DNA sequences confirm low specificity to definitive host and wide distribution of the cat pathogen Platynosomum illiciens (= P. fastosum) (Trematoda: Dicrocoeliidae).

Although feline platynosomosis has been commonly reported in several parts of the world, the taxonomy and epidemiological chain related to cat liver flukes remain controversial. In this study, nuclear ribosomal ITS, 28S, and mitochondrial cox1 sequences obtained for Platynosomum illiciens from cat, marmoset, lizard, and snail found naturally infected in Brazil reveal no significant molecular differences between these isolates. Moreover, sequence data confirm that Brazilian P. illiciens from different hosts is conspecific with parasites obtained from cats in Vietnam, supporting wide distribution of the species. The lack of pronounced specificity of P. illiciens to definitive hosts is confirmed here for the first time using molecular approach. The results are discussed in context of the epizootology of platynosomosis.

New insights into the life cycle of Platynosomum (Trematoda: Dicrocoeliidae).

The platynosomiasis, a worldwide parasitic disease with importance for domestic cat, has an etiological agent species of trematodes of the genus Platynosomum, whose complete life cycles are not yet known. The real role of lizards in the transmission of this dicrocoeliid parasite (as obligatory intermediate or paratenic host) still needs to be defined. In the present study, oval-shaped encysted metacercariae obtained from terrestrial isopods (Oniscidea sp. and Nagurus nanus) and elongated excysted metacercariae found in biliary ducts and gallbladder of lizards (Hemidactylus mabouia) in Brazil were used for morphological characterization and experimental infection of mice. Adult parasites recovered from bile ducts and liver of mice inoculated orally with metacercariae from both hosts (isopods and lizards) were identified as Platynosomum illiciens (=Platynosomum fastosum), showing that lizards are paratenic (not obligatory) hosts involved in the life cycle of this parasite. Moreover, Subulina octona is reported as the first intermediate host of P. illiciens in South America, and terrestrial isopods are presented here as new natural second intermediate hosts of the parasite. Finally, it is pointed out that high prevalence and intensity of infection of intermediate and paratenic hosts were observed. These findings on the life cycle of P. illiciens are relevant considering that they may indicate possible control measures of platynosomiasis.

Metacercarial infection of wild Nile tilapia (Oreochromis niloticus) from Brazil.

Fingerlings of Oreochromis niloticus collected in an artificial urban lake from Belo Horizonte, Minas Gerais, Brazil, were evaluated for natural infection with trematodes. Morphological taxonomic identification of four fluke species was performed in O. niloticus examined, and the total prevalence of metacercariae was 60.7% (37/61). Centrocestus formosanus, a heterophyid found in the gills, was the species with the highest prevalence and mean intensity of infection (31.1% and 3.42 (1-42), resp.), followed by the diplostomid Austrodiplostomum compactum (29.5% and 1.27 (1-2)) recovered from the eyes. Metacercariae of Drepanocephalus sp. and Ribeiroia sp., both found in the oral cavity of the fish, were verified at low prevalences (8.2% and 1.6%, resp.) and intensities of infection (only one metacercaria of each of these species per fish). These species of trematodes are reported for the first time in O. niloticus from South America. The potential of occurrence of these parasites in tilapia farming and the control strategies are briefly discussed.

A putative new genus of avian schistosome transmitted by Biomphalaria straminea (Gastropoda: Planorbidae) in Brazil, with a discussion on the potential involvement in human cercarial dermatitis.

Human cercarial dermatitis (HCD) caused by avian schistosomes is an emerging health issue in different parts of the world. Nevertheless, parasite diversity, life cycle, and involvement in HCD remain poorly known or neglected in South America. Herein, we reported data obtained during a long-term malacological survey carried out in Pampulha Reservoir, an urban eutrophic waterbody from Brazil between 2009 and 2012. An ocellate brevifurcate cercaria emerged from 55 of 16,235 (0.34%) specimens of Biomphalaria straminea. Samples of the cercariae were subjected to morphological, experimental, and molecular study (analysis of partial sequences of nuclear 28S and mitochondrial cox1 genes). The molecular analysis revealed that the larva corresponds to an avian schistosome; however, it does not correspond to any named genus. A close related isolate was previously reported in Biomphalaria sudanica from Kenya (molecular divergences of 0.54% and 9.62% for 28S and cox1, respectively). The morphology of this cercaria was compared with other avian schistosome larvae from Biomphalaria spp. Attempts to infect experimentally ducks (Cairina moschata) and mice revealed cutaneous manifestations after exposure to cercariae, but adult parasites were not obtained in these hosts. Phylogenetic analysis suggests this parasite is a putative new genus and species of avian schistosome. The potential involvement of the larvae herein described in cases of HCD in Brazil cannot be ruled out. Surprisingly, HCD was not reported in the country so far, which can be related to difficulties in its diagnosis in areas of overlap with human schistosomes.

Acute infection with Platynosomum illiciens (Trematoda: Dicrocoeliidae) as a clinically relevant and potentially fatal disease in Falco sparverius (Aves: Falconidae) in Brazil.

Platynosomum illiciens is a dicrocoeliid trematode from the biliary tract of warm-blooded vertebrates (felines, primates, marsupials, and birds) reported in different parts of the world. Although the veterinary relevance of platynosomosis in mammals, especially in domestic felines, has been increasingly evidenced in the scientific literature, studies involving avian disease caused by P. illiciens are comparatively scarce. In the present study, a female specimen of the American kestrel, Falco sparverius L., found dead, in November 2019, in Brazil, was necropsied. Parietal biliary effusion in the celomatic cavity was observed, suggesting biliary transudation and gallbladder stasis, which possibily preceded the distension and rupture of gallbladder noted during necropsy. In the microscopic analysis of the bile content, small trematodes were found and characterized as immature stages of Platynosomum after the morphological study. Partial sequences of the cox-1 gene enabled the identification of P. illiciens, with 100% similarity with previously sequenced sympatric isolates from mammals. The finding of immature specimens in a ruptured gallbladder strongly suggests a role for the parasite in biliary flow dysfunction, indicating acute platynosomosis as a clinically relevant and potentially fatal condition that has not yet been discussed.

Oral immunization with Salmonella harboring a Sm14-based DNA vaccine does not protect mice against Schistosoma mansoni infection.

The protection against Schistosoma mansoni infection was evaluated in SWISS mice orally vaccinated with an attenuated strain of Salmonella carrying a Sm14-based DNA vaccine. Although this formulation was not able to afford a reduction in the worm burden, a non-antigen-specific decrease in schistosome-induced granulomatous reaction was verified in livers of mice that received Salmonella.

Exploring possibilities for an alternative approach in experimental schistosomiasis mansoni: the peritoneal cavity of mice.

The schistosome oviposition and granuloma constitution in the peritoneal cavity of AKR/J mice were evaluated. Groups of mice intraperitoneally infected with cercariae of Schistosoma mansoni were weekly euthanized during the acute (56 to 84 days post-infection (DPI)) and chronic (147 to 175 DPI) phase of infection. Schistosome developmental stages obtained via peritoneal lavage and perfusion of the portal system were inspected, counted and fixed, and peritoneal granulomata were then processed for histology. The morphological characterization and quantitative analysis of peritoneal schistosome eggs and granulomata were for the first time performed, such as the demonstration of the viability of miracidia obtained there from. Eutopic and ectopic mature schistosomes and normal pattern of worm oviposition were observed in all periods studied. However, the size of schistosome eggs from peritoneal cavity was smaller than observed for eggs laid by female worms from the portal system. The numbers of S. mansoni eggs and/or granulomata recovered from the peritoneal cavity was higher in chronic than acute infection, while the mean diameter of peritoneal chronic granulomata was smaller than for peritoneal acute granulomata. The constitution and evolution of these cellular reactions at histology were similar to that of hepatic granuloma, and peritoneal granulomata were subject to the host immunomodulation. In addition to the standardization of this experimental approach, which allows the obtaining of free schistosomal granulomata from peritoneal cavity of AKR/J mice, the potential use of these granulomata in ex vivo and in vivo studies is discussed.

Platynosomum illiciens (Trematoda: Dicrocoeliidae) in Captive Black-Tufted Marmoset Callithrix penicillata (Primates: Cebidae) from Brazil: A Morphometric Analyses with Taxonomic Comments on Species of Platynosomum from Nonhuman Primates.

The trematodes belonging to the genus Platynosomum are biliary parasites of birds and mammals (domestic and wildlife) in tropical and subtropical areas of the globe. Despite several reports on platynosomosis in captive nonhuman primates, mainly in South America, the taxonomy of species of Platynosomum that infect these hosts remains confused, and it is not clear whether the species found in cats is the same that infects nonhuman primates. Because a detailed morphological study of Platynosomum from nonhuman primates is lacking, in this study we analyzed specimens of Platynosomum recovered from the biliary system of Callithrix penicillata kept in captivity in an animal facility. The helminths were submitted to morphological and morphometric analyses in a light microscope and measurements of 16 morphological traits were taken. A kernel density estimation (KDE) was used to estimate density distributions of the measurements obtained as well as the occurrence of overlap with the ranges of the measurements known to 2 other species of Platynosomum previously described from South American marmosets, Platynosomum amazonensis and Platynosomum marmoseti. A principal component analysis (PCA) was also performed in order to evaluate the position of each of the 3 species in the multivariate gradient of morphometric measurements. The occurrence of a growth gradient was also evaluated by analysis of correlation between the measurements. Besides a great morphological variability, all specimens obtained from marmosets in this study were identified as Platynosomum illiciens (Braun, 1901). In addition, the published ranges of the measurements of P. amazonensis and P. marmoseti were completely contained within the ranges found in this study as revealed by KDE. The PCA did not show the formation of groups, and the 3 species were distributed along a growth continuum, also corroborated by correlation analysis. Therefore, P. amazonensis and P. marmoseti are here synonymized with P. illiciens. The involvement of wildlife hosts to the epidemiology of feline platynosomosis and implications for its control are briefly discussed.

Oral administration of a live Aro attenuated Salmonella vaccine strain expressing 14-kDa Schistosoma mansoni fatty acid-binding protein induced partial protection against experimental schistosomiasis.

We report the oral vaccination of SWISS mice with an Aro attenuated Salmonella enterica var. Typhimurium vaccine strain expressing the 14-kDa Schistosoma mansoni antigen, Sm14. Bacterial adjuvants, including (i) Lactococcus lactis expressing interleukin-12 (IL-12) and (ii) Lactobacillus delbrueckii UFV-H2b20, were also employed in oral immunization assays. Detection assays to specific IgG and IgA anti-Sm14 antibodies were performed to evaluate humoral immune responses in vaccinated mice. An increase in specific IgG titers was observed; however, no IgA production was detected. The protection levels against schistosomiasis (34.9-49.5%) obtained with all experimental formulations in this work were very similar to values reported by previous studies, which used purified recombinant Sm14 for parenteral vaccination of mice. There was a slight reduction in hepatic granulomas of mice vaccinated with Salmonella. Oogram studies showed diminished numbers of S. mansoni eggs in the intestinal wall of vaccinated mice, but individual female worm fecundity did not seem to be affected by our immunization protocol.

Experimental platynosomosis: Characterization of parasite development in the mouse model.

Despite the veterinary importance of species of Platynosomum, biliary trematode parasites of birds and mammals with worldwide distribution and a growing role in feline practice, the basic parasitological aspects of platynosomosis is still not completely understood due to the scarcity of studies in experimental models. In the present study, metacercariae of Platynosomum illiciens obtained from naturally infected tropical house geckos (Hemidactylus mabouia) in an urban area of Brazil were force-fed to mice of the AKR/J strain (100 metacercariae/animal). Groups of mice were euthanized at 60, 120, 160 and 240 days post-infection (DPI), and the biliary tree of the animals (intrahepatic biliary ducts, common hepatic and bile ducts, cystic duct and gallbladder) were examined for the presence of adult parasites. Recovered flukes were counted, classified by their site of origin (i.e., intrahepatic or extrahepatic biliary ducts) and morphologically analyzed under light microscope. The number of adult parasites obtained at 60, 120, 160 and 240 DPI was 22 ± 6 (16-32), 41 ± 14 (18-48), 27 ± 11 (18-40) and 20 ± 6 (13-30), respectively, and no significant differences in total worm burden at the different experimental times were observed. However, 41%, 51%, 75% and 95% of the parasites were found in the common hepatic and bile ducts at 60, 120, 160 and 240 DPI, respectively, suggesting the occurrence of parasitic migration to the extrahepatic biliary tree during infection; however, no parasites were observed in the gallbladder or cystic duct. Regarding the morphometric analysis, progressive growth of P. illiciens during the experimental time was observed, and the parasites collected from the extrahepatic bile ducts were larger than those obtained from the intrahepatic ducts at the same time of infection. Parasites obtained from the extrahepatic biliary tree of the mice at 160 DPI had similar measurements to those of parasites obtained at 240 DPI, and those measurements were equivalent to those reported for parasites from natural hosts (cats, birds and nonhuman primates). The results obtained provide new insights into the biology of P. illiciens, and the kinetics of the parasite development of this species is presented here for the first time. The potential use of mice as an experimental model for P. illiciens is presented and the implications of the results obtained in that model for feline platynosomosis are briefly discussed.

Schistosoma mansoni tegument protein Sm29 is able to induce a Th1-type of immune response and protection against parasite infection.

BACKGROUND: Schistosomiasis continues to be a significant public health problem. This disease affects 200 million people worldwide and almost 800 million people are at risk of acquiring the infection. Although vaccine development against this disease has experienced more failures than successes, encouraging results have recently been obtained using membrane-spanning protein antigens from the tegument of Schistosoma mansoni. Our group recently identified Sm29, another antigen that is present at the adult worm tegument surface. In this study, we investigated murine cellular immune responses to recombinant (r) Sm29 and tested this protein as a vaccine candidate. METHODS AND FINDINGS: We first show that Sm29 is located on the surface of adult worms and lung-stage schistosomula through confocal microscopy. Next, immunization of mice with rSm29 engendered 51%, 60% and 50% reduction in adult worm burdens, in intestinal eggs and in liver granuloma counts, respectively (p<0.05). Protective immunity in mice was associated with high titers of specific anti-Sm29 IgG1 and IgG2a and elevated production of IFN-gamma, TNF-alpha and IL-12, a typical Th1 response. Gene expression analysis of worms recovered from rSm29 vaccinated mice relative to worms from control mice revealed a significant (q<0.01) down-regulation of 495 genes and up-regulation of only 22 genes. Among down-regulated genes, many of them encode surface antigens and proteins associated with immune signals, suggesting that under immune attack schistosomes reduce the expression of critical surface proteins. CONCLUSION: This study demonstrates that Sm29 surface protein is a new vaccine candidate against schistosomiasis and suggests that Sm29 vaccination associated with other protective critical surface antigens is the next logical strategy for improving protection.