program in attaining an adequate malaria control strategy towards malaria elimination., Competing Interests: The authors declare that they have no competing interests. LC and JEC are members of the Sistema Nacional de Investigación (SNI), SENACYT, Panama. Any opinions, findings, conclusions or recommendations expressed in this publication are those of the author(s) and do not necessarily reflect the view of the U.S. Department of Agriculture. The mention of firm names or trade products does not imply that they are endorsed or recommended by the U.S. Department of Agriculture over other firms or similar products not mentioned; the USDA is an equal opportunity employer. This does not alter our adherence to PLOS ONE policies on sharing data and materials.

Background: The genetic heterogeneity of Leishmania parasites is a major factor responsible for the wide variety of Leishmania-associated manifestations. Consequently, understanding the genetic make-up of Leishmania species using suitable molecular markers is an important component of realising local and regional scale disease risk. The cytochrome b (cytb) is frequently used to type New World Leishmania species. However, its potential to discriminate Leishmania species and variants requires further evaluation., Objectives: To explore the capacity of cytb gene to identify New World Leishmania species and variants and to develop an approach able to type local Leishmania species and variants., Methods: We retrieved 360 partial and complete Leishmania cytb gene sequences publicly available in GenBank database to study all single nucleotide polymorphisms (SNPs) across the cytb gene that differentiate New World Leishmania species. This information was used to develop an approach based upon the polymorphisms found in a DNA segment of 948bp. We also compared the typing results found with this technique with the polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) profiling obtained using HSP70 gene as target. One hundred Panamanian isolates were used to both typed Leishmania species and assess local genetic variability., Findings: We found complete agreement between our cytb approach and the PCR-RFLP profiling method based on HSP70 for Leishmania species identification. Ninety-two isolates were identified as L. panamensis, although other Viannia species were found circulating at a lower frequency. Three L. panamensis haplotypes were identified in Panamanian provinces. We also provide an initial report of L. guyanensis haplotypes circulating in Panama., Main Conclusions: Cytb gene sequence encompasses key main SNPs that aid to identify Leishmania species. The cytb approach developed with this information was able to identify and assess genetic variability of local Leishmania species found in this study.

Trypanosoma cruzi , a zoonotic kinetoplastid protozoan parasite, is the causative agent of American trypanosomiasis (Chagas disease). Having a very plastic, repetitive and complex genome, the parasite displays a highly diverse repertoire of surface molecules, with pivotal roles in cell invasion, immune evasion and pathogenesis. Before 2016, the complexity of the genomic regions containing these genes impaired the assembly of a genome at chromosomal level, making it impossible to study the structure and function of the several thousand repetitive genes encoding the surface molecules of the parasite. We here describe the genome assembly of the Sylvio X10/1 genome sequence, which since 2016 has been used as a reference genome sequence for T. cruzi clade I (TcI), produced using high coverage PacBio single-molecule sequencing. It was used to analyze deep Illumina sequence data from 34 T. cruzi TcI isolates and clones from different geographic locations, sample sources and clinical outcomes. Resolution of the surface molecule gene distribution showed the unusual duality in the organization of the parasite genome, a synteny of the core genomic region with related protozoa flanked by unique and highly plastic multigene family clusters encoding surface antigens. The presence of abundant interspersed retrotransposons in these multigene family clusters suggests that these elements are involved in a recombination mechanism for the generation of antigenic variation and evasion of the host immune response on these TcI strains. The comparative genomic analysis of the cohort of TcI strains revealed multiple cases of such recombination events involving surface molecule genes and has provided new insights into T. cruzi population structure., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Talavera-López, Messenger, Lewis, Yeo, Reis-Cunha, Matos, Bartholomeu, Calzada, Saldaña, Ramírez, Guhl, Ocaña-Mayorga, Costales, Gorchakov, Jones, Nolan, Teixeira, Carrasco, Bottazzi, Hotez, Murray, Grijalva, Burleigh, Grisard, Miles and Andersson.)

Aims: Leishmaniasis is considered a disease with multiple clinical/immunopathological characteristics, depending on the immunity of the host and the species of the parasite. In Panama, the most prevalent species that causes localized cutaneous leishmaniasis (LCL) is Leishmania (Viannia) panamensis, and its immune response is poorly studied. Therefore, we evaluated by immunohistochemistry, the in situ immune response during this infection., Methods and Results: Biopsies from Panamanian patients with LCL were collected and processed by histological techniques. Infection by L. (V.) panamensis was demonstrated by isolation in culture and molecular characterization by Hsp70-RFLP. The in situ immune response was assessed by immunohistochemistry. The immune response was characterized by predominance of T cells, mainly CD8 cells that showed positive correlation with IFN-γ and Granzyme B. CD4 cells presented positive correlation with both IFN-γ and IL-13, pointed by mixed cellular immune response. Regulatory response was characterized by FoxP3 cells, which showed positive correlation to IL-10 but not with TGF-β., Conclusions: L. (V.) panamensis infection triggers a mixed cellular immune response, characterized by the presence of pro-inflammatory, anti-inflammatory and regulatory elements in the skin lesion of Panamanian patients. These data contribute to a better understanding of the immunopathogenesis of Leishmania Viannia infection in Panama., (© 2020 John Wiley & Sons Ltd.)

Cutaneous Leishmaniasis transmission in the New World is observed in areas with rich sand fly species' faunas. The diversity and composition of sand fly species can change in response to seasonal weather and land use changes. Here, we present results from a two-year-long study where we collected, using Centers for Disease Control (CDC) light traps, sand flies from two rural areas, Las Pavas (LP) and Trinidad de las Minas (T) in western Panamá. Over 710 trap-nights, we collected 16,156 sand flies from 15 genera and 35 species. We identified 34 species in T, and the most abundant species collected was Nyssomyia trapidoi (Fairchild and Hertig, 1952) (n = 2278, 37%), followed by Psychodopygus panamensis (Shannon, 1926) (n = 1112, 18%), and Trichopygomyia triramula (Fairchild and Hertig, 1952) (n = 1063, 17%). In LP, we identified 26 species, and the most abundant species collected were Ty. triramula (n = 4729, 48%), and Ps. panamensis (n = 3444, 35%). We estimated a higher species' richness in T (Chao2 ± S.E.: 36.58 ± 3.84) than in LP (27.49 ± 2.28). In T, species' richness was significantly higher in the rainy season, but no seasonal differences were observed in LP. Species' assemblages were nested in the two areas. Phlebotomine sand fly species' abundance increased at the two sites during the rainy season. Our data suggest that seasonality is more important than land use as a factor driving sand fly species' diversity at the studied sites.

Isolates from 475 cutaneous leishmaniasis (CL) patients from three endemic regions were studied by three typing techniques. The molecular analysis from lesion scrapings based on hsp70 PCR-restriction fragment length polymorphism (RFLP) showed that 78.1% (371/475) restriction patterns corresponded to Leishmania (Viannia) panamensis, 19% (90/475) to Leishmania (Viannia) guyanensis, and 3.0% (14/475) to Leishmania (Viannia) braziliensis. Promastigotes isolated by culture from lesions of 228 patients (48.0%, 228/475) were identified by multi-locus enzyme electrophoresis. Of them, 95.2% (217/228) were typified as L. (V.) panamensis, 1.3% (3/228) as L. (V.) guyanensis, 2.2% (5/228) as L. (V.) braziliensis, and 1.3% (3/228) as hybrids (L. [V.] braziliensis/L. [V.] panamensis). However, a partial sequencing analysis of the hsp70 gene from 77 selected samples showed 16.9% (13/77) typified as L. (V.) panamensis, 68.8% (53/77) as Leishmania (V.) sp., 1, 3.9% (3/77) as L. (V.) guyanensis, 1.3% (1/77) as L. (V.) braziliensis outlier, 2.6% (2/77) as Leishmania (Viannia) naiffi, 2.6% as (2/77) Leishmania (V.) sp., and 2 and 3.9% (3/77) hybrid isolates of L. (V.) braziliensis/L. (V.) guyanensis. These results confirm L. (V.) panamensis as the predominant species and cause of CL lesions in Panama and that L. (V.) guyanensis, L. (V.) braziliensis, and L. (V.) naiffi are circulating to a lower degree. Furthermore, the determination of parasite isolates belonging to atypical clusters and hybrid isolates suggests the circulation of genetic variants with important implications for the epidemiology and clinical follow-up of CL in Panama. No evidence of the existence of parasites of the Leishmania (Leishmania) mexicana complex in Panamanian territory was found in this study.

Background: Increased Attalea butyracea palm propagation, notable for its role as key habitat for the primary Chagas disease vector in Panama, has been linked to landscape disturbance in single-palm observations in this region. Close proximity of these palms to human dwellings is proposed to increase risk of Chagas disease transmission from sylvatic transmission cycles to domestic transmission involving human populations. This study examines the relationship between landscape disturbance and mature A. butyracea spatial distribution, density, and proximity to human populations and vector and reservoir species' movement corridors at a regional scale in a 300 km 2 heterogeneous tropical landscape in central Panama., Methods: We remotely identified the locations of over 50,000 mature A. butyracea palms using high-resolution WorldView2 satellite imagery. A local Getis-Ord Gi* spatial analysis identified significant clusters of aggregated palms. Associations between palm and cluster abundance and a landscape disturbance gradient, derived from official Panama land cover data, were tested using Chi-square tests for Homogeneity and Z-test for proportions. Kruskall-Wallis non-parametric analysis of variance tests were run to assess whether palm cluster area varied by disturbance level, or whether disturbance was associated with proximity of palms and palm clusters to susceptible populations or vector movement corridors., Results: Our findings indicate a regional relationship between landscape disturbance and A. butyracea occurrence. We observe a significant increase in both individual and clustered A. butyracea in secondary forest, but a reduction of palms in agricultural settings. We do not detect evidence of any reduction in abundance of palms in residential settings. The majority of residential and commercial buildings in our study area are within vector flight distance of potential vector habitat in palm crowns., Conclusions: We observe probable anthropogenic elimination of A. butyracea palms in agricultural, but not residential, settings. Even in heavily deforested regions, significant concentrations of mature palms remain in close proximity to human establishments.

Panamá, together with all the nations in Mesoamerica, has committed to eliminate malaria from the region by 2020. As these countries approach malaria elimination and local transmission decreases, an active molecular surveillance to identify genotypes circulating along the border areas is particularly needed to accurately infer infection origin, drug resistance and disease propagation patterns in the region. This study evaluated the genetic diversity and allele frequencies of msp -1, msp -2 and glurp genes using different molecular analyses (nested PCR, PCR-restriction fragment length polymorphism (RFLP) and sequencing) from 106 autochthonous and imported P. falciparum isolates collected from different endemic areas in Panamá between 2003 and 2019. We also explored if P. falciparum genotypes assessed with these molecular markers were associated with relevant malaria epidemiological parameters using a multiple correspondence analysis. A strong association of certain local haplotypes with their geographic distribution in endemic areas, but also with parasite load and presence of gametocytes, was evidenced. Few multiclonal infections and low genetic diversity among locally transmitted P. falciparum samples were detected, consequent with the low transmission intensity of this parasite in Panamá, a pattern likely to be extended across Mesoamerica. In addition, several imported cases were genetically dissimilar to local infections and representative of more diverse extra-continental lineages.

Localized cutaneous leishmaniasis (LCL) caused by Leishmania ( Viannia ) panamensis is an endemic disease in Panama. This condition causes ulcerated skin lesions characterized by a mixed Th1/Th2 immune response that is responsible for disease pathology. However, the maintenance of the in situ inflammatory process involves other elements, such as Th17 and inflammasome responses. Although these processes are associated with parasite elimination, their role in the increase in disease pathology cannot be discarded. Thus, the role in Leishmania infection is still unclear. In this sense, the present study aimed at characterizing the Th17 and inflammasome responses in the skin lesions of patients with LCL caused by L. ( V. ) panamensis to help elucidate the pathogenesis of this disease in Panama. Th17 and inflammasome responses were evaluated by immunohistochemistry (IHQ) in 46 skin biopsies from patients with LCL caused by L. ( V. ) panamensis . The Th17 immune response was assessed using CD3, CD4, RoR γ t, IL-17, IL-6, IL-23, and TGF- β 1 antibodies, and the inflammasome response was assessed by IL-1 β , IL-18, and caspase-1 antibodies. The presence of the Th17 and inflammasome responses was evidenced by a positive reaction for all immunological markers in the skin lesions. An inverse correlation between the density of amastigotes and the density of RoR γ t + , IL-17 + , IL-1 β + , and caspase-1 + cells was observed, but no correlation between Th17 and the inflammasome response with evolutionary disease pathology was reported. These data showed the participation of Th17 cells and the inflammasome in the inflammatory response of the skin lesions of LCL caused by L. ( V. ) panamensis infection. These results suggest a role in the control of tissue parasitism of IL-17 and the activation of the NLRP3 inflammasome dependent on IL-1 β but cannot exclude their role in the development of disease pathology., Competing Interests: The authors declare that they have no conflict of interest., (Copyright © 2020 K. Gonzalez et al.)

Rhodnius pallescens is the principal vector of Chagas disease in Panama. Recently a dark chromatic morph has been discovered in the highlands of Veraguas Province. Limited genetic studies have been conducted with regards to the population structure and dispersal potential of Triatominae vectors, particularly in R. pallescens. Next generation sequencing methods such as RADseq and complete mitochondrial DNA (mtDNA) genome sequencing have great potential for examining vector biology across space and time. Here we utilize a RADseq method (3RAD), along with complete mtDNA sequencing, to examine the population structure of the two chromatic morpho types of R. pallescens in Panama. We sequenced 105 R. pallescens samples from five localities in Panama. We generated a 2216 SNP dataset and 6 complete mtDNA genomes. RADseq showed significant differentiation among the five localities (F CT  = 0.695; P = .004), but most of this was between localities with the dark vs. light chromatic morphs (Veraguas vs. Panama Oeste). The mtDNA genomes showed a 97-98% similarity between dark and light chromatic morphs across all genes and a 502 bp insert in light morphs. Thus, both the RADseq and mtDNA data showed highly differentiated clades with essentially no gene flow between the dark and light chromatic morphs from Veraguas and central Panama respectively. We discuss the growing evidence showing clear distinctions between these two morpho types with the possibility that these are separate species, an area of research that requires further investigation. Finally, we discuss the cost-effectiveness of 3RAD which is a third of the cost compared to other RADseq methods used recently in Chagas disease vector research., Competing Interests: Declaration of Competing Interest Jose E. Calzada and Azael Saldana are members of the SNI (Sistema Nacional de Investigación from SENACYT of Panama). All other authors declare no competing interest., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Background: The present study provides a countrywide perspective of the malaria situation in Panamá over a long-term framework, with the purpose of identifying historical malaria resurgence events and their potential causes., Methods: A descriptive-ecological study was conducted by analysing demographic and epidemiological annual malaria time series data in Panamá (1884-2019) using several data sources. Malaria intensity indicators were calculated during the study period. The effects of El Niño Southern Oscillation on malaria transmission were also analysed using a retrospective analysis of malaria cases between 1957 and 2019., Results: Several factors were identified responsible for malaria resurgence in Panamá, mostly related with Malaria Control Programme weakening. During the past 20 years (2000-2019) malaria has progressively increased in prevalence within indigenous settlements, with a predominance of male cases and a high proportion (15% of total cases) in children less than 5 years old. During this period, a significant and increasing proportion of the Plasmodium falciparum cases were imported. Retrospective analysis (1957-2019) evidenced that ENSO had a significant impact on malaria transmission dynamics in Panamá., Conclusions: Data analysis confirmed that although authorities have been successful in focalizing malaria transmission in the country, there are still neglected issues to be solved and important intercultural barriers that need to be addressed in order to achieve elimination of the disease by 2022. This information will be useful for targeting strategies by the National Malaria Elimination Programme.

Blastocystis sp. is a commonly reported intestinal parasite with a worldwide distribution. Phylogenetic analyses describe at least 17 subtypes for this parasite, and nine of them have been found in humans. However, the prevalence and some epidemiological characteristics of this parasitic infection in rural communities are not well known. The objective of this cross-sectional study was to evaluate the prevalence, subtypes, and epidemiological factors related to Blastocystis sp. Infection in children from of a small rural community in the central area of Panama. For this, 66 fecal samples from children (1 to 12 years old), were initially analyzed for the presence of parasites by a formalin-ethyl acetate/concentration method. Molecular detection and identification of Blastocystis sp. subtypes were carried out by amplification and sequencing of a partial fragment of the small-subunit ribosomal RNA gene. Using data from a questionnaire, analyses of epidemiological conditions potentially associated with Blastocystis sp. transmission were also conducted. Microscopic diagnostics showed that 33.3% (22/66) of the analyzed samples presented entero-parasites. Among them, Blastocystis sp. was the most prevalent, with 21.2% (14/66), followed by the E. histolytica/dispar/moshkovskii complex 4.5% (3/66), Giardia lamblia 1.5% (1/66) and Strongyloides stercoralis 1.5% (1/66). PCR-based analyses detected a prevalence of Blastocystis sp. infection of 74.2% (49/66) in apparently healthy children. Phylogenetic analysis revealed two different subtypes of this parasite: ST1 with 42.2% (28/66) infected, and ST3 with 31.8% (21/66) infected. In addition, recent diarrhea was significantly associated with Blastocystis sp. infection. None of the other risk factors evaluated was statistically associated with infection. These results highlight the need to further investigate clinical, epidemiological, and genetic characteristics of Blastocystis sp. infections in this community., (© 2020 The Authors. Published by Elsevier Ltd on behalf of World Federation of Parasitologists.)

Background: Malaria is endemic in Darién and an assessment of the different factors affecting its epidemiology is crucial for the development of adequate strategies of surveillance, prevention, and disease control. The objective of this study was to determine the main characteristics of the epidemiological behavior of malaria in the Darien region., Methods: This research was comprised of a retrospective analysis to determine the incidence and malaria distribution in the Darien region from 2015 to 2017. We evaluated malaria indicators, disease distribution, incidence (by age group and sex), diagnostic methods, treatment, and control measures. In addition, we examined the cross-border migration activity and its possible contribution to the maintenance and distribution of malaria., Results: During the period of 2015-2017, we examined 41,141 thick blood smear samples, out of which 501 tested positive for malaria. Plasmodium vivax was responsible for 92.2% of those infections. Males comprised 62.7% of the total diagnosed cases. Meanwhile, a similar percentage, 62.7%, of the total cases were registered in economically active ages. The more frequent symptoms included fever (99.4%) and chills (97.4%), with 53.1% of cases registering between 2,000 and 6,000 parasites/μl of blood. The annual parasitic incidence (API) average was 3.0/1,000 inhabitants, while the slide positivity rate (SPR) was 1.2% and the annual blood examination rate (ABER) 22.5%. In Darién there is a constant internal and cross-border migration movement between Panama and Colombia. Malaria control measures consisted of the active and passive search of suspected cases and of the application of vector control measures., Conclusion: This study provides an additional perspective on malaria epidemiology in Darién. Additional efforts are required to intensify malaria surveillance and to achieve an effective control, eventually moving closer to the objective of malaria elimination. At the same time, there is a need for more eco-epidemiological, entomological and migratory studies to determine how these factors contribute to the patterns of maintenance and dissemination of malaria., Competing Interests: The authors have declared that no competing interests exist. LC is member of the Sistema Nacional de Investigación (SNI), SENACYT, Panama. Any opinions, findings, conclusions or recommendations expressed in this publication are those of the author(s) and do not necessarily reflect the view of the U.S. Department of Agriculture. The mention of firm names or trade products does not imply that they are endorsed or recommended by the U.S. Department of Agriculture over other firms or similar products not mentioned; the USDA is an equal opportunity employer.

Background: Triatomine bugs are vectors of the protozoan parasite Trypanosoma cruzi, which causes Chagas disease. Rhodnius pallescens is a major vector of Chagas disease in Panama. Understanding the microbial ecology of disease vectors is important in the development of vector management strategies that target vector survival and fitness. In this study we examined the whole-body microbial composition of R. pallescens from three locations in Panama., Methods: We collected 89 R. pallescens specimens using Noireau traps in Attalea butyracea palms. We then extracted total DNA from whole-bodies of specimens and amplified bacterial microbiota using 16S rRNA metabarcoding PCR. The 16S libraries were sequenced on an Illumina MiSeq and analyzed using QIIME2 software., Results: We found Proteobacteria, Actinobacteria, Bacteroidetes and Firmicutes to be the most abundant bacterial phyla across all samples. Geographical location showed the largest difference in microbial composition with northern Veraguas Province having the most diversity and Panama Oeste Province localities being most similar to each other. Wolbachia was detected in high abundance (48-72%) at Panama Oeste area localities with a complete absence of detection in Veraguas Province. No significant differences in microbial composition were detected between triatomine age class, primary blood meal source, or T. cruzi infection status., Conclusions: We found biogeographical regions differ in microbial composition among R. pallescens populations in Panama. While overall the microbiota has bacterial taxa consistent with previous studies in triatomine microbial ecology, locality differences are an important observation for future studies. Geographical heterogeneity in microbiomes of vectors is an important consideration for future developments that leverage microbiomes for disease control.

Cutaneous leishmaniasis (CL) is one of the most frequent parasitic zoonoses in Panama. Currently, conventional, molecular and histopathological tests are performed to diagnose CL. Immunohistochemistry (IHC) has proven to be a valuable tool to facilitate the diagnosis of leishmaniasis and to study the cellular immune response developed during the infection. Therefore, considering the absence of IHC in the diagnostic routine in Panama, the objective of this study is to demonstrate the usefulness of this test as a complementary diagnostic tool for improving the sensitivity of histopathology (HP) and helping to study the cellular immune response of patients. Samples from patients with suspected CL were analysed by intradermal reaction of Montenegro (IDRM), smears, culture, PCR (Viannia, Hsp-70), HP and IHC. According to the diagnostic criteria, 95.8% of patients were positive for Leishmania sp., that was characterized as Leishmania (V.) panamensis by PCR-HSP70/RFLP. From positive samples, Leishmania was detected by the tested diagnostic methods in the following degrees: 100% by IDRM, 60% by smears, 93.3% by culture, 100% by kDNA PCR, 78.3% by PCR Hsp-70, 50% by HP and 73.9% by IHC. Although IHC had a poor correlation (k = 0.191) with the diagnostic criteria, the sensitivities of both HP (76.1%) and smears (89.1%) were improved by combining them with IHC. IHC considerably improved the detection of the Leishmania parasites in the histopathological sections, supporting the need to implement this diagnostic tool in Panama. In addition, immunohistochemistry allows evaluation of the patient's immune response and thus provides new guidelines for the treatment and control of CL in Panama., (Copyright © 2019 Elsevier B.V. All rights reserved.)

American cutaneous leishmaniasis (ACL) is a common and important vector-borne parasitic zoonosis in Panamá. Here, we study Leishmania spp. infection rates and blood-feeding patterns among common sand flies in Trinidad de Las Minas, a rural community with hyperendemic ACL transmission, and where a deltamethrin fogging trial was performed. Sand flies were collected from April 2010 to June 2011 with light traps installed inside and in the peridomicile of 24 houses. We restricted our analysis to the most abundant species at the study site: Lutzomyia trapidoi , Lutzomyia gomezi , Lutzomyia panamensis , Lutzomyia triramula , and Lutzomyia dysponeta . We detected Leishmania spp. infection in sand flies by polymerase chain reaction (PCR) amplification of the internal transcribed spacer region 1 (ITS-1) in pooled females (1-10 females per pool). Host species of engorged sand flies were identified using a cytochrome b PCR. From 455 sand fly pools analyzed, 255 pools were positive for Leishmania spp., with an estimated infection rate (confidence interval) of 0.096 [0.080-0.115] before the deltamethrin fogging which slightly, but not significantly ( P > 0.05), increased to 0.116 [0.098-0.136] after the deltamethrin fogging. Blood meal analysis suggested that pigs, goats, and birds were the most common sand fly blood sources, followed by humans and domestic dogs. DNA sequencing from a subsample of ITS-1 positive pools suggests that Leishmania panamensis , Leishmania naiffi , and other Leishmania spp. were the parasite species infecting the most common vectors at the study site. Our data confirm an association between sand fly species, humans, domestic dogs, and pigs and Leishmania spp. parasites in rural Panamá.

The Panamá Canal construction encompassed one of the first examples of malaria elimination. Nevertheless, malaria has uninterruptedly persisted in Native American populations living within a few kilometers of the Panamá Canal. Here, we present results from a monthly longitudinal study (May 2016 to March 2018), whose goal was to quantitatively describe seasonal patterns of Plasmodium spp. infection in Anopheles albimanus Wiedemann, and its association with environmental covariates, at Ipetí-Guna, a village within a region targeted for malaria elimination in Panamá. To detect Plasmodium spp. infections we employed a standard nested PCR on DNA extracts from mosquito pools of varying size, which were then used to estimate monthly infection rates using a maximum likelihood method. The infection rate estimates (IR) were analyzed using time series analysis methods to study their association with changes in rainfall, temperature, NDVI (a satellite derived vegetation index), malaria cases and human biting rates (HBR). We found that mosquitoes were infected by Plasmodium vivax mainly from September to December, reaching a peak in December. Time series modeling showed malaria IR in An albimanus increased, simultaneously with HBR, and IR in the previous month. These results suggest that elimination interventions, such as mass drug administration, are likely to be more effective if deployed from the middle to the end of the dry season (March and April at Ipetí-Guna), when the likelihood of malaria infection in mosquitoes is very low and when curtailing human infections driving infections in mosquitoes can reduce malaria transmission, and increase the chance for elimination., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Anopheles albimanus Wiedemann is a major malaria vector in Mesoamerica and the Caribbean whose population dynamics, in response to changing environments, has been relatively poorly studied. Here, we present monthly adult and larvae data collected from May 2016 to December 2017 in Ipetí-Guna, a village within an area targeted for malaria elimination in the República de Panamá. During the study period we collected a total of 1678 Anopheles spp. mosquitoes (1602 adults and 76 larvae). Over 95% of the collected Anopheles spp. mosquitoes were An. albimanus . Using time series analysis techniques, we found that population dynamics of larvae and adults were not significantly correlated with each other at any time lag, though correlations were highest at one month lag between larvae and adults and four months lag between adults and larvae. Larvae population dynamics had cycles of three months and were sensitive to changes in temperature with 5 months lag, while adult abundance was correlated with itself (1 month lag) and with the normalized difference vegetation index (NDVI) with three months lag. A key observation from our study is the absence of both larvae and adults of An. albimanus between January and April from environments associated with Guna population's daily activities, which suggests this time window could be the best time to implement elimination campaigns aimed at clearing Plasmodium spp. parasites from Guna populations using, for example, mass drug administration.

Background: Rhodnius pallescens, the only species of this genus reported in Panama, has a wide geographical distribution and is associated with most cases of Chagas disease and human infections with Trypanosoma rangeli in this country. Thus far, no phenotypic variants of this triatomine have been registered. Similarly, genotyping of the trypanosomes that infect this vector has only been partially evaluated., Results: A total of 347 specimens of R. pallescens were collected in Attalea butyracea palm trees located near a mountainous community of the district of Santa Fe, province of Veraguas. Bugs were slightly longer and had a darker coloration compared to that reported for this species. Infection rates for trypanosomes performed with three PCR analyses showed that 41.3% of the adult triatomines were positive for T. cruzi, 52.4% were positive for T. rangeli and 28.6% had mixed T. cruzi/T. rangeli infections. Based on cox2 analysis, TcI was the single T. cruzi discrete typing unit (DTU) detected, and a genetic variant of KP1(-)/lineage C was the only genetic group found for T. rangeli., Conclusions: A darker chromatic variation of R. pallescens predominates in a mountainous region of Panama. These triatomines show high trypanosome infection rates, especially with T. rangeli. Regarding T. rangeli genetic diversity, complementary studies using other molecular markers are necessary to better define its phylogenetic position.

Ecoepidemiological scenarios for Trypanosoma cruzi Chagas transmission are partially shaped by kissing bug vector ecology. The presence of Attalea butyracea Kunth, the 'royal palm', is a major risk factor for Chagas disease transmission in Panamá given their frequent infestations by Rhodnius pallescens Barber, a major neotropical T. cruzi vector. It was assumed that in Panamá this relationship was very close and unique, limiting the niche of R. pallescens to that of Att. butyracea. However, here we present observations about T. cruzi-infected R. pallescens infesting coyol palms, Acrocomia aculeata Jacquin, in Pedasí district, Los Santos Province, Panamá. Between May 2015 and August 2016, we sampled kissing bugs from 83 coyol palms using mice-baited traps placed at the crown of each palm during the dry and wet season. We collected 62 R. pallescens and one Eratyrus cuspidatus Stål kissing bugs. Using logistic regression, we found that the probability of kissing bug infestation in coyol palms increased during the rainy season, with infructescence number and palm height. We examined adult R. pallescens bugs (n = 30) and found T. cruzi in 67% of the samples. We were able to isolate and characterize T. cruzi from parasites present in the feces from R. pallescens, all parasites belonging to the TC I lineage. We found that green fronds number and house proximity increased T. cruzi infection probability in kissing bugs collected in coyol palms. These results highlight coyol palms as a potential risk factor for Chagas disease transmission in the dry arch of Panamá.

Background: Malaria has historically been entrenched in indigenous populations of the República de Panamá. This scenario occurs despite the fact that successful methods for malaria elimination were developed during the creation of the Panamá Canal. Today, most malaria cases in the República de Panamá affect the Gunas, an indigenous group, which mainly live in autonomous regions of eastern Panamá. Over recent decades several malaria outbreaks have affected the Gunas, and one hypothesis is that such outbreaks could have been exacerbated by climate change, especially by anomalous weather patterns driven by the EL Niño Southern Oscillation (ENSO)., Results: Monthly malaria cases in Guna Yala (1998-2016) were autocorrelated up to 2 months of lag, likely reflecting parasite transmission cycles between humans and mosquitoes, and cyclically for periods of 4 months that might reflect relapses of Plasmodium vivax, the dominant malaria parasite transmitted in Panamá. Moreover, malaria case number was positively associated (P < 0.05) with rainfall (7 months of lag), and negatively with the El Niño 4 index (15 months of lag) and the Normalized Difference Vegetation Index, NDVI (8 months of lag), the sign and magnitude of these associations likely related to the impacts of weather patterns and vegetation on the ecology of Anopheles albimanus, the main malaria vector in Guna Yala. Interannual cycles, of approximately 4-year periods, in monthly malaria case numbers were associated with the El Niño 4 index, a climatic index associated with weather and vegetation dynamics in Guna Yala at seasonal and interannual time scales., Conclusion: The results showed that ENSO, rainfall and NDVI were associated with the number of malaria cases in Guna Yala during the study period. These results highlight the vulnerability of Guna populations to malaria, an infection sensitive to climate change, and call for further studies about weather impacts on malaria vector ecology, as well as the association of malaria vectors with Gunas paying attention to their socio-economic conditions of poverty and cultural differences as an ethnic minority.

We examined the mitogenomes of a large global collection of human malaria parasites to explore how and when Plasmodium falciparum and P. vivax entered the Americas. We found evidence of a significant contribution of African and South Asian lineages to present-day New World malaria parasites with additional P. vivax lineages appearing to originate from Melanesia that were putatively carried by the Australasian peoples who contributed genes to Native Americans. Importantly, mitochondrial lineages of the P. vivax-like species P. simium are shared by platyrrhine monkeys and humans in the Atlantic Forest ecosystem, but not across the Amazon, which most likely resulted from one or a few recent human-to-monkey transfers. While enslaved Africans were likely the main carriers of P. falciparum mitochondrial lineages into the Americas after the conquest, additional parasites carried by Australasian peoples in pre-Columbian times may have contributed to the extensive diversity of extant local populations of P. vivax.

Cutaneous leishmaniasis (CL) is an endemic disease in the Republic of Panama, caused by Leishmania (Viannia) parasites, whose most common clinical manifestation is the presence of ulcerated lesions on the skin. These lesions usually present a chronic inflammatory reaction, sometimes granulomatous, with the presence of lymphocytes, plasma cells and macrophages. This study describes the histopathological characteristics found in the skin lesions of patients with CL caused by Leishmania (V.) panamensis in Panama. We analyzed 49 skin biopsy samples from patients with clinical suspicion of CL, by molecular tests (PCR for subgenus Viannia and HSP-70) and by Hematoxylin-Eosin staining. Samples were characterized at the species level by PCR-HSP-70/RFLP. From the 49 samples studied, 46 (94%) were positive by PCR and were characterized as Leishmania (V.) panamensis. Of these, 48% were positive by Hematoxylin-Eosin staining with alterations being observed both, in the epidermis (85%) and in the dermis (100%) of skin biopsies. The inflammatory infiltrate was characterized according to histopathological patterns: lymphohistiocytic (50%), lymphoplasmacytic (61%) and granulomatous (46%) infiltration, being the combination of these patterns frequently found. The predominant histopathological characteristics observed in CL lesions caused by L. (V.) panamensis in Panama were: an intense inflammatory reaction in the dermis with a combination of lymphohistiocytic, lymphoplasmacytic and granulomatous presentation patterns and the presence of ulcers, acanthosis, exocytosis and spongiosis in the epidermis.

Species composition of wild reservoir hosts can influence the transmission and maintenance of multi-host vector borne pathogens. The 'pace of life' hypothesis proposes that the life history strategy of reservoir hosts can influence pathogen transmission of vector borne generalist pathogens. We use empirical data to parameterize a mathematical model that investigates the impacts of host life history traits on vector transmission dynamics of the vector-borne multi-host parasite Trypanosoma cruzi in habitats characterized by different degrees of deforestation and varying host community structure. The model considers susceptible and infected vector and host populations. When comparing the proportion of vectors infected with T. cruzi predicted by the model with empirical data, we found a trend of increasing vector infection as anthropogenic landscape disturbance increases for both data and model output. The model's vector infection rates were significantly lower than empirical results, but when incorporating host congenital transmission in the model, vector infection approaches field data. We conclude that intervened habitats associated with r-selected host species communities predict higher proportions of infected vectors.

Accurate blood meal identification is critical to understand hematophagous vector-host relationships. This study describes a customizable Next-Generation Sequencing (NGS) approach to identify blood meals from Rhodnius pallescens (Hemiptera: Reduviidae) triatomines using multiple barcoded primers and existing software to pick operational taxonomic units and match sequences for blood meal identification. We precisely identified all positive control samples using this method and further examined 74 wild-caught R. pallescens samples. With this novel blood meal identification method, we detected 13 vertebrate species in the blood meals, as well as single and multiple blood meals in individual bugs. Our results demonstrate the reliability and descriptive uses of our method., (© The Authors 2017. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Background: The purpose of the study was to recognize the social representation of the Guna indigenous population by identifying cultural elements related to malaria, in order to create an intercultural approach to any health intervention to control and prevent the disease., Methods: This qualitative study has an anthropological focus that used participant observation, informal conversations, focus groups, interviews, in-depth interviews with key actors. Analyses included review, classification and categorization of interviews., Results: Malaria within the Guna culture is in harmony with several cultural factors, such as magic, religion, beliefs, myths, and nature. The health system must include these factors in its intercultural approach to ensure the sustainability of anti-malarial intervention measures. Until this is not properly addressed, the Guna population of Madungandi will remain a permanent source of risk for malaria transmission in this region and for the rest of the country., Conclusion: The findings of this study contribute new variables that can facilitate an intercultural approach to improve the perception of malaria in the indigenous population of Comarca Guna de Madungandi, Panama.

Synanthropic wild mammals can be important hosts for many vector-borne zoonotic pathogens. The aim of this study was determine the exposure of synanthropic mammals to two types of tick-borne pathogens in Panama, spotted fever group Rickettsia (SFGR) and Borrelia relapsing fever (RF) spirochetes. One hundred and thirty-one wild mammals were evaluated, including two gray foxes, two crab-eating foxes (from zoos), four coyotes, 62 opossum and 63 spiny rats captured close to rural towns. To evaluate exposure to SFGR, serum samples from the animals were tested by indirect immunofluorescence assay (IFA) using Rickettsia rickettsii and Candidatus Rickettsia amblyommii antigen. Immunoblotting was performed using Borrelia turicatae protein lysates and rGlpQ, to assess infection caused by RF spirochetes. One coyote (25%) and 27 (43%) opossums showed seroreactivity to SFGR. Of these opossums, 11 were seroreactive to C. R. amblyommii. Serological reactivity was not detected to B. turicatae in mammal samples. These findings may reflect a potential role of both mammals in the ecology of tick-borne pathogens in Panama., Competing Interests: The authors have declared that no competing interests exist.

A precise identification of Leishmania species involved in human infections has epidemiological and clinical importance. Herein, we describe a preliminary validation of a restriction fragment length polymorphism assay, based on the calmodulin intergenic spacer region, as a tool for detecting and typing Leishmania species. After calmodulin amplification, the enzyme HaeIII yielded a clear distinction between reference strains of Leishmania mexicana, Leishmania amazonensis, Leishmania infantum, Leishmania lainsoni, and the rest of the Viannia reference species analyzed. The closely related Viannia species: Leishmania braziliensis, Leishmania panamensis, and Leishmania guyanensis, are separated in a subsequent digestion step with different restriction enzymes. We have developed a more accessible molecular protocol for Leishmania identification/typing based on the exploitation of part of the calmodulin gene. This methodology has the potential to become an additional tool for Leishmania species characterization and taxonomy., (© The American Society of Tropical Medicine and Hygiene.)

American Cutaneous Leishmaniasis (ACL) is a neglected vector-borne zoonosis that persists despite increasing socio-economic development and urbanization in Panamá. Here, we investigate the association between environmental changes and spatio-temporal ACL transmission in the Republic of Panamá (1980-2012). We employ a macroecological approach, where patterns of variation in ACL incidence at the spatially coarse-grained scale of health areas are studied considering factors linked to the ecology of ACL transmission. We specifically study impacts of climatic variability, measured by the different phases of El Niño Southern Oscillation (ENSO), within diverse ecosystems and sand fly (Diptera: Psychodidae) vector species, as well as heterogeneous local climatic patterns, deforestation, population growth rates, and changes in social marginalization. We found that over the study period, patterns of ACL incidence: (i) were asynchronous with clusters changing from east to west of the Panamá Canal, (ii) trends increased in the west, and decreased or remained nearly constant in the east, independent of human population growth, (iii) generally increased in years following El Niño, and (iv) decreased as forest cover increased. We found no significant association between changes in socio-economic indicators and ACL transmission. Regarding vector abundance and presence, we found that studies had been biased to locations east of the Panamá canal, and that, in general, the abundance of dominant vector species decreased during the cold phase of ENSO. Finally, our results indicate that a macroecological approach is useful to understand heterogeneities related to environmental change impacts on ACL transmission.

Background: This study aims to describe the epidemiological and entomological factors associated with a recent malaria outbreak that occurred in 2012 in a socially marginalized population from Guna Yala Comarca in Panama., Methods: A descriptive and observational study was conducted by analysing demographic and epidemiological data from all malaria cases registered during 2012 in the Comarca Guna Yala, Panama. Malaria intensity indicators were calculated during the study period. Entomological evaluations were performed monthly, from October to December 2012, in the three communities that presented the most intense malaria transmission during the first semester of 2012. Anopheles breeding habitats were also characterized., Results: During the studied period, 6754 blood smears were examined (17.8 % of the total population), and 143 were confirmed as positive for Plasmodium vivax. A significant increase of malaria transmission risk indicators (API: 3.8/1000, SPR: 2.1 %) was observed in Guna Yula, when compared with previous years, and also in comparison with estimates from the whole country. Anopheles albimanus was the most abundant and widespread (877; 72.0 %) vector species found in the three localities, followed by Anopheles punctimacula (231; 19.0 %) and Anopheles aquasalis (110; 9.0 %). Three An. albimanus pools were positive for P. vivax, showing an overall pooled prevalence estimate of 0.014., Conclusions: Data analysis confirmed that during 2012 a malaria epidemic occurred in Guna Yala. Panama. This study provides baseline data on the local epidemiology of malaria in this vulnerable region of Panamá. This information will be useful for targeting control strategies by the National Malaria Control Programme.

Chagas disease, caused by Trypanosoma cruzi infection, is a zoonosis of humans, wild and domestic mammals, including dogs. In Panama, the main T. cruzi vector is Rhodnius pallescens, a triatomine bug whose main natural habitat is the royal palm, Attalea butyracea. In this paper, we present results from three T. cruzi serological tests (immunochromatographic dipstick, indirect immunofluorescence and ELISA) performed in 51 dogs from 24 houses in Trinidad de Las Minas, western Panama. We found that nine dogs were seropositive (17.6% prevalence). Dogs were 1.6 times more likely to become T. cruzi seropositive with each year of age and 11.6 times if royal palms where present in the peridomiciliary area of the dog's household or its two nearest neighbours. Mouse-baited-adhesive traps were employed to evaluate 12 peridomestic royal palms. All palms were found infested with R. pallescens with an average of 25.50 triatomines captured per palm. Of 35 adult bugs analysed, 88.6% showed protozoa flagellates in their intestinal contents. In addition, dogs were five times more likely to be infected by the presence of an additional domestic animal species in the dog's peridomiciliary environment. Our results suggest that interventions focused on royal palms might reduce the exposure to T. cruzi infection.

American cutaneous leishmaniasis (ACL) is a complex disease with a rich diversity of animal host species. This diversity imposes a challenge, since understanding ACL transmission requires the adequate identification of reservoir hosts, those species able to be a source of additional infections. In this study we present results from an ACL cross-sectional serological survey of 51 dogs (Canis familiaris), where we used diagnostic tests that measure dog's exposure to Leishmania spp. parasites. We did our research in Panamá, at a village that has undergone significant ecosystem level transformations. We found an ACL seroprevalence of 47% among dogs, and their exposure was positively associated with dog age and abundance of sand fly vectors in the houses of dog owners. Using mathematical models, which were fitted to data on the proportion of positive tests as function of dog age, we estimated a basic reproductive number (R 0 ± s.e.) of 1·22 ± 0·09 that indicates the disease is endemically established in the dogs. Nevertheless, this information by itself is insufficient to incriminate dogs as ACL reservoirs, given the inability to find parasites (or their DNA) in seropositive dogs and previously reported failures to experimentally infect vectors feeding on dogs with ACL parasites.

Identifying the source of resurgent parasites is paramount to a strategic, successful intervention for malaria elimination. Although the malaria incidence in Panama is low, a recent outbreak resulted in a 6-fold increase in reported cases. We hypothesized that parasites sampled from this epidemic might be related and exhibit a clonal population structure. We tested the genetic relatedness of parasites, using informative single-nucleotide polymorphisms and drug resistance loci. We found that parasites were clustered into 3 clonal subpopulations and were related to parasites from Colombia. Two clusters of Panamanian parasites shared identical drug resistance haplotypes, and all clusters shared a chloroquine-resistance genotype matching the pfcrt haplotype of Colombian origin. Our findings suggest these resurgent parasite populations are highly clonal and that the high clonality likely resulted from epidemic expansion of imported or vestigial cases. Malaria outbreak investigations that use genetic tools can illuminate potential sources of epidemic malaria and guide strategies to prevent further resurgence in areas where malaria has been eliminated., (© The Author 2014. Published by Oxford University Press on behalf of the Infectious Diseases Society of America. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.)

The eco-epidemiology of American cutaneous leishmaniasis (ACL) is driven by animal reservoir species that are a source of infection for sand flies that serve as vectors infecting humans with Leishmania spp parasites. The emergence and re-emergence of this disease across Latin America calls for further studies to identify reservoir species associated with enzootic transmission. Here, we present results from a survey of 52 individuals from 13 wild mammal species at endemic sites in Costa Rica and Panama where ACL mammal hosts have not been previously studied. For Leishmania spp. diagnostics we employed a novel PCR technique using blood samples collected on filter paper. We only found Leishmania spp parasites in one host, the two-toed sloth, Choloepus hoffmanni. Our findings add further support to the role of two-toed sloths as an important ACL reservoir in Central America.

Background: Cutaneous Leishmaniasis (CL) is a neglected tropical vector-borne disease. Sand fly vectors (SF) and Leishmania spp parasites are sensitive to changes in weather conditions, rendering disease transmission susceptible to changes in local and global scale climatic patterns. Nevertheless, it is unclear how SF abundance is impacted by El Niño Southern Oscillation (ENSO) and how these changes might relate to changes in CL transmission., Methodology and Findings: We studied association patterns between monthly time series, from January 2000 to December 2010, of: CL cases, rainfall and temperature from Panamá, and an ENSO index. We employed autoregressive models and cross wavelet coherence, to quantify the seasonal and interannual impact of local climate and ENSO on CL dynamics. We employed Poisson Rate Generalized Linear Mixed Models to study SF abundance patterns across ENSO phases, seasons and eco-epidemiological settings, employing records from 640 night-trap sampling collections spanning 2000-2011. We found that ENSO, rainfall and temperature were associated with CL cycles at interannual scales, while seasonal patterns were mainly associated with rainfall and temperature. Sand fly (SF) vector abundance, on average, decreased during the hot and cold ENSO phases, when compared with the normal ENSO phase, yet variability in vector abundance was largest during the cold ENSO phase. Our results showed a three month lagged association between SF vector abundance and CL cases., Conclusion: Association patterns of CL with ENSO and local climatic factors in Panamá indicate that interannual CL cycles might be driven by ENSO, while the CL seasonality was mainly associated with temperature and rainfall variability. CL cases and SF abundance were associated in a fashion suggesting that sudden extraordinary changes in vector abundance might increase the potential for CL epidemic outbreaks, given that CL epidemics occur during the cold ENSO phase, a time when SF abundance shows its highest fluctuations.

ABSTRACT Flight dispersal of the triatomine bug species Rhodnius pallescens Barber, the principal vector of Chagas disease in Panama, is an important mechanism for spreading Trypanosoma cruzi, causative agent of Chagas disease. This study measures R. pallescens flight performance using a tethered flight mill both when uninfected, and when infected with T. cruzi or Trypanosoma rangeli. Forty-four out of the 48 (91.7%) insects initiated flight across all treatments, and trypanosome infection did not significantly impact flight initiation. Insects from all treatments flew a cumulative distance ranging from 0.5 to 5 km before fatiguing. The median cumulative distance flown before insect fatigue was higher in T. cruzi- and T. rangeli-infected insects than in control insects; however, this difference was not statistically significant. There was a positive relationship between parasite load ingested and time until flight initiation in T. rangeli-infected bugs, and T. rangeli- and T. cruzi-infected females flew significantly faster than males at different time points. These novel findings allow for a better understanding of R. pallescens dispersal ability and peridomestic management strategies for the prevention of Chagas disease in Panama.

This study was designed to examine the height-for-age z-scores (HAZ), and the prevalence of intestinal inflammation, gastrointestinal infections with parasites, and enteroaggregative Escherichia coli (EAEC) in rural Panamanian children. Stool microscopy and polymerase chain reaction (PCR) testing for EAEC detected Giardia lamblia (32%, 32 of 100) and EAEC (13%, 11 of 87) in the study participants, respectively. Anthropometric analyses showed that those children who were > 12 months of age had lower HAZ scores (mean of -1.449) than the reference population. As a group, the children in the study 1 to 5 years of age did not show recovery from the previously mentioned decline in terms of their HAZ. The HAZ means of the children infected with G. lamblia, EAEC, and Ascaris lumbricoides were -1.49, -1.67, and -2.11, respectively. Furthermore, the study participants with A. lumbricoides and EAEC infections in the presence of lactoferrin showed another decrease of 0.19 and 0.13, respectively, in their HAZ means., (© The American Society of Tropical Medicine and Hygiene.)

Although the geographic origin of malaria cases imported into the United States can often be inferred from travel histories, these histories may be lacking or incomplete. We hypothesized that mitochondrial haplotypes could provide region-specific molecular barcodes for tracing the origin of imported Plasmodium vivax infections. An analysis of 348 mitochondrial genomes from worldwide parasites and new sequences from 69 imported malaria cases diagnosed across the United States allowed for a geographic assignment of most infections originating from the Americas, southeast Asia, east Asia, and Melanesia. However, mitochondrial lineages from Africa, south Asia, central Asia, and the Middle East, which altogether contribute the vast majority of imported malaria cases in the United States, were closely related to each other and could not be reliably assigned to their geographic origins. More mitochondrial genomes are required to characterize molecular barcodes of P. vivax from these regions., (© The American Society of Tropical Medicine and Hygiene.)

The prevalence and distribution of Anaplasma and Ehrlichia were estimated in 201 symptomatic dogs from Panama by nested PCR and DNA sequencing of the 16S rRNA gene. In total, 163 dogs (70.6%; 163/201) were infected with either Anaplasma or Ehrlichia. On the basis of PCR results, the majority of cases were infected with E. canis (64.2%; 129/201) followed by 21.4% (43/201) with A. platys, whereas 7.5% (15/201) had Anaplasma/Ehrlichia co-infections. Further analyses of 16S rDNA partial sequences show sequence homology with E. canis and A. platys from other countries. Hematology findings from 79 E. canis PCR-positive dogs included anemia (74.7%), thrombocytopenia (81.9%), macroplatelets (29.1%), and leukopenia (6.3%). Among 16 A. platys PCR-positive dogs with available hematology, 62.5% were anemic, 75% had thrombocytopenia, and 100% had macroplatelets. On the basis of E.canis serology (enzyme-linked immunosorbent assay [ELISA]) (n=92 dogs), 30 dogs that were seropositive for E. canis were also PCR-positive, whereas among seronegatives (n=62), 10 were PCR-positive for E. canis. This study provides the first characterization of canine anaplasmosis and ehrlichiosis infections in Panama and is important to veterinary public health and comparative studies of these pathogens in the Americas.