Population genetic structure and morphological diversity of Cruzia tentaculata (Nematoda: Ascaridida), a parasite of marsupials (Didelphinae), along the Atlantic Forest on the eastern coast of South America.

Cruzia tentaculata is a helminth parasite of marsupials and has a wide geographic distribution from Mexico to Argentina. The aim of this study was to analyse the genetic population structure of this nematode along the Atlantic Forest biome. Cruzia tentaculata specimens were recovered from Didelphis aurita, Didelphis albiventris and Philander quica in 9 localities. Morphological and morphometric data were investigated for phenotypic diversity among localities and hosts using multivariate discriminant analysis of principal components. Phylogenetic relationships of C. tentaculata were determined using maximum likelihood and Bayesian inference. The population structure was analysed by fixation indices, molecular variance analysis, Tajima's D and Fu's Fs neutrality tests, Mantel tests and Bayesian clustering analysis. A higher significant morphometric difference for males was observed between localities. In the haplogroup networks, 2 groups were recovered, separating locations from the north and from the south/southeast. The morphometric variation in C. tentaculata between different localities was compatible with this north and southeast/south pattern, suggesting adaptation to different ecological conditions. Population genetic analyses suggested a pattern of evolutionary processes driven by Pleistocene glacial refugia in the northeast and southeast of the Atlantic Forest based on the distribution of genetic diversity.

Population genetic structure and phenotypic diversity of Aspidodera raillieti (Nematoda: Heterakoidea), a parasite of Didelphini marsupials in Brazil's South and Southeast Atlantic Forest.

BACKGROUND: The population genetics of parasites may be influenced by host specificity, life cycle, host geographical range, evolutionary history, and host population structure. The nematode Aspidodera raillieti infects different marsupial and rodent hosts in the Nearctic and Neotropical regions, implying a gene flow among populations. However, niche diversification of the main hosts of A. raillieti in superimposed areas may provide conditions for population genetic structuring within this parasite species. We examined the genetic structuring of A. raillieti infecting three marsupial species co-occurring along the South and Southeast Brazilian Atlantic Forest, a hotspot of biodiversity. METHODS: We employed morphometric analyses and partial mitochondrial cytochrome c oxidase I gene sequences (MT-CO1) to characterize populations via phylogenetic and phylogeographic analyses. RESULTS: Among 175 A. raillieti specimens recovered from the marsupial hosts Didelphis aurita, D. albiventris, and Philander quica, we identified 99 MT-CO1 haplotypes forming four haplogroups and four clades in networks and phylogenetic trees, respectively. Clades I and II encompassed parasites of D. albiventris from the South region, clade III comprised parasites of D. aurita from the South and Southeast regions, and clade IV encompassed parasites of D. aurita and D. albiventris from the South and Southeast regions and parasites of P. quica from the South region. High genetic differentiation between clades, with a high fixation index and greater genetic variation in the analysis of molecular variance (AMOVA), indicated low gene flow between clades. Haplotypes shared among host species revealed a lack of host specificity. A significant correlation in the Mantel test suggested parasite isolation by distance, while there was no evidence of geographical structure between populations. Negative neutrality test values for clades III and IV suggested recent population expansion. Morphometric differentiation between A. raillieti specimens recovered from different host species, as well as from different localities, was more evident in males. CONCLUSION: The genetic structure of A. raillieti populations in the South and Southeast Atlantic Forest resulted from historical events rather than from current geographical distribution or host specificity. We also demonstrate morphometric variation associated with host species and localities, suggesting phenotypic plasticity to host attributes and to spatial variables.

Bats from the Pedra Branca Forest, Rio de Janeiro, Brazil.

The Pedra Branca Forest is located in a highly-urbanised region of the central portion of Rio de Janeiro City, comprises the largest urban forest on the continent and is isolated from other Atlantic Forest remnants. The local flora and fauna are protected by three conservation units (Pedra Branca State Park, Prainha Municipal Natural Park and Guaratiba State Biological Reserve) and one biological station (Fiocruz Atlantic Forest Biological Station-EFMA). Here, we provide an updated list of the bat fauna for the remnant. The results are based on samplings at EFMA and literature data from Pedra Branca State Park and Prainha Natural Park. The three sampling sites combined resulted in 31 species, 23 genera and four families. Phyllostomidae was the richest family with 24 species, followed by Vespertilionidae with five species (3%) and Molossidae and Noctilionidae with one species. The local bat fauna was predominantly composed of species with a broad geographic distribution.

Three new species of Eimeria Schneider 1875 in the montane grass mouse, Akodon montensis (Rodentia: Cricetidae: Sigmodontinae), and redescription of Eimeria zygodontomyis Lainson and Shaw 1990 from southeastern Brazil.

We describe three new coccidian species of the genus Eimeria Schneider 1875 (Apicomplexa: Eimeriidae) and redescribe and report Eimeria zygodontomyis Lainson and Shaw, 1990 in the montane grass mouse, Akodon montensis Thomas, 1913 from the Serra dos Órgãos National Park in southeastern Brazil. Sporulated oocysts of Eimeria zygodontomyis are ellipsoidal to cylindrical with a 0.6 (0.5-0.8) µm thick very delicate bi-layered wall; length × width (n = 49) 18.3 × 12.5 (16-20 × 11-13) µm; length/width ratio of 1.4 (1.2-1.6); 1 polar granule occasionally present; micropyle, residuum both absent. Sporocysts are ellipsoidal; length × width 8.5 × 5.2 (8-11 × 5-6) µm; length/width ratio of 1.5 (1.3-1.7) µm; Stieda body is prominent; sub-Stieda body is absent; sporocyst residuum is compact. Sporulated oocysts of Eimeria montensis n. sp. are spheroidal to subspheroidal with a 1.2 (1.0-1.4) µm thick bi-layered wall; outer layer lightly pitted; length × width (n = 30) 16.3 × 12.5 (15-17 × 13-15) µm; length/width ratio of 1.3 (1.0-1.4); 1 polar granule present; micropyle, residuum both absent. Sporocysts are ellipsoidal; length × width 7.2 × 5.1 (6-8 × 4-6) µm; length/width ratio of 1.4 (1.2-1.6); Stieda body is present, sub-Stieda body is absent; sporocyst residuum consists of small, scattered granules. Sporulated oocysts of Eimeria uricanensis n. sp. are ovoidal to pyriform with a 1.4 ( 1.3-1.6) µm thick bi-layered wall; outer layer lightly pitted; length × width (n = 40) 26.6 × 18.6 (23-30 × 17-20) µm; length/width ratio of 1.4 (1.3-1.6); 1 polar granule present; micropyle, residuum both absent. Sporocysts are ellipsoidal, length × width 13.3 × 8.0 (10-16 × 7-9) µm; length/width ratio of 1.7 (1.5-1.9); Stieda body, sub-Stieda body both absent; sporocyst residuum consists of a cluster of granules, forming a spheroid mass. Sporulated oocysts of Eimeria parnasiensis n. sp. are subspheroidal to ellipsoidal with a 1.8 ( 1.3-2.4) µm thick bi-layered wall; outer layer lightly pitted; length × width (n = 54) 28.2 × 21.9 (26-32 × 19-28) µm; length/width ratio of 1.3 (1.2-1.4); 1 polar granule present; micropyle is absent; oocyst residuum is present and consists of a cluster of granules of varying thickness. Sporocysts are ovoidal, tapering towards the Stieda body; length × width 12.2 × 7.6 (10-13 × 6-9) µm; length/width ratio of 1.6 (1.4-1.9); Stieda body is present; sub-Stieda body is absent; sporocyst residuum is present and consists of an aggregate of thin granules.

Metacommunity Structure of Helminths of Necromys lasiurus (Rodentia: Sigmodontinae) in Different Land Use Areas in the Brazilian Cerrado.

The ecology of host-parasite interactions can be studied in the infracommunity and component community scales, which may show different patterns in species distributions, interacting and affecting each other on a regional scale. Few studies have been carried out concerning the structure and variation of the helminth communities of wild rodents in Brazil. The rodent Necromys lasiurus is typical from the mammalian fauna of the Cerrado biome; however, the environmental disturbances are making this species occur in rural areas and in other biomes where it may act as host/reservoir of many diseases. This study aimed to describe the composition and structure of the helminth metacommunity in the rodent N. lasiurus in the Brazilian Cerrado, Uberlândia, State of Minas Gerais, using the analysis of metacommunity structure. Rodents were sampled in 3 habitat types: borders of soybean and corn plantations, pasturelands, and preserved areas of Cerrado borders. Adult helminths of 8 species were found in the gastrointestinal tract: Protospirura numidica criceticola, Physaloptera sp., Pterygodermatites ( Paucipectines) zygodontomis (Spirurida), Stilestrongylus freitasi (Rhabditida), Trichuris navonae (Trichurida) and Syphacia ( Syphacia) alata (Oxiurida) of the Phylum Nematoda; Rodentolepis akodontis (Cyclophyllidea) of the Phylum Platyhelminthes; and Moniliformis sp. (Moniliformida) of the Phylum Acanthocephala. Season and the kind of land use favored some helminths species in this rodent, especially in the plantation area, although diversity was not largely influenced by the land use. Plantation areas could provide an increase in the host abundance and the occurrence of other rodent species, favoring a higher rate of parasite exchange among different hosts. A checkerboard structure of metacommunity was found on the infracommunity scale, which suggests the existence of interspecific competition. A quasi-nested structure of metacommunity was observed on the component community scale showing that most species were influenced by the same environmental gradient and that the species-poor communities were subsets of species-rich communities. Syphacia alata, P. zygodontomis, S. freitasi, and R. akodontis were dominant species in all habitats and represented the core-species in the metacommunity.

A new species of Eimeria Schneider, 1875 from the Serra dos Órgãos National Park, Rio de Janeiro, Brazil, with notes on its endogenous development in the montane grass mouse, Akodon montensis Thomas, 1913 (Rodentia: Sigmodontinae).

A total of 53 specimens of the montane grass mouse, Akodon montensis Thomas, 1913 were collected in the Serra dos Órgãos National Park (SONP) in November 2014 and July 2015. The fecal material was analyzed, and a prevalence of 7.5% was recorded for a new coccidian species of the genus Eimeria Schneider, 1875, with part of its endogenous development recorded in the small intestine. The oocysts of a new coccidian species of genus Eimeria are ellipsoidal to subspherical. The wall is bi-layered, c. 1.5 µm (1.3-1.6 µm) thick, outer layer rough. Oocyst (n = 126) mean length is 25.3 µm (21.0-28.0 µm), with a width of 20.2 µm (17.0-22.0 µm) and mean length/width (L:W) ratio of 1.3 (1.2-1.4). Polar granule is present, with the oocyst residuum as a large spherical to subspherical globule. Sporocyst shape (n = 126) is ellipsoidal, with a mean length of 11.8 µm (9.3-14.4 µm), width of 7.9 µm (6.7-9.3 µm), and mean L:W ratio of 1.5 (1.4-1.7). Sporocysts with nipple-like Stieda body and sub-Stieda body are absent. A sporocyst residuum formed by several globules, usually along the sporocyst wall. This is the first record of Eimeria in the montane grass mouse from Brazil.

Lungworm Heterostrongylus heterostrongylus Travassos, 1925 from the black-eared opossum in South America: Morphologic, histopathological and phylogenetic aspects.

Nematode parasites of the cardiopulmonary system of livestock and pet animals have been receiving attention due to the pathogenic effects they produce in the course of the infection. However, parasitism in wild animals by metastrongilid nematodes has been neglected, resulting in potential risk to wildlife. Heterostrongylus heterostrongylus is the etiological agent of bronchial pneumonia in the black-eared opossum, Didelphis aurita, a widely distributed marsupial frequently reported to inhabit areas from wild environments to peri-urban spaces. In this study, we reassessed the taxonomy of H. heterostrongylus, describing and comparing morphology of L1 and L3 larvae and adult worms with closely related angiostrongilids, and inferring phylogenetic affinities within the family Angiostrongylidae; we also detailed histopathological reactions under natural infection. Ultrastructural morphology and light microscopy confirmed cephalic structures with well-developed trilobed lips, patterns of caudal bursa rays, spicules shape, terminal anal aperture and presence of the ventral protuberance in L3 larvae, characteristics present for the family Angiostrongylidae. Our molecular phylogenetic analyses, based on the partial small subunit (SSU) 18S rRNA gene, suggested that H. heterostrongylus and Didelphostrongylus hayesi are closely related. Pathological analyses agreed with previous findings, showing that cardiopulmonary parasitism provokes verminous pneumonia and mucous bronchiolitis with hypertrophy of the smooth bronchiolar muscle. These data confirmed that H. heterostrongylus represents a pulmonary pathogen in D. aurita, produces severe pathological effects, and may represent risk to domestic and wild animals.

Serologic evidence of the exposure of small mammals to spotted-fever Rickettsia and Rickettsia bellii in Minas Gerais, Brazil.

INTRODUCTION: Sources of pathogenic Rickettsia in wildlife are largely unknown in Brazil. In this work, potential tick vectors and seroreactivity of small mammals against four spotted-fever group Rickettsia (R. rickettsii, R. parkeri, R. amblyommii and R. rhipicephali) and Rickettsia bellii from peri-urban areas of Uberlândia, a major town in Brazil, are described for the first time. METHODOLOGY: Small mammals were captured and blood samples collected. Ticks were collected from the surface of the host and the environment and posteriorly identified. Reactivity of small mammal sera to Rickettsia was tested by indirect immunofluorescence assay (IFA) using crude antigens from five Brazilian Rickettsia isolates. RESULTS: Information was obtained from 416 small mammals (48 Marsupialia and 368 Rodentia). Forty-eight animals were parasitized and two tick species, Ixodes loricatus and Amblyomma dubitatum, were found on several host species, with a few tick-host relationships described for the first time. From the 416 tested sera, 70 reacted to at least one Rickettsia antigen (prevalence of 16.8%) and from these, 19 (27.1%) reacted to two or more antigens. Seroprevalence was higher for marsupials (39.6%) than for rodents (13.8%). Marsupial and Rhipidomys spp. sera reacted mainly (highest seroprevalence and titers) to R. bellii, and that of Necromys lasiurus mainly to R. rickettsii. CONCLUSIONS: Although the serologic assays poorly discriminate between closely related spotted-fever group Rickettsia, the observed small mammal seroreactivity suggests the circulation of Rickettsia in the peri-urban area of Uberlândia, albeit at low levels.

Distantiae transmission of Trypanosoma cruzi: a new epidemiological feature of acute Chagas disease in Brazil.

BACKGROUND: The new epidemiological scenario of orally transmitted Chagas disease that has emerged in Brazil, and mainly in the Amazon region, needs to be addressed with a new and systematic focus. Belém, the capital of Pará state, reports the highest number of acute Chagas disease (ACD) cases associated with the consumption of açaí juice. METHODOLOGY/PRINCIPAL FINDINGS: The wild and domestic enzootic transmission cycles of Trypanosoma cruzi were evaluated in the two locations (Jurunas and Val-de Cães) that report the majority of the autochthonous cases of ACD in Belém city. Moreover, we evaluated the enzootic cycle on the three islands that provide most of the açaí fruit that is consumed in these localities. We employed parasitological and serological tests throughout to evaluate infectivity competence and exposure to T. cruzi. In Val-de-Cães, no wild mammal presented positive parasitological tests, and 56% seroprevalence was observed, with low serological titers. Three of 14 triatomines were found to be infected (TcI). This unexpected epidemiological picture does not explain the high number of autochthonous ACD cases. In Jurunas, the cases of ACD could not be autochthonous because of the absence of any enzootic cycle of T. cruzi. In contrast, in the 3 island areas from which the açaí fruit originates, 66.7% of wild mammals and two dogs displayed positive hemocultures, and 15.6% of triatomines were found to be infected by T. cruzi. Genotyping by mini-exon gene and PCR-RFLP (1f8/Akw21I) targeting revealed that the mammals and triatomines from the islands harbored TcI and Trypanosoma rangeli in single and mixed infections. CONCLUSION/SIGNIFICANCE: These findings show that cases of Chagas disease in the urban area of Belém may be derived from infected triatomines coming together with the açaí fruits from distant islands. We term this new epidemiological feature of Chagas disease as "Distantiae transmission".

Ecological study of hantavirus infection in wild rodents in an endemic area in Brazil.

A 3-year ecological study of small mammals was carried out in an endemic area for hantavirus pulmonary syndrome in the state of Santa Catarina in Southern Brazil. A total of 994 rodents of 14 different species corresponding to the subfamilies of Sigmodontinae, Murinae, Eumysopinae, and Caviinae were captured during 2004-2006. Oligoryzomys nigripes and Akodon montensis were the most abundant species and showed a clear seasonal pattern with higher population sizes during the winter. Rodent population outbreaks, associated within bamboo mast seeding events, were detected predominantly in areas where hantavirus pulmonary syndrome cases were notified in the state. Antibody reactivity to Hantavirus was detected in five sigmodontine species: O. nigripes (39/435), A. montensis (15/318), Akodon paranaensis (4/37), Thaptomys nigrita (1/86) and Sooretamys angouya (1/12). The highest hantavirus antibody prevalence occurred during the period of highest population size in A. montensis. For O. nigripes, hantavirus prevalence was higher in late spring, when reproduction was more frequent. Co-circulation of Juquitiba (JUQV) and Jabora (JABV) viruses was observed - JABV in A. paranaensis and A. montensis; JUQV in O. nigripes and T. nigrita. JABV occurrence was associated to gender and population size of the rodent while JUQV was related to gender, season, temperature, and locality.

Biochemical and histological changes in liver of Nectomys squamipes naturally infected by Schistosoma mansoni.

The South American water rat Nectomys squamipes is a wild mammal reservoir of Schistosoma mansoni in Brazil. In the present study, wild rodents were collected in the field and categorized into two groups: infected and uninfected by S. mansoni. Blood was collected to analyze changes in the serum glucose level (mg/dL) and liver fragments were used to determine the hepatic glycogen content (mg of glucose/g tissue). The histological examination showed inflammatory granulomatous lesions in different phases of development in the liver of rodents naturally infected with S. mansoni, in some cases with total or partial occlusion of the vascular lumen. Early lesions were characterized by the presence of inflammatory infiltrate around morphologically intact recently deposited eggs. Despite the significance of these histological lesions, the biochemical changes differed in extent. N. squamipes naturally infected by S. mansoni showed no variation in hepatic glycogen reserves. These findings were accompanied by a significant increase in plasma glucose contents, probably as a consequence of amino acids deamination, which are degraded, resulting in the formation of intermediates used as precursors for the glucose formation, without compromising the reserves of liver glycogen. In the wild, naturally infected N. squamipes can maintain S. mansoni infections without undergoing alterations in its carbohydrate metabolism, which minimizes the deleterious effects of S. mansoni.

Evaluation of immunological, parasitological and molecular methods for the diagnosis of Schistosoma mansoni infection before and after chemotherapy treatment with praziquantel in experimentally infected Nectomys squamipes.

In low endemicity areas of schistosomiasis, the recommended diagnostic method of coprological examination results in an underestimation of infection cases. Alternative diagnostic methods have been developed, such as immunodiagnostic and molecular techniques. In this study we evaluated three methods used in the diagnosis of Schistosoma mansoni infection: parasitological (Kato-Katz), immunological (ELISA) and molecular (real time PCR), and also investigated the sensitivity of each technique in the cure determination after treatment with praziquantel using the water rat Nectomys squamipes, a natural reservoir for S. mansoni, as an experimental model. Two infection laboratory experiments were carried out. The first experiment aimed to observe the evolution of the immunological response in the first moments after infection and in the first months after treatment. The second experiment aimed to compare the efficacy of the three diagnostic techniques after infection and after treatment over a more extended time period. In the first experiment, 44% of the infected animals showed IgG reactivity after two weeks of infection, and 94% were positive based on serology 30 days after infection. The serological IgG titers increased just after infection but decreased gradually after treatment. In the second experiment, 89% of the animals showed positive IgG titers 22 days after infection. Only 68% of the animals showed positive results on the coproscopic diagnostic analysis and 79% did so by qPCR, 50 days after infection. Treated animals showed negative results on coproscopy one month after treatment but remained positive by serology even 12 months after treatment, although showing a decline in immunologic reaction after treatment. By qPCR analysis, all animals showed negative results three months after treatment, except for one animal. The parasitosis can be detected by coproscopy only six weeks after infection, and by serology 14 days after infection. The qPCR was a better diagnostic method for confirming the infection cure of S. mansoni. In early infection, this method was less efficient than serology but was slightly more efficient than the Kato-Katz method. We suggest that the methods should be used in low endemic areas as follows: serology should be used in the initial diagnosis in a population with potential positive cases; subsequently, coproscopy should be used in IgG positive cases to confirm the current infection; and qPCR should be used to evaluate the infection cure after treatment and is also a very valuable tool when there are cases showing positive IgG and negative coproscopy.