Culicidae (Diptera: Culicomorpha) in the southern Brazilian collection "Ana Leuch Lozovei" with notes on distribution and diversity

Biological collections are of extreme importance in acquiring knowledge of the biodiversity of a specific environment. In this article, we organise, list and catalogue the adult specimens belonging to the family Culicidae housed in the Parasitology Collection of the Basic Pathology Department at the Federal University of Paraná, southern Brazil. To this end, a data bank was created, containing information on the taxonomy and collecting of each sample. The culicids were collected using different methodologies in 18 municipalities in the state of Paraná, between 1967 and 1999. There are 5,739 catalogued specimens of which 4,704 (81.96%) are identified at a specific level, with a diversity of 205 species. Of these, 18 are new recorded samples for the state of Paraná and 3 for Brazil. This being the case, we propose, in honour of the 30 years dedicated to the study of culicids in the state of Paraná, the creation of the Ana Leuch Lozovei Entomological Collection, which is composed of insect vectors or potential vectors, of agents that cause diseases in humans. 

Culicidae (Diptera: Culicomorpha) in the southern Brazilian 'Ana Leuch Lozovei' collection, with notes on distribution and diversity.

Biological collections are important for acquiring knowledge of the biodiversity of a specific environment. Here, we organize, list and catalog adult specimens of Culicidae from the Parasitology Collection of the Basic Pathology Department at the Federal University of Paraná, Brazil, and present a databank of taxonomic and collection information for each sample. Culicids were collected using different methodologies in 18 municipalities in Paraná state, between 1967 and 1999. There are 5,739 catalogued specimens, of which 4,703 (81.94%) are identified at species level, with a diversity of 100 species. Of these, 18 are new recorded samples for Paraná, and three are new for Brazil. This collection is named the Ana Leuch Lozovei Entomological Collection in honor of the 30 years Prof Lovozei dedicated to culicid study in Paraná. The collection comprises insect vectors or potential vectors, of agents causing human diseases.

Identification and Molecular Characterization of Four New Blastocystis Subtypes Designated ST35-ST38.

Three recent studies of Blastocystis epidemiology in mammalian hosts identified four novel sequences that appeared to share B. lapemi as the most similar sequence. However, full-length ssu rRNA gene sequences were not available to confirm the validity of these new subtypes. In the present study, Nanopore MinION sequencing was used to obtain full-length reference sequences for each of the new subtypes. Additionally, phylogenetic analyses and pairwise distance comparisons were performed to confirm the validity of each of these new subtypes. We propose that the novel sequences described in this study should be assigned the subtype designations ST35-ST38. The full-length reference sequences of ST35-ST38 will assist in accurate sequence descriptions in future studies of Blastocystis epidemiology and subtype diversity.

Selective isolation of agents of chromoblastomycosis from insect-associated environmental sources.

Chromoblastomycosis is a neglected disease characterized by cutaneous, subcutaneous or disseminated lesions. It is considered an occupational infectious disease that affects mostly rural workers exposed to contaminated soil and vegetal matter. Lesions mostly arise after a traumatic inoculation of herpotrichiellaceous fungi from the Chaetothyriales order. However, the environmental niche of the agents of the disease remains obscure. Its association with insects has been predicted in a few studies. Therefore, the present work aimed to analyze if social insects, specifically ants, bees, and termites, provide a suitable habitat for the fungi concerned. The mineral oil flotation method was used to isolate the microorganisms. Nine isolates were recovered and phylogenetic analysis identified two strains as potential agents of chromoblastomycosis, i.e., Fonsecaea pedrosoi CMRP 3076, obtained from a termite nest (n = 1) and Rhinocladiella similis CMRP 3079 from an ant exoskeleton (n = 1). In addition, we also identified Fonsecaea brasiliensis CMRP 3445 from termites (n = 1), Exophiala xenobiotica CMRP 3077 from ant exoskeleton (n = 1), Cyphellophoraceae CMRP 3103 from bees (n = 1), Cladosporium sp. CMRP 3119 from bees (n = 1), Hawksworthiomyces sp. CMRP 3102 from termites (n = 1), and Cryptendoxyla sp. from termites (n = 2). The environmental isolate of F. pedrosoi CMRP 3076 was tested in two animal models, Tenebrio molitor and Wistar rat, for its pathogenic potential with fungal retention in T. molitor tissue. In the Wistar rat, the cells resembling muriform cells were observed 30 d after inoculation.

Vaccuuming method as a successful strategy in the diagnosis of active infestation by Pediculus humanus capitis.

Most human epidemiological and clinical studies use visual inspection of the hair and scalp to diagnose Pediculus humanus capitis , however this method has low sensitivity to diagnose active infestations (presence of nymphs and adult lice). Vacuuming the hair and scalp has been used as a diagnostic method, but there are no previous data comparing its effectiveness with visual inspection. The aim of this study was to determine the prevalence of overall infestation (nits and trophic stages), of active infestation by Pediculus humanus capitis , and to evaluate the effectiveness of vacuuming in comparison with the visual inspection. Visual inspection was performed by three examiners and vacuuming of the scalp by one investigator, with an adapted vacuum cleaner. A total of 166 children aged 4 to 10 years old were randomly selected from public schools in Southern Brazil. Considering the positive results obtained by both methods, the prevalence of overall infestation was 63.3%, whereas active infestation was 18.7%. The visual inspection was more effective on diagnosing overall infestation, however, its effectiveness to detect active infestation was lower, ranging from 0.6% (RR=3%, p<0.001) to 6.6% (RR=35%, p=0.001), depending on the number of examiners. The effectiveness of vacuuming to diagnose active infestation was higher than the one of visual inspection, with a prevalence rate of 16.3% (RR=87%, p=0.332). As presented in our study, the vacuuming method was 2.74 to 7.87 times most likely to detect active infestation, thus it could be adopted as a more accurate method to diagnose active pediculosis.

Reduced prevalence of soil-transmitted helminths and high frequency of protozoan infections in the surrounding urban area of Curitiba, Paraná, Brazil.

Human populations living in the surrounding urban areas of large Brazilian cities have increased vulnerability to intestinal parasites. However, the epidemiological scenario of soil-transmitted helminths (STH) in Curitiba, Paraná's main city, remains largely unknown. To bridge this gap of knowledge, this study aims to determine the prevalence of intestinal parasites and to investigate potential transmission pathways of the most prevalent species detected. We conducted a cross-sectional epidemiological study between July and September 2014 among schoolchildren in urban and peri-urban (deprived) areas of the municipality of Campo do Tenente, Curitiba. A total of 549 stool samples were used for coproparasitological diagnosis. Microscopy-positive samples of the most common species found were re-assessed by PCR and sequencing methods at the small subunit rRNA gene. Prevalence of infection by any given enteroparasite was 24.8%, but soil-transmitted helminths were only detected in 3.5% of the examined samples. Frequency of protozoan infections reached 90% and 97.8% in single and multiple infections, respectively. Blastocystis sp. (38.9%) was the most frequently species found in the surveyed schoolchildren population. A total of 41 Blastocystis-positive samples were unambiguously typed as ST1 (36.4%), ST2 (21.2%), ST3 (39.4%), and ST1 + ST3 mixed infection (3.0%). These results indicate that Blastocystis transmission is primarily anthroponotic in origin. This data highlights the importance of maintaining the anthelminthic control programs currently in place and of improving sanitary disposal of human excreta in poor-resource settings.

Abundancia y diversidad de vectores (Diptera: Psychodidae) en área de transmisión antigua de Leishmaniasis Cutánea en el nuevo mundo después de la construcción del gasoducto Bolivia-Brasil / Abundance and diversity of vectors (Diptera: Psychodidae) in an old transmission area of cutaneous leishmaniasis in the new world after Bolivia-Brazil gas pipeline construction

The objective of this study was to determine the abundance and diversity of the species and their presumed vectorial role in the transmission of Cutaneous Leishmaniasis (CL). Research on sand flies (Diptera: Psychodidae) was carried out in two municipalities in the Ribeira River Valley in the State of Paraná, southern Brazil. One, Adrianópolis, is an endemic area of CL and the other is a recent outbreak area (Cerro Azul). A total of 432 specimens were collected from residential, peri-domestic and wild environments. According to the data obtained, for each ecotope studied, the statistical analysis has shown that two lines of the Lutzomyia intermedia population coexist and that the quantity of specimens from L. intermedia s.l. is significantly different from L. intermedia s.s. in all environments (χ2=9.943; DF=2; p=0.07). The prevalence of L. intermedia (93.28%) in the ecotopes studied suggests that it is the main vector of leishmaniasis. The succession of vector species and their involvement in the epidemiological cycle of L. braziliensis is discussed in this article(AU)

El objetivo de este trabajo fue determinar la abundancia y diversidad de las especies y su supuesto papel vectorial en la transmisión de Leishmaniasis Cutánea (LC). La investigación sobre flebótomos (Diptera: Psychodidae) se llevó a cabo en dos municipios en el Valle del Río Ribeira en el Estado de Paraná, sur de Brasil. Las regiones estudiadas fueron Adrianópolis, área endémica de LC, y Cerro Azul, que se reporta como zona de brote reciente. Se recolectaron un total de 432 especímenes de ambientes residenciales, peri-domésticos y silvestres. El análisis estadístico realizado, a partir de los datos obtenidos para cada ecotopo estudiado, mostró que coexistían dos linajes de la población de Lutzomyia intermedia y que la cantidad de especímenes de L. intermedia s.l. fue significativamente diferente de L. intermedia s.s. en todos los entornos (χ2=9,943; GL=2; p=0,07). La prevalencia de L. intermedia (93,28%) en los ecotopos estudiados sugiere que es el principal vector de la leishmaniasis. La sucesión de especies de vectores y su participación en el ciclo epidemiológico de L. braziliensis se discute en este artículo(AU)

Scalp microbiota alterations in children with pediculosis.

Pediculosis is a disease caused by the insect Pediculus humanus capitis that mainly occurs in childhood. A comparative study was carried out evaluating groups of schoolchildren with (group A) and without pediculosis (group B) to analyse the characteristics of the scalp microbiota. Samples were collected by swab using Stuart transport medium and incubate in Sabouraud dextrose agar with tetracycline to analyse the fungal microbiota and in blood agar to assess the bacterial microbiota. The isolates identity was confirmed by sequencing of the 16S and 18S regions of the ribosomal DNA gene for bacteria and fungi, respectively. The analysis of the 186 isolates led to the identification of 35 bacteria and 40 fungi in group A and 47 bacteria and 64 fungi in group B. The results indicate differences in bacterial and fungal species in the groups analysed. In the observed bacterial microbiota, Staphylococcus capitis occurred more frequently than Staphylococcus epidermidis in group A vs B. Among fungal isolates, Debaryomyces sp. was more frequent in group B vs A. Our findings showed scalp microbiota alterations in children with pediculosis, meriting future studies to analyse the relationship between these agents and their impact on human health.

Prevalence of intestinal parasites, with emphasis on the molecular epidemiology of Giardia duodenalis and Blastocystis sp., in the Paranaguá Bay, Brazil: a community survey.

BACKGROUND: Intestinal protozoan parasites are major contributors to the global burden of gastrointestinal disease causing significant socioeconomic consequences. Children living in resource-poor settings with restricted access to water and sanitary services are particularly at risk of these infections. METHODS: A prospective, community-based, cross-sectional survey was conducted in Paraná (southern Brazil) between May 2015 and May 2016. A total of 766 stool samples were individually collected from volunteers (male/female ratio: 0.99; age range: 0-76 years) and used for investigating the presence of intestinal helminth and protozoan species by routine microscopic procedures including the Kato-Katz and modified Ritchie concentration methods and the Ziehl-Neelsen stain technique. Quantitative real-time PCR confirmed microscopy-positive samples for Giardia duodenalis and the assemblages and sub-assemblages determined by multilocus sequence-based genotyping of the glutamate dehydrogenase (gdh) and ß-giardin (bg) genes of the parasite. Identification of Blastocystis subtypes was carried out by amplification and sequencing of a partial fragment of the small-subunit ribosomal RNA gene (SSU rDNA) of this heterokont microorganism. RESULTS: Overall, 46.1% (353/766) of the participants were infected/colonised by at least one intestinal parasite/commensal species. Protozoan and helminth species were detected in 42.7% and 10.1% of the surveyed population, respectively. Blastocystis sp. (28.2%), Endolimax nana (14.9%), and Giardia duodenalis (11.0%) were the most prevalent species found among protozoans and Ascaris lumbricoides (5.0%), Trichuris trichiura (4.6%) and hookworms (1.0%) among helminths. A total of 38 G. duodenalis-positive samples were genotyped at gdh and bg markers, revealing the presence of the sub-assemblages AII (47.4%), AII/AIII (2.6%), BIII (5.3%), BIV (26.3%) and BIII/BIV (13.1%). Two samples (5.3%) were only identified as assemblage B. AII was predominantly found in females aged 5-9 years and was associated with a higher likelihood of reporting gastrointestinal symptoms. A total of 102 Blastocystis-positive samples were successfully subtyped at the SSU rRNA gene revealing the presence of ST1 (36.3%), ST2 (15.7%), ST3 (41.2%), ST4 (2.9%), ST6 (1.0%) and ST8 (2.9%). CONCLUSIONS: Data presented here indicate that enteric parasites still represent a pressing health concern in Paraná, Brazil, probably due to sub-optimal water, sanitation and hygiene conditions. A mostly anthroponotic origin is suspected for G. duodenalis and Blastocystis sp. infections.

Intestinal symptoms and Blastocystis load in schoolchildren of Paranaguá Bay, Paraná, Brazil.

The symptomatology of Blastocystis cannot be attributed to any particular subtype, although can be related to a high Blastocystis infection load. One stool sample from each of 217 schoolchildren of Vale de Sol Paranaguá Bay (Paraná, Brazil) was collected. Three milliliters of each fixed stool sample were processed applying the formalin-ether concentration technique. After obtaining the overall prevalence of intestinal parasites, quantification was carried out in Blastocystis positive samples. A total of 75/217 (34.6%) children suffered from intestinal symptoms (abdominal pain and/or persistent diarrhea), of whom 41.3% (31/75) presented moderate/heavy Blastocystis load with a statistically significant risk to present intestinal symptoms (OR 0.039 [0.006-0.15]; p<0.001) Moreover, those symptomatic schoolchildren monoparasitized only by Blastocystis (10/75, 13.3%), and those polyparasitized by Blastocystis with other non-pathogenic species (15/75, 20%) with moderate/heavy loads, also entail a statistically significant risk of intestinal symptomatology, both in monoparasitism (12%, OR 0.10 [0.004-0.63]; p=0.021) and in polyparasitism with a non-pathogenic parasite (18.6%, OR 0.059 [0.002-0.35]; p=0.001). For the first time in Brazil, using data from schoolchildren of Paranaguá Bay, we demonstrated that moderate/ heavy loads of Blastocystis could be related to intestinal symptoms.

Limiar de positividade e sensibilidade dos métodos de Faust et al. e Lutz para detecção de cistos de Giardia duodenalis / Positivity threshold and sensitivity of methods of Faust et al. and Lutz for detection of Giardia duodenalis cysts

Objetivo: O presente estudo teve como objetivo determinar e comparar o limiar de positividade e a sensibilidade dos métodos de centrífugo-flutuação em sulfato de zinco (Faust et al.) e sedimentação espontânea (Lutz) para o diagnóstico de cistos de Giardia duodenalis. Métodos: Para obtenção de amostras fecais com quantidades conhecidas de cistos de G. duodenalis, amostras positivas para o parasito foram purificadas e quantificadas, e posteriormente alíquotas com diferentes quantidades foram adicionadas a amostras fecais negativas para parasitos. Após a contaminação de oito amostras negativas com quantidades variando entre 1.000 e 200.000 cistos por grama de fezes (c/g/f), as mesmas foram submetidas aos métodos de Faust et al. e Lutz, onde o primeiro se mostrou mais sensível para a detecção de cistos de G. duodenalis. Resultados: O limiar de positividade do método de Faust et al. foi de 11.000 c/g/f, e do método de Lutz foi de 100.000 c/g/f, portanto, cargas parasitárias inferiores a esses limiares levariam a resultados falso-negativos. Conclusão: O método de Lutz não é adequado para o diagnóstico de giardíase, portanto deve ser sempre utilizado em conjunto com o método de Faust et al.

Objective: The present study aimed to determine and compare the positivity threshold and sensitivity of the methods of zinc sulfate centrifugal flotation (Faust et al.) and spontaneous sedimentation (Lutz) for the diagnosis of Giardia duodenalis. Methods: To obtain fecal samples containing known amounts of G. duodenalis cysts, the samples with the parasite were purified and quantified, then aliquots with different amounts were added to fecal samples negative for parasites. After the contamination of eight negative samples with amounts ranging between 1.000 and 200.000 cysts per gram of feces, they were subjected to methods of Faust et al. and Lutz, where the first was more sensitive for the detection of G. duodenalis cysts. Results: The positivity threshold of the method of Faust et al. was 11.000 c/g/f, and the method of Lutz was 100.000 c/g/f, so parasitic loads below those thresholds would lead to false-negative results. Conclusion: The method of Lutz is not suitable for the diagnosis of giardiasis, therefore must always be associated with the method Faust et al.

Trypanosoma cruzi vector infection rate is underestimated in some localities in the state of Bahia

Bahia was the last Brazilian state declared free of Chagas disease transmission by Triatoma infestans in 2006. The program designed to control vector transmission of Chagas is currently active, and all potential triatomines collected by the Bahia State Department of Health officials are most frequently diagnosed as negative for Trypanosoma cruzi when analyzed by the conventional parasitological direct method. The aim of the current study was to investigate whether triatomines from Bahia are free of T. cruzi infection using a more sensitive diagnostic methodology, namely the kinetoplastid-DNA polymerase chain reaction (kDNA-PCR). With the help of health officials, 51 triatomines were analyzed from peridomicile areas within the central north region of the state of Bahia. The majority (60.8%) were Triatoma brasiliensis, 29.4% were Triatoma pseudomaculata, and 9.8% were unidentified nymphs. Only one insect tested potentially positive for T. cruzi by the conventional parasitological direct method, and 31.4% were positive for T. cruzi DNA by kDNA-PCR. Almost half the infected insects (41.9%) were T. brasiliensis, a species with high potential for T. cruzi transmission. These results demonstrate that the number of infected triatomines with high transmission potential of T. cruzi may be greater than expected in four localities in the state of Bahia

Detection of Streptococcus mutans using padlock probe based on Rolling Circle Amplification (RCA)

The aim of this study was to develop and evaluate a padlock probe based on the Rolling Circle Amplification (RCA), which targeted to 16S-23S rDNA region of S. mutans. The specificity of developed padlock probe was tested for DNA within a panel strains, including S. mutans isolated from the saliva and reference strains of the genus Streptococcus, as well as total DNA samples of biofilm and saliva. The results were positive either for DNA samples of S. mutans or DNA samples recovered from the biofilm and saliva revealing the specificity of designed padlock probe. The padlock probe based on the RCA was proved to be an effective, reproducible method for S. mutans detection and demonstrated the possibility of a rapid detection and accurate identification of S. mutans infection.

Molecular detection of the blood meal source of sand flies (Diptera: Psychodidae) in a transmission area of American cutaneous leishmaniasis, Paraná State, Brazil.

The feeding behavior of sand flies provides valuable information about the vector/host interactions and elucidates the epidemiological patterns of American cutaneous leishmaniasis (ACL) transmission. The aim of this study was to identify the blood meal sources of sand flies in endemic areas of leishmaniasis in Paraná State through polymerase chain reaction (PCR) amplification of a prepronociceptin (PNOC) gene fragment and its subsequent DNA sequencing. Moreover, molecular assays were conducted to evaluate the sensitivity and reproducibility of the PNOC gene amplification. Besides that, a time-course digestion test of the blood using sand flies that fed artificially on BALB/c mice was performed. Of 1263 female sand flies collected in the field, 93 (3.6%) specimens were engorged and 27 allowed efficient amplification of the PNOC gene. These flies had fed on equine (Equus caballus), porcine (Sus scrofa) and canine (Canis lupus familiaris) species. The results also showed that the identification of the blood meal sources of the sand flies using the molecular method was directly linked to the level of digestion of the blood (time-course) and not to the amount of blood that had been ingested or to the presence of inhibitors in the blood.

Using molecular markers to assess Streptococcus mutans variability and the biological risk for caries

AIM: To characterize the genetic variability of Streptococcus mutans isolates and to correlate this variability with different colonization profiles observed during dental caries in a sample of children. METHODS: S. mutans samples were isolated from the saliva of 30 children with varying histories of dental caries, and they were characterized according to morphological and biochemical markers and the sequences of their 16S-23S intergenic spacer region. The genetic variability of the isolates was first assessed using Random Amplified Polymorphic DNA (RAPD) markers. Next, the isolates were differentiated by sequencing a specific region of the gene encoding the enzyme glucosyltransferase B (gtfB). RESULTS: Characterization using RAPD markers uncovered significant genetic variability among the samples and indicated the existence of clusters, which allowed us to reconstruct both the origin and clinical history of the disease. By sequencing the 16S-23S intergenic region, it was found that all of the isolates belonged to the species S. mutans. Based on the genetic similarity of the isolates and pattern of amino acid variations identified by partial sequencing of the gtfB gene, base-pair changes were identified and correlated with different virulence patterns among the isolates. CONCLUSIONS: The partial sequencing of the gtfB gene can be a useful tool for elucidating the colonization patterns of S. mutans. As amino acid variations are likely to be correlated with differences in biological risk, molecular characterization, such as that described in this paper, could be the key for assessing the development of dental caries in children...

High occurrence of Calodium hepaticum (syn. Capillaria hepatica) spurious infection in a village in the Atlantic Forest of southern Brazil

Calodium hepaticum (syn. Capillaria hepatica) is a nematode of the Capillariidae family that infects rodents and other mammals. In Brazil, human spurious infections of C. hepaticum have been detected in indigenous or rural communities from the Amazon Basin, but not in the southern states of the country. Here, we report the highest occurrence (13.5% of 37 residents) of C. hepaticum human spurious infection detected in Brazil and the first record in a southern region, Guaraqueçaba. The finding is explained by the area being located in the Atlantic Forest of the state of Paraná, surrounded by preserved forests and because the inhabitants consume the meat of wild mammals.

High occurrence of Calodium hepaticum (syn. Capillaria hepatica) spurious infection in a village in the Atlantic Forest of southern Brazil.

Calodium hepaticum (syn. Capillaria hepatica) is a nematode of the Capillariidae family that infects rodents and other mammals. In Brazil, human spurious infections of C. hepaticum have been detected in indigenous or rural communities from the Amazon Basin, but not in the southern states of the country. Here, we report the highest occurrence (13.5% of 37 residents) of C. hepaticum human spurious infection detected in Brazil and the first record in a southern region, Guaraqueçaba. The finding is explained by the area being located in the Atlantic Forest of the state of Paraná, surrounded by preserved forests and because the inhabitants consume the meat of wild mammals.

Morphological, genotypic, and physiological characterization of Acanthamoeba isolates from keratitis patients and the domestic environment in Vitoria, Espírito Santo, Brazil.

Amoebae of the genus Acanthamoeba are free-living protozoa that can cause granulomatous encephalitis and keratitis in humans. In this study, four clinical and three household dust isolates obtained in Vitória, Espírito Santo, Brazil were characterized by their morphological, genotypic, and physiological properties. All isolates belonged to group II according to Pussard and Pons' cyst morphology. Analysis of their 18S rDNA sequence identified one isolate from household dust as genotype T11 and the others six samples as genotype T4. Five T4 isolates presented a highly variable region (DF3) in 18S rDNA identical to those previously described. Physiological assays carried out with trophozoites in co-culture with bacteria or in axenic conditions showed all samples tolerated temperatures up to 37°C, regardless of culture method. One keratitis isolate grew at 42°C in co-culture with bacteria. Most isolates in co-culture survived at 1.0M, except a T11 isolate, which tolerated up to 0.5M. The isolates did not grow at 42°C and did not tolerate 0.5M and 1.0M under axenic condition. This is the first report of 18S rRNA gene genotyping applied to Acanthamoeba isolated from keratitis patients in Brazil. The results also indicated that osmo-tolerance is dependent on the culture system.

Phenetic analysis of Panstrongylus megistus Burmeister, 1835 (Hemiptera: Reduviidae: Triatominae) in the State of Paraná-Brazil

Panstrongylus megistus is an important Chagas Disease vector and is said to be one of the species that might replace Triatoma infestans as the main vector of that disease in Brazil. The different degrees of P. megistus domiciliation in Brazil and its epidemiological relevance draw forth the need for the development of genetic studies that make it possible to analyze and understand the interchange of individual and gene fluxes among different populations. Thus, the present work aimed at studying the genetic variability of P. megistus in the State of Paraná - south of Brazil- and at comparing it with populations of the same species from five other states in Brazil (SP, MG, SC, RS, SE). In order to attain the proposed objective, 25 populations were studied using fifteen isoenzymatic systems (6PGD, G6PD, ME¹, ME², ICD, PGM, GPI, GOT¹, GOT², NP¹, NP², DIA, MPI, F, and MDH). The phenetic analysis allowed the individuation of 22 electromorphs and five zymodemes. The G6PD enzyme was the only polymorphic one presenting four electromorphs for the studied populations, all of them described for the State of Paraná-BR. The P. megistus populations from other states grouped with those from Paraná-BR, evidencing a low genetic variability in that species. Despite the existing geographic barriers, sub-samples - away from one another by at most 570km - were grouped in one and the same zymodeme. The epidemiological implications of such results are discussed in the present work.

Panstrongylus megistus é um importante vetor da Doença de Chagas e é apontado como uma das espécies com potencial para substituir Triatoma infestans como principal vetor desta doença no Brasil. Os diferentes graus de domiciliação por P. megistus - no Brasil - e sua importância epidemiológica evocam a necessidade de estudos com bases genéticas que possibilitem analisar e compreender os intercâmbios de indivíduos e os fluxos gênicos entre as distintas populações. Assim, o presente trabalho tem como objetivo estudar a variabilidade genética de P. megistus no Estado do Paraná e compará-los com populações da mesma espécie de cinco estados do Brasil (SP, MG, SC, RS, SE). Para atingir o objetivo proposto, 25 populações foram estudadas empregando quinze sistemas isoenzimáticos (6PGD, G6PD, ME¹, ME², ICD, PGM, GPI, GOT¹, GOT², NP¹, NP², DIA, MPI, FH e MDH). A análise fenética permitiu a individualização de 22 eletromorfos e quatro zimodemas. A enzima G6PD foi a única polimórfica que apresentou quatro eletromorfos para as populações estudadas, todas descritas para o Estado do Paraná. As populações de P. megistus procedentes dos outros estados agruparam-se com as do Paraná, demonstrando haver baixa variabilidade genética na espécie. Apesar das barreiras geográficas existentes, sub-amostras - distantes entre si por até 570 km - ficaram reunidas num mesmo zimodema. As implicações epidemiológicas destes resultados são discutidas no presente trabalho.

Phylogeography and genetic variation of Triatoma dimidiata, the main Chagas disease vector in Central America, and its position within the genus Triatoma.

BACKGROUND: Among Chagas disease triatomine vectors, the largest genus, Triatoma, includes species of high public health interest. Triatoma dimidiata, the main vector throughout Central America and up to Ecuador, presents extensive phenotypic, genotypic, and behavioral diversity in sylvatic, peridomestic and domestic habitats, and non-domiciliated populations acting as reinfestation sources. DNA sequence analyses, phylogenetic reconstruction methods, and genetic variation approaches are combined to investigate the haplotype profiling, genetic polymorphism, phylogeography, and evolutionary trends of T. dimidiata and its closest relatives within Triatoma. This is the largest interpopulational analysis performed on a triatomine species so far. METHODOLOGY AND FINDINGS: Triatomines from Mexico, Guatemala, Honduras, Nicaragua, Panama, Cuba, Colombia, Ecuador, and Brazil were used. Triatoma dimidiata populations follow different evolutionary divergences in which geographical isolation appears to have had an important influence. A southern Mexican-northern Guatemalan ancestral form gave rise to two main clades. One clade remained confined to the Yucatan peninsula and northern parts of Chiapas State, Guatemala, and Honduras, with extant descendants deserving specific status. Within the second clade, extant subspecies diversity was shaped by adaptive radiation derived from Guatemalan ancestral populations. Central American populations correspond to subspecies T. d. dimidiata. A southern spread into Panama and Colombia gave the T. d. capitata forms, and a northwestern spread rising from Guatemala into Mexico gave the T. d. maculipennis forms. Triatoma hegneri appears as a subspecific insular form. CONCLUSIONS: The comparison with very numerous Triatoma species allows us to reach highly supported conclusions not only about T. dimidiata, but also on different, important Triatoma species groupings and their evolution. The very large intraspecific genetic variability found in T. dimidiata sensu lato has never been detected in a triatomine species before. The distinction between the five different taxa furnishes a new frame for future analyses of the different vector transmission capacities and epidemiological characteristics of Chagas disease. Results indicate that T. dimidiata will offer problems for control, although dwelling insecticide spraying might be successful against introduced populations in Ecuador.