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15 pages, 3796 KiB  
Article
Coprological Survey of Helminths in Reindeer (Rangifer tarandus) in 50 Selected Zoos and Menageries in Russia
by Olga A. Loginova, Svetlana V. Akulova, Dmitry N. Egorov, Natalia S. Egorova, Sergei E. Spiridonov, Iuliia K. Plotnikova, Larisa M. Belova, Yuriy E. Kuznetsov, Daria I. Chuprak, Anna A. Krutikova, Iuliia V. Vasilkova, Diana A. Gelashvili, Yuri A. Shchepanovsky, Ivan A. Mizin, Danila V. Panchenko, Mikhail G. Bondar and Taras P. Sipko
J. Zool. Bot. Gard. 2024, 5(3), 492-506; https://doi.org/10.3390/jzbg5030033 (registering DOI) - 12 Aug 2024
Abstract
Zoo conditions are unique for reindeer, since domestic reindeer are not kept in captive facilities like cattle. In the zoo, reindeer are usually surrounded by many different animals that they would never encounter naturally. Thus, they might be infected with new helminths. Numerous [...] Read more.
Zoo conditions are unique for reindeer, since domestic reindeer are not kept in captive facilities like cattle. In the zoo, reindeer are usually surrounded by many different animals that they would never encounter naturally. Thus, they might be infected with new helminths. Numerous petting zoos raise concerns about the safety of tactile interactions for human visitors. Our study is the first large-scale one. Qualitative and quantitative fecal analyses were carried out for 233 reindeer distributed over 50 Russian zoos according to the National Standard of the Russian Federation (GOST R 54627-2011) Ruminant animals—Methods of Laboratory Helminthological Diagnostics. Where possible, DNA analyses of helminths were performed targeting internal transcribed spacer region. As a result, F. hepatica, Paramphistomum sp., Moniezia sp. (including M. expansa), gastrointestinal strongylids (including Nematodirus spp.), Dictyocaulus sp., E. rangiferi, Trichuris sp., and Capillaria sp. were found in 106 (45%) zoo reindeer. All these helminths were previously reported in reindeer and pose no direct danger for humans. The intensity of invasions was mostly low. Fecal examination might be considered as an indirect method for mange diagnostics, as Chorioptes and Demodex mites were found in reindeer fecal samples. The latter may represent a novel species of mite specific for reindeer. Full article
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Graphical abstract

Graphical abstract
Full article ">Figure 1
<p>Diagnostic stages of helminths obtained from feces of zoo reindeer. (<b>A</b>) <span class="html-italic">Fasciola hepatica</span> egg; (<b>B</b>) <span class="html-italic">Moniezia expansa</span> egg; (<b>C</b>) <span class="html-italic">Nematodirus</span> sp. egg at late stage of embryonic development; (<b>D</b>) rounded strongyle-type egg; (<b>E</b>) <span class="html-italic">Trichuris</span> sp. egg; (<b>F</b>) <span class="html-italic">Paramphistomum</span> sp. egg; (<b>G</b>) <span class="html-italic">Moniezia</span> sp. egg; (<b>H</b>) <span class="html-italic">Nematodirus</span> sp. egg at early stage of embryonic development; (<b>I</b>) elongated strongyle-type egg; (<b>J</b>) <span class="html-italic">Capillaria</span> sp. egg; (<b>K</b>) <span class="html-italic">Dictyocaulus</span> sp. L1; (<b>L</b>) <span class="html-italic">Elaphostrongylus rangiferi</span> L1. Bright field microscopy, 400× magnification. Scale bar equals 50 μm.</p>
Full article ">Figure 2
<p>Prevalence rates (%) of helminths found in feces of zoo reindeer.</p>
Full article ">Figure A1
<p>Map of Russia indicating the zoos. Insert on the right represents magnified area marked with a rectangle. Numbers correspond to the zoo ID numbers in <a href="#jzbg-05-00033-t0A1" class="html-table">Table A1</a>.</p>
Full article ">Figure A2
<p>Mites of reindeer. (<b>A</b>) <span class="html-italic">Demodex</span> sp. obtained from the feces of a reindeer from zoo #12; (<b>B</b>) egg of mite at early developmental stage obtained from the feces of a reindeer from zoo #30; (<b>C</b>) egg of mite at late developmental stage obtained from the feces of a reindeer from zoo #48; (<b>D</b>) <span class="html-italic">Chorioptes</span> sp. obtained from the feces of a reindeer from zoo #40; (<b>E</b>) manifestation of mange caused by mites in a reindeer from zoo #30 (furless patches in the front leg and chest are shown); photo courtesy: Kristina Zabarina. Light micrographs were made via bright field microscopy, 400× magnification. Scale bar equals 50 μm.</p>
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14 pages, 3047 KiB  
Article
Investigation of a Serine Protease Inhibitor Active in the Infectious Stage of the Human Liver Fluke Opisthorchis viverrini
by Rosnanee Salang, Wansika Phadungsil, Amornrat Geadkaew-Krenc and Rudi Grams
Pathogens 2024, 13(8), 678; https://doi.org/10.3390/pathogens13080678 - 10 Aug 2024
Viewed by 268
Abstract
Serine protease inhibitors (serpins) participate in the regulation of inflammation, blood coagulation, and complement activation in humans. This research aimed to identify and characterize such inhibitors of the human liver fluke Opisthorchis viverrini. Parasite proteins that might contribute to the modulation of [...] Read more.
Serine protease inhibitors (serpins) participate in the regulation of inflammation, blood coagulation, and complement activation in humans. This research aimed to identify and characterize such inhibitors of the human liver fluke Opisthorchis viverrini. Parasite proteins that might contribute to the modulation of host physiology are of particular interest, especially as chronic opisthorchiasis increases the risk of developing biliary cancer. BLAST was used to find hypothetical serpins predicted from the parasite genome data. RNA extraction and reverse transcriptase PCR were used to isolate a serpin cDNA and to determine developmental transcript abundance. The evolutionary relation to other trematode serpins was revealed by phylogenetic analysis. Recombinant serpin was expressed in Escherichia coli and used to test the immunoreactivity of human opisthorchiasis sera and the inhibition of human serine proteases. A substantial serpin family with high sequence divergence among the members was found in the genus Opisthorchis. A serpin, different from previously analyzed trematode serpins, was cloned. The transcript was only detected in metacercariae and newly excysted juveniles. Human opisthorchiasis sera showed statistically significant reactivity to recombinant serpin. The serpin caused moderate inhibition of thrombin and low inhibition of kallikrein and chymotrypsin. This parasite serpin could be further evaluated as a diagnostic tool for early infection. Kallikrein and thrombin are involved in fibrinolysis; therefore, further research should explore the effects of the parasite serpin on this process. Full article
(This article belongs to the Section Parasitic Pathogens)
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Graphical abstract

Graphical abstract
Full article ">Figure 1
<p>Multiple sequence alignment of <span class="html-italic">S. haematobium</span> SPI (UniProt: Q26502), <span class="html-italic">O. viverrini</span> SIS (UniProt: A0A075AEP2), <span class="html-italic">C. sinensis</span> SIS (UniProt: A0A419PW56), and <span class="html-italic">O. felineus</span> SIS (UniProt: A0A4S2MGE0). The α-helical and β-strand regions of <span class="html-italic">Sh</span>SPI and <span class="html-italic">Ov</span>SIS based on their AlphaFold native state models [<a href="#B23-pathogens-13-00678" class="html-bibr">23</a>] are indicated at the top and bottom, respectively. The numbering of the secondary structures follows α<sub>1</sub>-antitrypsin [<a href="#B24-pathogens-13-00678" class="html-bibr">24</a>]. The region of the reactive center loop (RCL) with the P1 residue is indicated by a gray bar. Strand 4A in the RCL was manually added as it forms and integrates into β-sheet A after cleavage at P1. Sequence conservation using the BLOSUM 62 similarity matrix and <span class="html-italic">Sh</span>SPI as the reference sequence is indicated by color shading (fully conserved: purple background, ≥50% conserved: blue background, and similar: red background). The graphical representation of the aligned sequences was created using TEXshade v1.28 [<a href="#B25-pathogens-13-00678" class="html-bibr">25</a>].</p>
Full article ">Figure 2
<p><span class="html-italic">Ov</span>SIS structure model predicted in AlphaFold [<a href="#B23-pathogens-13-00678" class="html-bibr">23</a>] and obtained from the AlphaFold Protein Structure Database [<a href="#B26-pathogens-13-00678" class="html-bibr">26</a>]. The <span class="html-italic">Ov</span>SIS model with the three β-sheets A (red), B (green), and C (yellow), RCL (dark blue), and threonine 365 in the P1 position is shown on the left. The <span class="html-italic">Ov</span>SIS model prediction confidence from high to low is indicated by red to blue shading, respectively. Overlays with <span class="html-italic">Sh</span>SPI (PDB: 3STO) and <span class="html-italic">Sm</span>SPI (PDB: 6SSV) demonstrate structural conservation between the three serpins. Graphical representations were created using ChimeraX [<a href="#B27-pathogens-13-00678" class="html-bibr">27</a>].</p>
Full article ">Figure 3
<p>(<b>a</b>) Phylogenetic tree of trematode serpins specified by their UniProt accession numbers. The three previously described serpins of <span class="html-italic">C. sinensis</span> are indicated with a trailing •. Schistosome serpins are shown in blue, with the SPI in bold lettering. <span class="html-italic">Ov</span>SIS and its orthologs are shown in red. <span class="html-italic">Opisthorchis</span>/<span class="html-italic">Clonorchis</span> SIS is not orthologous to schistosome SIP. (<b>b</b>) Sequence conservation of the reactive center loop in evolutionarily close serpins (specified by their UniProt accession numbers) in the species <span class="html-italic">O. viverrini</span> (Ov), <span class="html-italic">O. felineus</span> (Of), <span class="html-italic">C. sinensis</span> (Cs), <span class="html-italic">S. haematobium</span> (Sh), <span class="html-italic">S. japonicum</span> (Sj), and <span class="html-italic">S. mansoni</span> (Sm). The P1 position in the protease interaction is indicated. The color shading indicates fully conserved: purple background, ≥50% conserved: blue background, and similar: red background.</p>
Full article ">Figure 4
<p>(<b>a</b>) RT-PCR products resolved in 0.8% agarose gels. M: marker, 1: newly excysted juveniles, 2: 2-week-old juveniles, and 3: mature parasites. <span class="html-italic">O. viverrini</span> β-actin (292 bp) was used as a positive control. A 640 bp <span class="html-italic">Ov</span>SIS cDNA was only produced with RNA of newly excysted juveniles. (<b>b</b>) Micrograph of a newly excysted <span class="html-italic">O. viverrini</span> juvenile (top) and a metacercaria (bottom), the developmental stages in which <span class="html-italic">Ov</span>SIS is present. The dark granule-filled excretory bladder, which can also be seen through the cyst wall, is the most prominent. Other outstanding morphological features are the oral and ventral suckers. It is thought that parasite-secreted serine proteases participate in the release of the encysted juveniles and that cognate serpins regulate their excess activity.</p>
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<p>(<b>a</b>) SDS-PAGE showing pre-induced (1) and IPTG-induced bacterial proteins (2) and affinity-purified recombinant <span class="html-italic">Ov</span>SIS (3). The protein migrates at the expected 46 kDa molecular weight. (<b>b</b>) Absorption values of two groups of human sera against recombinant <span class="html-italic">Ov</span>SIS obtained by indirect ELISA. n: normal sera (<span class="html-italic">n</span> = 10). i: sera of <span class="html-italic">O. viverrini</span>-infected individuals (<span class="html-italic">n</span> = 30). The minimum, 25th percentile, median, 75th percentile, and maximum are indicated in the box plots. The star symbols indicate that all infected sera were statistically significantly different (*** <span class="html-italic">p</span> &lt; 0.05) from the normal sera. (<b>c</b>) Human infection with <span class="html-italic">O. viverrini</span>. The metacercariae are released from infested undercooked fish in the stomach. The acidic environment activates the excystment of the juveniles. The newly excysted juveniles are released with the stomach content into the duodenum and migrate via the sphincter of Oddi into the biliary system (green).</p>
Full article ">Figure 6
<p>Inhibition kinetics of <span class="html-italic">Ov</span>SIS against human serine proteases. Proteases without inhibitors are indicated in black, proteases incubated with recombinant <span class="html-italic">Ov</span>SIS are shown in red, and proteases incubated with 20 mM PMSF are shown in teal. After 30 min of preincubation with 0.5 µM recombinant <span class="html-italic">Ov</span>SIS and 30 min of incubation with the substrate, the activities of 1 nM chymotrypsin, 1 nM kallikrein, and 0.1 U thrombin were lowered to 69.21, 66.76, and 48.84%, respectively. The enzyme substrates are listed in <a href="#pathogens-13-00678-t001" class="html-table">Table 1</a>.</p>
Full article ">
27 pages, 5317 KiB  
Article
ARGONAUTE2 Localizes to Sites of Sporocysts in the Schistosome-Infected Snail, Biomphalaria glabrata
by Phong Phan, Conor E. Fogarty, Andrew L. Eamens, Mary G. Duke, Donald P. McManus, Tianfang Wang and Scott F. Cummins
Genes 2024, 15(8), 1023; https://doi.org/10.3390/genes15081023 - 3 Aug 2024
Viewed by 671
Abstract
MicroRNAs (miRNAs) are a class of small regulatory RNA that are generated via core protein machinery. The miRNAs direct gene-silencing mechanisms to mediate an essential role in gene expression regulation. In mollusks, miRNAs have been demonstrated to be required to regulate gene expression [...] Read more.
MicroRNAs (miRNAs) are a class of small regulatory RNA that are generated via core protein machinery. The miRNAs direct gene-silencing mechanisms to mediate an essential role in gene expression regulation. In mollusks, miRNAs have been demonstrated to be required to regulate gene expression in various biological processes, including normal development, immune responses, reproduction, and stress adaptation. In this study, we aimed to establishment the requirement of the miRNA pathway as part of the molecular response of exposure of Biomphalaria glabrata (snail host) to Schistosoma mansoni (trematode parasite). Initially, the core pieces of miRNA pathway protein machinery, i.e., Drosha, DGCR8, Exportin-5, Ran, and Dicer, together with the central RNA-induced silencing complex (RISC) effector protein Argonaute2 (Ago2) were elucidated from the B. glabrata genome. Following exposure of B. glabrata to S. mansoni miracidia, we identified significant expression up-regulation of all identified pieces of miRNA pathway protein machinery, except for Exportin-5, at 16 h post exposure. For Ago2, we went on to show that the Bgl-Ago2 protein was localized to regions surrounding the sporocysts in the digestive gland of infected snails 20 days post parasite exposure. In addition to documenting elevated miRNA pathway protein machinery expression at the early post-exposure time point, a total of 13 known B. glabrata miRNAs were significantly differentially expressed. Of these thirteen B. glabrata miRNAs responsive to S. mansoni miracidia exposure, five were significantly reduced in their abundance, and correspondingly, these five miRNAs were determined to putatively target six genes with significantly elevated expression and that have been previously associated with immune responses in other animal species, including humans. In conclusion, this study demonstrates the central importance of a functional miRNA pathway in snails, which potentially forms a critical component of the immune response of snails to parasite exposure. Further, the data reported in this study provide additional evidence of the complexity of the molecular response of B. glabrata to S. mansoni infection: a molecular response that could be targeted in the future to overcome parasite infection and, in turn, human schistosomiasis. Full article
(This article belongs to the Special Issue Evolution of Non-coding Elements in Genome Biology)
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Figure 1

Figure 1
<p>Identification and expression analysis of core pieces of protein machinery of the <span class="html-italic">Biomphalaria glabrata</span> miRNA pathway 16 and 42 h after its exposure to <span class="html-italic">Schistosoma mansoni</span> miracidia. (<b>A</b>) Schematic of the production (nucleus; grey-colored shaded region) and action (cytoplasm; pale-yellow-colored shaded region) stages of the <span class="html-italic">B. glabrata</span> miRNA pathway with the functional position within the pathway of <span class="html-italic">Bgl</span>-Drosha, <span class="html-italic">Bgl</span>-DGCR8, <span class="html-italic">Bgl</span>-Exp5, <span class="html-italic">Bgl</span>-Ran, <span class="html-italic">Bgl</span>-Dcr, and <span class="html-italic">Bgl</span>-Ago2 indicated. (<b>B</b>) Schematic outlining the functional domain structure of the core pieces of protein machinery of the <span class="html-italic">B. glabrata</span> miRNA pathway, including <span class="html-italic">Bgl</span>-Drosha, <span class="html-italic">Bgl</span>-DGCR8, <span class="html-italic">Bgl</span>-Exp5, <span class="html-italic">Bgl</span>-Ran, <span class="html-italic">Bgl</span>-Dcr, and <span class="html-italic">Bgl</span>-Ago2. RIBOc, ribonuclease III C terminal domain; DSRM, double-stranded RNA motif; OBD/WW, origin-binding domain/tryptophan-tryptophan domain; XPO, Exportin domain; IBN_N, Importin-β N-terminal domain; Ran, Ras-related nuclear domain; Dcr/Dcr, Dicer dimerization domain, HELICc, helicase superfamily C-terminal domain; PAZ, PIWI, Argonaute, and Zwille domain; N, Ago protein amino-terminal region domain; DUF1785, domain of unknown function 1785 domain; PIWI, P-element-induced wimpy testis domain; L2, linker region 2; MID, middle domain. (<b>C</b>) RNA-Seq assessment of the altered expression of <span class="html-italic">Bgl-Drosha</span>, <span class="html-italic">Bgl-DGCR8</span>, <span class="html-italic">Bgl-Exp5</span>, <span class="html-italic">Bgl-Ran</span>, <span class="html-italic">Bgl-Dcr</span>, and <span class="html-italic">Bgl-Ago2</span>, 16 and 42 h after exposure of <span class="html-italic">B. glabrata</span> animals to the <span class="html-italic">S. mansoni</span> parasite. Error bars represent the standard error of the mean, and an asterisk (*) denotes significantly altered transcript abundance (<span class="html-italic">p</span>-value ≤ 0.05) at either the 16 or 42 h time point compared to the level of expression of each core piece of protein machinery in control (unexposed) <span class="html-italic">B. glabrata</span> animals.</p>
Full article ">Figure 2
<p>Expression analysis and tissue localization of the Ago2 protein in <span class="html-italic">Biomphalaria glabrata</span> animals following their short- and long-term exposure to the <span class="html-italic">Schistosoma mansoni</span> parasite. (<b>A</b>) Western blot hybridization analysis of Ago2 protein abundance in whole-protein extracts from unexposed (control) and exposed <span class="html-italic">B. glabrata</span> whole animals and <span class="html-italic">S. mansoni</span> miracidia. (<b>B</b>) Western blot hybridization analysis of Ago2 protein abundance in unexposed <span class="html-italic">B. glabrata</span> whole animals (control) and in exposed animals 16 and 42 h after their exposure to <span class="html-italic">S. mansoni</span> miracidia, an analysis that was performed in triplicate for quantification (<b>C</b>) to definitively demonstrate significantly altered Ago2 protein abundance in <span class="html-italic">S. mansoni</span>-exposed <span class="html-italic">B. glabrata</span> whole animals. An asterisk (*) denotes significantly altered Ago2 protein abundance (<span class="html-italic">p</span>-value ≤ 0.05) at either the 16 or 42 h time point compared to the level of Ago2 protein in control (unexposed) <span class="html-italic">B. glabrata</span> animals. (<b>D</b>) Light-field and fluorescent microscopic analysis of the intestinal digestive gland of sectioned <span class="html-italic">B. glabrata</span> animals 16 h post exposure to <span class="html-italic">S. mansoni</span> miracidia. Specifically, (<b>Di</b>,<b>Dii</b>) show different magnifications of bright-field microscopic analysis of H&amp;E-stained regions of the <span class="html-italic">B. glabrate</span> intestinal digestive gland. No fluorescence was observed in the intestinal digestive gland of sectioned <span class="html-italic">B. glabrata</span> animals 16 h post exposure to <span class="html-italic">S. mansoni</span> miracidia (<b>Diii</b>,<b>Div</b>), nor were fluorescent signals observed in negative control samples (<b>Dv</b>,<b>Dvi</b>). (<b>E</b>) Light-field microscopic analysis and fluorescent microscopic analysis of the intestinal digestive gland of sectioned <span class="html-italic">B. glabrata</span> animals 20 days post exposure to the <span class="html-italic">S. mansoni</span> parasite. Specifically, (<b>Ei</b>,<b>Eii</b>) show different magnifications of bright-field microscopic analysis of H&amp;E-stained regions of the intestinal digestive gland of sectioned <span class="html-italic">B. glabrate</span> animals, with the black arrows indicating sporocysts. Readily observable fluorescence was observed in the intestinal digestive gland of sectioned <span class="html-italic">B. glabrata</span> animals (yellow arrows), specifically around the sporocysts that had formed at this long-term exposure time point (<b>Eiii</b>,<b>Eiv</b>). As shown in (<b>Dv</b>,<b>Dvi</b>), no fluorescence was observed in the intestinal digestive gland sectioned of <span class="html-italic">B. glabrata</span> animals 20 days post exposure to <span class="html-italic">S. mansoni</span> miracidia in the negative control samples (<b>Eiv</b>,<b>Ev</b>).</p>
Full article ">Figure 3
<p>Profiling of the microRNA landscape of <span class="html-italic">Biomphalaria glabrata</span> whole animals 16 h after their exposure to <span class="html-italic">Schistosoma mansoni</span> miracidia. (<b>A</b>) Heat map of the 66 miRNAs with altered abundance (up- (orange) or down-regulated (blue) abundance) 16 h post exposure of <span class="html-italic">B. glabrata</span> to <span class="html-italic">S. mansoni</span> miracidia, with the intensity of shading of each tile indicating the degree of change to miRNA abundance. (<b>B</b>) Elevated (n = 8) or reduced (n = 5) levels of the 13 miRNAs with significantly altered abundance 16 h post exposure of <span class="html-italic">B. glabrata</span> to <span class="html-italic">S. mansoni</span> miracidia, with orange-colored columns showing up-regulated miRNAs and blue-colored columns representing down-regulated miRNAs. (<b>C</b>) Schematic demonstrating that miRNA abundance was altered in <span class="html-italic">B. glabrata</span> whole animals after their exposure to <span class="html-italic">S. mansoni</span> miracidia regardless of the genomic context of their encoding gene, with altered miRNAs originating from <span class="html-italic">MIR</span> gene clusters or positioned at isolated <span class="html-italic">MIR</span> gene loci in both intragenic and intergenic genomic contexts. (<b>D</b>) Pie chart outlining the genomic context of <span class="html-italic">MIR</span> genes from which the 66 miRNAs with altered abundance 16 h post the exposure of <span class="html-italic">B. glabrata</span> whole animals to <span class="html-italic">S. mansoni</span> miracidia are derived.</p>
Full article ">Figure 4
<p>Interaction map of the putative target genes of the five <span class="html-italic">Biomphalaria glabrata</span> miRNAs with significantly reduced abundance 16 h post exposure to <span class="html-italic">Schistosoma mansoni</span> miracidia. (<b>A</b>) Venn diagram showing the number of predicted target genes for the five significantly reduced <span class="html-italic">B. glabrata</span> miRNAs 16 h post exposure to <span class="html-italic">S. mansoni</span> miracidia following target gene assessment using the miRanda and RNAhybrid prediction tools, respectively. (<b>B</b>) miRNA/target gene interaction map for the five significantly reduced <span class="html-italic">B. glabrata</span> miRNAs (blue blocks) post exposure to <span class="html-italic">S. mansoni</span> miracidia, including putative target genes both with down-regulated (yellow blocks) and up-regulated (red blocks) levels of expression.</p>
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20 pages, 109182 KiB  
Article
In Vivo Antischistosomicidal and Immunomodulatory Effects of Dietary Supplementation with Taraxacum officinale
by Amany Ebrahim Nofal, Amal Mohamed Shaaban, Hany Mohammed Ibrahim, Faten Abouelmagd and Azza Hassan Mohamed
J. Xenobiot. 2024, 14(3), 1003-1022; https://doi.org/10.3390/jox14030056 - 29 Jul 2024
Viewed by 290
Abstract
Bilharziasis is a widespread trematode parasite that poses a severe public health burden. Dandelion (Taraxacum officinale) has several pharmacological and traditional properties critical for treating several hepatic disorders. The present study was designed to assess the potential efficacy of T. officinale [...] Read more.
Bilharziasis is a widespread trematode parasite that poses a severe public health burden. Dandelion (Taraxacum officinale) has several pharmacological and traditional properties critical for treating several hepatic disorders. The present study was designed to assess the potential efficacy of T. officinale root (TOR) dietary supplementation with or without praziquantel (PZQ) against liver and intestinal disorders in mice infected with Schistosoma mansoni. This study was conducted on five groups; G1: uninfected control, G2: untreated S. mansoni-infected mice, G3: infected animals treated with 250 mg/kg PZQ for three alternative days, G4: infected animals were orally administered 600 mg/kg bw TOR daily for 10 days, and G5: infected animals that received both PZQ and TOR as previously described. The current findings after different treatments indicated topographical scanning electron microscopy alterations of male adult worms and a critical reduction in worm burden, ova count, granuloma diameter, hepatic and intestinal histological abnormalities, fibrosis, immunohistochemical expression of CD3+ and CD20+ cells, oxidative stress, and interleukin-10, also upregulation of interferon-gamma, and antioxidant enzymes, when compared to the infected untreated mice. The best results were obtained in mice administered PZQ+TOR together because of their antioxidant properties and ability to promote the host immune response to parasitic infection. Full article
(This article belongs to the Section Natural Products/Herbal Medicines)
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Figure 1

Figure 1
<p>Schematic diagram of the study design.</p>
Full article ">Figure 2
<p>Representative scanning electron micrographs of the dorsal surface of adult male <span class="html-italic">S. mansoni</span> worms of the different groups (n = 3). (<b>A</b>,<b>B</b>): the worm from an infected untreated mouse has a rough dorsal tegument with numerous large tubercles (TU) bearing sharp spines (S) with small tegumental blebs (Tb) and tegumental ridges between them (R). (<b>C</b>,<b>D</b>): worm from the infected group treated with PZQ alone exhibiting worms with marked destruction of the tegument, erosion of the sloughing surface, swollen tubercles (TU) with a proximal loss of spines, extensive ulceration of the outer surface (black arrows), and loss of intertubercular tegumental ridges (stars). (<b>E</b>,<b>F</b>): the worm from an infected mouse supplemented with TOR alone showing distortion of the tubercles with loss of spines and wrinkling in the areas between them (yellow arrows). (<b>G</b>,<b>H</b>): the worm from PZQ+TOR-treated group exhibiting tegumental wrinkles (yellow arrows), distorted tubercles with markedly decreased spines, and many surrounding blebs (red arrows).</p>
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<p>Representative scanning electron micrographs of the anterior region of adult male <span class="html-italic">S. mansoni</span> worms showing oral sucker (OS) and ventral sucker (VS) of the different groups (n = 3). (<b>A</b>–<b>C</b>): the worm from an infected untreated mouse showing normal morphology with oral and ventral suckers, respectively, with numerous sharp spines (S). (<b>D</b>–<b>F</b>) the worm from an infected mouse treated with PZQ exhibiting abnormal oral and ventral suckers with marked disturbances in the direction of the spines and a decrease in the number of spines (arrows). (<b>G</b>–<b>I</b>) the worm from an infected mouse supplemented with TOR displaying damaged oral and ventral suckers that lost their normal structure with marked disturbance in the direction of spines and a marked decrease in spines (arrows). (<b>J</b>–<b>L</b>) the worm from an infected mouse dual treated with both PZQ and TOR showing abnormal appearance of oral and ventral suckers with a marked decrease in the number of spines (arrows).</p>
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<p>Representative photomicrographs from H. &amp; E. mice liver sections of the studied groups (n = 5): uninfected control (<b>A</b>), infected untreated (<b>B</b>,<b>C</b>), PZQ-monotherapy (<b>D</b>), TOR-monotherapy (<b>E</b>), PZQ+TOR-dual therapy (<b>F</b>), demonstrating central vein (CV), hepatocytes (HC), sinusoids (S), cellular infiltration around the granuloma (G) with trapped parasitic ova (thin arrows), fiber deposition (black arrowheads), hypereosinophilic hepatocyte (green arrows), hypertrophied Kupffer cell (blue arrows), necrotic area (N), leucocytic infiltration (LI), portal vein (PV), and bile canaliculi (Bc) (H. &amp; E. stain).</p>
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<p>Representative photomicrographs from Masson trichrome mice liver sections of the studied groups (n = 5): uninfected control group (<b>A</b>), infected untreated group (<b>B</b>,<b>C</b>), PZQ-treated group (<b>D</b>), TOR-treated group (<b>E</b>), PZQ+TOR-dual therapy group (<b>F</b>), demonstrating blue–green collagen fibers distribution (arrows), and central vein (CV) (Masson’s trichrome stain).</p>
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<p>Representative photomicrographs from H. &amp; E. mice small intestinal sections of the studied groups (n = 5): uninfected control (<b>A</b>), infected untreated (<b>B</b>,<b>C</b>), PZQ-monotherapy (<b>D</b>), TOR-monotherapy (<b>E</b>), PZQ+TOR-dual therapy (<b>F</b>), showing that the muscularis (M) and villi (V) extended from the submucosa to the lumen, cellular infiltration around the granuloma (G) with trapped parasitic ova (thin arrows), fiber deposition (black arrowheads), and cellular infiltration around the empty positions devoid of eggs (thick arrow) (H. &amp; E. stain).</p>
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<p>Representative photomicrographs from Masson trichrome mice small intestinal sections of the studied groups (n = 5): uninfected control (<b>A</b>), infected untreated (<b>B</b>,<b>C</b>), PZQ-monotherapy (<b>D</b>), TOR-monotherapy (<b>E</b>), and PZQ+TOR-dual therapy (<b>F</b>), demonstrating blue–green collagen fibers distribution (arrows) (Masson’s trichrome stain).</p>
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<p>Representative photomicrographs of the immunohistochemical hepatic sections (n = 5) of the infected mice without treatment (<b>A</b>), with PZQ monotherapy (<b>B</b>), with TOR monotherapy (<b>C</b>), and with PZQ+TOR dual therapy (<b>D</b>), demonstrating positive brown CD3<sup>+</sup> reactions (arrows).</p>
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<p>Representative photomicrographs of the immunohistochemical hepatic sections (n = 5) of infected mice without treatment (<b>A</b>), with PZQ monotherapy (<b>B</b>), with TOR monotherapy (<b>C</b>), and with PZQ+TOR dual therapy (<b>D</b>), demonstrating positive brown CD20<sup>+</sup> reactions (arrows).</p>
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<p>Representative photomicrographs of the immunohistochemical intestinal sections (n = 5) of infected mice without treatment (<b>A</b>), with PZQ monotherapy (<b>B</b>), with TOR monotherapy (<b>C</b>), and with PZQ+TOR dual therapy (<b>D</b>), demonstrating positive brown CD3<sup>+</sup> reactions (arrows).</p>
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<p>Representative photomicrographs of the immunohistochemical intestinal sections (n = 5) of infected mice without treatment (<b>A</b>), with PZQ monotherapy (<b>B</b>), with TOR monotherapy (<b>C</b>), and with PZQ+TOR dual therapy (<b>D</b>), demonstrating positive brown CD20<sup>+</sup> reactions (arrows).</p>
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<p>Histogram of the CD4<sup>+</sup> and CD8<sup>+</sup> blood phenotypic analysis after different therapies. The data are expressed as the means ± SD of the following groups (n = 3): the uninfected control group (G1), the infected untreated group (G2), the PZQ monotherapy group (G3), the TOR monotherapy group (G4), and the PZQ+TOR dual therapy group (G5). The significant (<span class="html-italic">p</span> &lt; 0.05) differences are (*) compared to G2, (#) against G3, and (<span>$</span>) compared with G1.</p>
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<p>Histograms of the immune response and antioxidant status after different therapies. The data are expressed as the means ± SD of the following groups (n = 4): the uninfected control group (G1), the infected untreated group (G2), the PZQ monotherapy group (G3), the TOR monotherapy group (G4), and the PZQ+TOR dual therapy group (G5). Abbreviations: total immunoglobulin G (Total IgG), interleukin 10 (IL-10), interferon-gamma (IFN-γ), nitric oxide (NO), malondialdehyde (MDA), superoxide dismutase (SOD), and catalase (CAT). A significant (<span class="html-italic">p</span> &lt; 0.05) difference is shown as (*) compared to G2, (#) against G3, and (<span>$</span>) compared with G1.</p>
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13 pages, 1114 KiB  
Article
DNA Prevalence of Eukaryotic Parasites with Zoonotic Potential in Urban-Associated Birds
by Xabier Cabodevilla, Juan E. Malo, Daniel Aguirre de Carcer, Julia Zurdo, Rubén Chaboy-Cansado, Alberto Rastrojo and Juan Traba
Birds 2024, 5(3), 375-387; https://doi.org/10.3390/birds5030025 - 24 Jul 2024
Viewed by 504
Abstract
Synanthropic birds might play an important role as reservoirs of many zoonotic endoparasites; however, little information is available on many parasites and their prevalence. Here, we use an approach based on targeted metagenomic detection through the use of DNA metabarcoding of faecal samples [...] Read more.
Synanthropic birds might play an important role as reservoirs of many zoonotic endoparasites; however, little information is available on many parasites and their prevalence. Here, we use an approach based on targeted metagenomic detection through the use of DNA metabarcoding of faecal samples to screen for circulating parasites in alien parakeets (Myiopsitta monachus and Psittacula krameri) and urban landfill-feeding storks (Ciconia ciconia) and gulls (Larus fuscus). We focus especially on potentially zoonotic parasites, with the aim of better understanding the zoonotic risk that these birds’ faeces may pose. We detected a total of 23 genera of eukaryotic parasites: six fungi, three protists, five nematodes, two cestodes and seven trematodes. Among them, six stood out for their relevance to human health: Cryptococcus spp., Aspergillus spp. and Candida spp. (fungi); Cryptosporidium spp. (a protist); and Ascaris spp. and Halicephalobus spp. (nematodes). In parakeets, we detected Cryptococcus spp. and Ascaris spp., the latter being detected in 10–20% of the samples. In the White Stork and the Lesser Black-backed Gull, we found a high prevalence of Aspergillus spp. (in 15% and 50% of the samples, respectively) and Candida spp. (in 63% and 82% of the samples, respectively), and the presence of Cryptosporidium spp. in 10% of the samples. We detected Halicephalobus spp. in one gull sample (2%). Our results show that synanthropic birds may act as vectors and reservoirs of zoonotic parasites and their faeces could pose a risk to human health associated with the zoonotic parasites present in them. This should be taken into account when developing management plans for urban populations of these bird species. Full article
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<p>Prevalence of zoonotic fungi genera detected in bird faeces. All genera could parasitise humans; in black are those described as zoonotic in <a href="#birds-05-00025-t002" class="html-table">Table 2</a> and in grey are those that, according to the literature, may have some effect on human health.</p>
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<p>Prevalence of zoonotic protist genera. Genera that could parasitise humans are highlighted in bold; in black are those described as zoonotic in <a href="#birds-05-00025-t002" class="html-table">Table 2</a>, and in grey are those that, according to the literature, may have some effect on human health. The genera that are not in bold are bird parasites.</p>
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<p>Prevalence of zoonotic metazoan genera. Genera that could parasitise humans are highlighted in bold; in black are those described as zoonotic in <a href="#birds-05-00025-t002" class="html-table">Table 2</a>, and in grey are those that, according to the literature, may have some effect on human health. The genera that are not in bold are bird parasites; most of them are harmless to humans or have been very rarely reported in humans.</p>
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13 pages, 6802 KiB  
Article
The Natural Infection of Freshwater Snails with the Avian Air Sac Fluke, Cyclocoelum mutabile (Trematoda: Cyclocoelidae), in Brazil
by Jordana Costa Alves de Assis and Hudson Alves Pinto
Diversity 2024, 16(7), 422; https://doi.org/10.3390/d16070422 - 19 Jul 2024
Viewed by 321
Abstract
Trematodes of the family Cyclocoelidae are parasites mainly of the respiratory system of birds and present a cosmopolitan distribution. Although infection with these flukes can result in pathological changes and even bird death, information on their life cycles is scarce and almost entirely [...] Read more.
Trematodes of the family Cyclocoelidae are parasites mainly of the respiratory system of birds and present a cosmopolitan distribution. Although infection with these flukes can result in pathological changes and even bird death, information on their life cycles is scarce and almost entirely based on experimental infection data. Thus, the generation of knowledge on the mollusks that act as natural intermediate hosts of cyclocoelids is necessary and can aid control measures against these air sac trematodes. In the present study, gastropod mollusks collected in an urban stream from Belo Horizonte, Minas Gerais, Brazil, were subjected to the compression technique for the detection of non-emerging larval trematodes. Tailless cercariae with confluent ceca were found in 8/30 (26.7%) specimens of Biomphalaria glabrata and 3/33 (9.1%) specimen of Physella acuta. Samples of the cercariae were subjected to morphological characterization and genetic study (28S, Cox-1, and Nad-1). For comparative purposes, adult trematodes previously collected in the air sac of a common gallinule (Gallinula galeata) found dead in another waterbody from the same region were also characterized. The molecular sequences obtained revealed a high degree of similarity (100% in 28S, 99.2% in Cox-1, and 99.5% in Nad-1) between larval stages found in mollusks and adult parasites found in G. galeata and morphologically identified as Cyclocoelum mutabile. The conspecificity with this widely distributed cyclocoelid was also corroborated by phylogenetic analysis and comparison with isolates of this species previously characterized in Peru and the Czech Republic (99.4–100% and 96.7–97.0% of similarity in Nad-1, respectively). Thus, the integrative analysis carried out in the present work enabled us to identify C. mutabile in mollusks in South America for the first time. The finding of B. glabrata and P. acuta as new intermediate hosts corroborates the importance of freshwater gastropods in the transmission of C. mutabile, as well as the low specificity to the mollusk group, as previously characterized through experimental studies. Full article
(This article belongs to the Special Issue Diversity of Wildlife Pathogens)
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<p>Larval stages of the <span class="html-italic">Cyclocoelum mutabile</span> found in <span class="html-italic">Biomphalaria glabrata</span> and <span class="html-italic">Physella acuta</span> from Brazil. (<b>A</b>) Redia with developing cercariae. (<b>B</b>) Detail of cercaria. (<b>C</b>) Anterior end of the cercaria with the anterior organ and pharynx. (<b>D</b>) Metacercaria. Scale bars: (<b>A</b>)—500 µm, (<b>B</b>)—150 µm, (<b>C</b>,<b>D</b>)—50 µm.</p>
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<p>Air sac flukes in a juvenile specimen of the common gallinule (<span class="html-italic">Gallinula galeata</span>) from Brazil. Scale bar: 1 cm.</p>
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<p><span class="html-italic">Cyclocoelum mutabile</span> found in <span class="html-italic">Gallinula galeata</span> from Brazil. (<b>A</b>) Specimen in toto. (<b>B</b>) Detail of the anterior end, with the pre-pharyngeal genital pore, oral opening, and pharynx. (<b>C</b>,<b>D</b>) Mature egg with occulate miracidium presenting a developed redia inside. Scale bars: (<b>A</b>)—2 mm, (<b>B</b>)—500 m, (<b>C</b>,<b>D</b>)—50 μm.</p>
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<p>Phylogenetic relationships between the <span class="html-italic">Cyclocoelum mutabile</span> found in mollusks (<span class="html-italic">Biomphalaria glabrata</span> and <span class="html-italic">Physella acuta</span>) and common gallinule (<span class="html-italic">Gallinula galeata</span>) from Brazil and selected members of the family Cyclocoelidae, inferred from partial sequences of the genes (<b>A</b>) 28S (1173 bp, evolutive model: GTR + G), (<b>B</b>) Cox-1 (306 bp, evolutive model: TN93 + G) and (<b>C</b>) Nad-1 (437 bp, evolutive model: HKY + G). The phylogenetic trees were generated by Bayesian Inference, and nodal supports are indicated by posterior probability. New sequences from the current study are shown in bold. Scale bars indicate the number of substitutions per site.</p>
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19 pages, 5479 KiB  
Article
Expression of Transposable Elements throughout the Fasciola hepatica Trematode Life Cycle
by Elizaveta K. Skalon, Nick V. Panyushev, Olga I. Podgornaya, Anastasia R. Smolyaninova and Anna I. Solovyeva
Non-Coding RNA 2024, 10(4), 39; https://doi.org/10.3390/ncrna10040039 - 3 Jul 2024
Viewed by 669
Abstract
Background: Transposable elements (TEs) are major components of eukaryotic genomes. The extensive body of evidence suggests that although they were once considered “genomic parasites”, transposons and their transcripts perform specific functions, such as regulation of early embryo development. Understanding the role of TEs [...] Read more.
Background: Transposable elements (TEs) are major components of eukaryotic genomes. The extensive body of evidence suggests that although they were once considered “genomic parasites”, transposons and their transcripts perform specific functions, such as regulation of early embryo development. Understanding the role of TEs in such parasites as trematodes is becoming critically important. Fasciola hepatica, a parasite affecting humans and livestock, undergoes a complex life cycle in diverse environments and hosts, and knowledge about its life cycle regulation is scarce so far. Methods: We summarized the data regarding the repetitive elements in F. hepatica and conducted bulk RNA-seq analysis across its life cycle stages. TE expression profiles were analyzed, focusing on differential expression and potential homology with previously described long non-coding RNAs (lncRNAs). Results: Differential expression analysis revealed stage-specific TE transcription patterns, notably peaking during egg and metacercariae stages. Some TEs showed homology with known lncRNAs and contained putative transcription factor binding sites. Interestingly, TE transcription levels were highest in eggs and metacercariae compared to adults, suggesting regulatory roles in trematode life cycle transitions. Conclusions: These findings suggest that TEs may play roles in regulating trematode life cycle transitions. Moreover, TE homology with lncRNAs underscores their significance in gene regulation. Full article
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<p>The pie chart showing the mean composition of the <span class="html-italic">F. hepatica</span> genome.</p>
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<p>Venn diagram showing candidate TEs with stage-specific expression in <span class="html-italic">F. hepatica</span> life cycle. The figures at the intersection of the diagram correspond to the number of commonly transcribed TEs, while the figures outside the intersections show the numbers of stage-specific TEs.</p>
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<p>The analysis of TE expression profiles between transcriptomes throughout <span class="html-italic">F. hepatica</span> life cycle stages. (<b>A</b>) The heatmap shows Jensen–Shannon divergence based on TE expression profiles for each sample; the darker the color, the greater the divergence. (<b>B</b>) The principal component analysis (PCA). Based on the TE expression profiles, the dot plot shows the distribution of samples between principal components 1 (<span class="html-italic">X</span>-axis) and 2 (<span class="html-italic">Y</span>-axis). Ad—adult; egg—egg; juv—21-day juvenile; met—metacercaria; NEJ1 h, 3 h, 24 h—newly emerged juveniles after 1, 3, and 24 h of excystment.</p>
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<p>The heatmap of the top 20 differentially expressed transposable elements sorted by <span class="html-italic">p</span>-values: (<b>A</b>)—LINE, (<b>B</b>)—LTR retroelements, (<b>C</b>)—DNA transposons, (<b>D</b>)—Unknown TE. Colored squares on the top indicate stages of the life cycle. Ad—adult, egg—egg, juv—21-day juvenile, met—metacercaria, NEJ1h, 3h, 24h—newly emerged juveniles after 1, 3, and 24 h after excystment.</p>
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<p>Volcano plots for TE differential expression—adult stage vs. the other stages of the life cycle. The X-axis is a beta value, an analog of fold change, and Y-axis is −log(10)q-val, where qval—q-value, a false discovery rate-adjusted <span class="html-italic">p</span>-value in Wald test. The blue dotted line corresponds to q-value &lt; 0.05. Each point in the chart represents a different TE. Black dots indicate false positives, while red dots indicate significant changes in expression. The positive beta value indicates upregulation, and negative is downregulation.</p>
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<p>Number of upregulated and downregulated TEs at each stage in comparison to the <span class="html-italic">F. hepatica</span> adults. NEJ1h, 3h, 24h—newly emerged juveniles 1, 3 and 24 h after excystment.</p>
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<p>Transcription factor binding site predictions in stage-specific upregulated TE transcripts in egg, metacercaria, and 21-day juvenile. The figure is the layout showing the localization of the identified TF binding sites (colored rectangles) on the forward and reverse TE chain (labelled to the left of the schemes) with the combined match <span class="html-italic">p</span>-value. The block height corresponds to the site significance, i.e., taller blocks are more significant. The consensus sequences of the TF binding sites are indicated in boxes under the schemes. Blue asterisks indicate the similar lncRNA TEs.</p>
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<p>Transcription factor binding site predictions in stage-specific upregulated TE transcripts in newly emerged juveniles of 1, 3, and 24 h age. The figure is the layout showing the localization of the identified TF binding sites (colored rectangles) on the forward and reverse TE chain (labelled to the left of the schemes) with the combined match <span class="html-italic">p</span>-value. The block height corresponds to the site significance, i.e., taller blocks are more significant. The consensus sequences of the TF binding sites are indicated in boxes under the schemes. Blue asterisks indicate the similar lncRNA TEs.</p>
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<p>Pipeline for differential expression (DE) analysis of mobile elements within various Fasciola hepatica life cycle stages.</p>
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23 pages, 42252 KiB  
Article
Gastrointestinal Helminths in Wild Felids in the Cerrado and Pantanal: Zoonotic Bioindicators in Important Brazilian Biomes
by Iago de Sá Moraes, Victória Luiza de Barros Silva, Beatriz Elise de Andrade-Silva, Ana Paula Nascimento Gomes, Nicoly Ferreira de Urzedo, Vitória Breda Abolis, Renata de Souza Gonçalves, Karina Varella Arpon, Zara Mariana de Assis-Silva, Lizandra Fernandes da Silva, Ellen Amanda Zago, Michelle Benevides Gonçalves, Ísis Assis Braga, Klaus Casaro Saturnino, Edson Moleta Colodel, Arnaldo Maldonado Júnior, Richard de Campos Pacheco and Dirceu Guilherme de Souza Ramos
Animals 2024, 14(11), 1622; https://doi.org/10.3390/ani14111622 - 30 May 2024
Viewed by 663
Abstract
Environmental changes in the Brazilian Pantanal and Cerrado facilitate the spread of parasitic diseases in wildlife, with significant implications for public health owing to their zoonotic potential. This study aimed to examine the occurrence and diversity of gastrointestinal parasites in wild felids within [...] Read more.
Environmental changes in the Brazilian Pantanal and Cerrado facilitate the spread of parasitic diseases in wildlife, with significant implications for public health owing to their zoonotic potential. This study aimed to examine the occurrence and diversity of gastrointestinal parasites in wild felids within these regions to assess their ecological and health impacts. We collected and analyzed helminth-positive samples from 27 wild felids using specific taxonomic keys. Diverse parasitic taxa were detected, including zoonotic helminths, such as Ancylostoma braziliense, Ancylostoma caninum, Ancylostoma pluridentatum, Toxocara cati, Toxocara canis, Dipylidium caninum, Taenia spp., Echinococcus spp., and Spirometra spp. Other nematodes, such as Physaloptera praeputialis and Physaloptera anomala, were identified, along with acanthocephalans from the genus Oncicola and a trematode, Neodiplostomum spp. (potentially the first record of this parasite in wild felids in the Americas). Human encroachment into natural habitats has profound effects on wild populations, influencing parasitic infection rates and patterns. This study underscores the importance of continuous monitoring and research on parasitic infections as a means of safeguarding both wildlife and human populations and highlights the role of wild felids as bioindicators of environmental health. Full article
(This article belongs to the Section Wildlife)
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<p>Geographical distribution of carcass collection of wild felids sites and biomes in Brazil.</p>
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<p>Municipalities in the Brazilian Cerrado and Pantanal biomes in which the remains of the wild felids were recovered for the analysis of gastrointestinal helminths are marked.</p>
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<p>Heatmap of total parasites found in each host by parasite type and a bar plot of total parasites by biome. Light yellow indicates lowest range of parasite counts, indicating relatively minor infections, and dark blue represents the highest rang of parasite counts, indicating severe infections.</p>
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<p>Analysis of parasite–host–environment relationships. Total of parasite count by Biome (Cerrado and Pantanal).</p>
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<p>Venn diagram representation of helminth co-infections observed in the studied wild felids. Colors denote different helminth taxa, and the numbers indicate the frequency of observed co-infections among the studied wild felids.</p>
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<p>Micrograph of <span class="html-italic">Oncicola</span> spp. (<b>A</b>) Anterior portion of male <span class="html-italic">Oncicola</span> spp. recovered from Panthera onca; (<b>B</b>) posterior portion. Blue arrowhead—proboscis with hooks; black arrowhead—testis; yellow arrowhead—lemnisci; green arrowhead—cement glands.</p>
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<p>Micrographs of <span class="html-italic">Toxocara</span> spp. detected in <span class="html-italic">Panthera onca</span>. (<b>A</b>) Cervical wing of <span class="html-italic">Toxocara cati</span>; (<b>B</b>) cervical wing of <span class="html-italic">Toxocara canis</span>; (<b>C</b>) anterior view of <span class="html-italic">T. cati</span> showing the three lips; (<b>D</b>) posterior view of a male <span class="html-italic">T. canis</span> with both spicules externalized. (<b>E</b>,<b>F</b>) Ventriculus that intercalated between the esophagus and the intestine in <span class="html-italic">T. canis</span>. Black arrowhead—lips; blue arrowhead—spicules; green arrowhead—digitiform process; white arrowhead—ventriculus.</p>
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<p>Micrograph of Ancilostomatids recovered from wild felids. (<b>A</b>,<b>D</b>) Anterior view of <span class="html-italic">Ancylostoma pluridentatum</span> from <span class="html-italic">Puma concolor</span>; (<b>B</b>,<b>E</b>) anterior view of <span class="html-italic">Ancylostoma caninum</span> from <span class="html-italic">P. concolor</span>; (<b>C</b>,<b>F</b>) anterior view of <span class="html-italic">Ancylostoma braziliense</span> from <span class="html-italic">Herpailurus yagouarandi</span>; (<b>G</b>) posterior view, male of <span class="html-italic">A. pluridentatum</span>; (<b>H</b>) posterior view, male of <span class="html-italic">A. braziliense</span>; (<b>I</b>) posterior view, female of <span class="html-italic">A. caninum</span>.</p>
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<p>Micrograph of <span class="html-italic">Physaloptera</span> spp. recovered from wild felids. (<b>A</b>,<b>B</b>) Anterior portion of <span class="html-italic">Physaloptera anomala</span> from <span class="html-italic">Leopardus pardalis</span>; (<b>C</b>) posterior portion of male <span class="html-italic">P. anomala</span>; (<b>D</b>) showing the sessile papillae; (<b>E</b>) posterior portion of female <span class="html-italic">P. anomala</span>; (<b>F</b>) posterior end, part of the uterus filled with eggs; (<b>G</b>) anterior portion; (<b>H</b>) posterior portion of female showing the cuticular sheath of <span class="html-italic">Physaloptera praeputialis</span> in <span class="html-italic">Herpailurus yagouarandi</span>.</p>
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<p>Micrograph of <span class="html-italic">Trichuris vulpis</span> recovered from wild felids. (<b>A</b>–<b>C</b>) Posterior portion of male <span class="html-italic">T. vulpis</span> from <span class="html-italic">Leopardus pardalis</span>; (<b>B</b>) highlighting the proximal portion of the sheath with spiny cuticle; (<b>C</b>) opening of the sheath with the tip of the spicule internalized. White arrowhead—spicule; black arrowhead—shealth; red arrowhead—tip of the spicule.</p>
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<p>Micrograph of cestodes recovered from wild felids. (<b>A</b>) Mature proglottid of <span class="html-italic">Spirometra</span> sp. from <span class="html-italic">Leopardus pardalis</span>; (<b>B</b>,<b>D</b>) anterior portion of <span class="html-italic">Taenia</span> spp. from a <span class="html-italic">Herpailurus yagouarandi</span> and a <span class="html-italic">Panthera onca</span>, respectively; (<b>C</b>) gravid proglottid of <span class="html-italic">Dipylidium caninum</span> from <span class="html-italic">H. yagouarandi</span>; (<b>E</b>) <span class="html-italic">Echinococcus</span> sp. from <span class="html-italic">P. onca</span>. Blue arrowhead—uterus; yellow arrowhead—suckers; green arrowhead—hooks; white arrowhead—genital pore.</p>
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<p>Micrograph of <span class="html-italic">Neodiplostomum</span> (syn. <span class="html-italic">Fibricola</span>) spp. recovered from <span class="html-italic">Leopardus pardalis</span>. (<b>A</b>) Adult <span class="html-italic">Neodiplostomum</span> spp.; (<b>B</b>) anterior portion; (<b>C</b>) middle portion with eggs and vitelline glands; (<b>D</b>) posterior portion highlighting the testes.</p>
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27 pages, 1176 KiB  
Article
Diversity of Helminths of Insectivorous Mammals (Mammalia: Eulipothyphla) from Large Forest Protected Areas of the Middle Volga Region (European Russia)
by Nadezhda Yu. Kirillova, Alexander A. Kirillov, Alexander B. Ruchin and Alexander I. Fayzulin
Diversity 2024, 16(5), 307; https://doi.org/10.3390/d16050307 - 20 May 2024
Viewed by 613
Abstract
Insectivores (Eulypotiphla) are a substantial component of Russian forest ecosystems. The parasites of these animals also form an important part of natural biocenoses and act as one of the factors in the formation of biodiversity. The Mordovia Nature Reserve and National Park “Smolny” [...] Read more.
Insectivores (Eulypotiphla) are a substantial component of Russian forest ecosystems. The parasites of these animals also form an important part of natural biocenoses and act as one of the factors in the formation of biodiversity. The Mordovia Nature Reserve and National Park “Smolny” are large, forested areas located in the center of European Russia. We studied the helminth fauna of insectivores in these protected areas in 2018–2022. In total, using the method of complete helminthological necropsy, we examined 478 individuals of shrews, moles, and hedgehogs and recorded 34 species of parasitic worms, i.e., 8 trematode, 7 cestode, 1 acanthocephalan, and 18 nematode species. The most diverse helminth fauna was found in Sorex araneus (22 species). The composition of helminths in S. isodon (12), Neomys fodiens (9), Sorex minutus, and Erinaceus roumanicus (8 species each) turned out to be less diverse. The lowest species diversity of helminths was observed in Neomys milleri (3) and Talpae europaea (2 species). Taking into account the newly obtained data, we conducted a review of the helminth diversity in shrews, hedgehogs, and moles in the Middle Volga region. According to our literature data, the helminth fauna of insectivores in this region consists of 52 species, including 14 cestodes, 13 trematodes, 22 nematodes, and 3 acanthocephalans. Most of them belong to the Palearctic faunal complex (36 species). The helminth fauna of insectivores in the studied protected areas was compared with the helminth fauna of micromammals in other areas of the Middle Volga region. Our comparative analysis showed a high and average degree of similarity in the helminth fauna within individual species and genera of Eulipotyphla. Full article
(This article belongs to the Section Animal Diversity)
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<p>Map of the Middle Volga region showing regions with cases of detection of helminth species in insectivorous mammals (dark red—more than 30 species of helminths detected; red—10 or more helminth species; pink—less than 10 species; white—no helminthological studies). The numbers show the number of helminth species identified.</p>
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<p>Dendrogram of the similarity of the helminth fauna of insectivores from different areas of the Middle Volga region, obtained using the Morisita index (UPGMA). 1–5—groups with the most similar helminth fauna. Correlation coefficient: r = 0.935.</p>
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11 pages, 1657 KiB  
Article
Diplostomum cf. vanelli Yamaguti, 1935 (Trematoda: Diplostomidae Poirier, 1886): Morpho-Molecular Data and Life Cycle
by Anna V. Izrailskaia, Vladimir V. Besprozvannykh and Michael Yu. Shchelkanov
Diversity 2024, 16(5), 286; https://doi.org/10.3390/d16050286 - 10 May 2024
Viewed by 619
Abstract
Furcocercariae, of the trematodes from the family Diplostomidae, were found in freshwater snails—Radix auricularia, which were collected in a reservoir located on Popov Island (Peter the Great Bay, Sea of Japan). The life cycle was experimentally reproduced for the first time, [...] Read more.
Furcocercariae, of the trematodes from the family Diplostomidae, were found in freshwater snails—Radix auricularia, which were collected in a reservoir located on Popov Island (Peter the Great Bay, Sea of Japan). The life cycle was experimentally reproduced for the first time, while morphometric data for the development stages were studied and described for the newly discovered trematode. Moreover, molecular data for nuclear and mitochondrial markers were also obtained. It was determined that the morphometric characteristics of the trematode coincided with the species Diplostomum cf. vanelli, the molecular data analysis validates the species independence. Furthermore, the study highlights the issue of species identification in the Diplostomum genus. Full article
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<p><span class="html-italic">Diplostomum cf. vanelli</span>: (<b>A</b>,<b>B</b>) adult worm; (<b>C</b>,<b>D</b>) cercaria; (<b>E</b>,<b>F</b>) metacercaria. Scale bars: µm.</p>
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<p>Phylogeny of the genera <span class="html-italic">Diplostomum</span> based on <span class="html-italic">28S</span> sequences using the Bayesian algorithm. A posterior probability of ≥50 was shown in the nodes. The scale bar indicates the number of substitutions per site. Samples from this study in bold.</p>
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<p>Phylogeny of the genera <span class="html-italic">Diplostomum</span> based on COX1 sequences using the Bayesian algorithm. A posterior probability of ≥50 was shown in the nodes. The scale bar indicates the number of substitutions per site. Samples from this study in bold.</p>
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12 pages, 668 KiB  
Article
Emerging and Reemerging Parasitic Diseases in Taiwan: A Retrospective Study of Clinical Case Reports in 2001~2018
by Shao-Lun Hsu and Chia-Kwung Fan
Pathogens 2024, 13(5), 383; https://doi.org/10.3390/pathogens13050383 - 5 May 2024
Viewed by 1192
Abstract
Emerging and re-emerging parasitic diseases can cause significant economic burdens at national and global levels. However, governments often underestimate or ignore these diseases, especially in developed countries. This retrospective, case-oriented study analyzed parasitic diseases reported in Taiwan between 2001 and 2018. One hundred [...] Read more.
Emerging and re-emerging parasitic diseases can cause significant economic burdens at national and global levels. However, governments often underestimate or ignore these diseases, especially in developed countries. This retrospective, case-oriented study analyzed parasitic diseases reported in Taiwan between 2001 and 2018. One hundred and thirty-two eligible clinical profiles of Taiwanese patients obtained from the NCBI, Scopus, Google Scholar, and Web of Science databases and local journals according to age, sex, source of infection, symptoms, risk factors, and geographical regions were analyzed. The analysis results showed that the number/frequency of cases caused by nematodes (46.97%) or protozoa (37.88%) was significantly higher than that of trematodes (9.85%) or cestodes (5.30%) (p < 0.0001). Northern Taiwan (46.97%) had a significantly higher rate than Southern Taiwan (33.33%), Central Taiwan (8.33%), and Eastern Taiwan (5.30%) (p < 0.05). The 15–65 age group (68.94%) had a significantly higher rate than the 65–90 age group (22.73%) and the 0–15 age group (8.33%) (p < 0.0001). Males (70.46%) had a significantly higher number/frequency of cases than females (29.54%) (p < 0.0001). People who acquired the infection through the food/soil route (32.58%) or who had a low immune status (32.58%) had a higher rate than travel-related infections (15.15%) (p < 0.001). The present study showed that emerging/reemerging parasitic infections continue to be of great concern to the lives and health of Taiwanese citizens and, if ignored, will threaten the health of the Taiwanese people; therefore, the delineation of preventive measures by health authorities is urgently warranted. Full article
(This article belongs to the Special Issue Parasite Infection and Tropical Infectious Diseases)
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<p>Study selection and data extraction were used from PubMed, Scopus, Google Scholar, Web of Science databases, and local journals, and the candidate articles from 2001 to 2018 were selected according to the inclusion criteria.</p>
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<p>(<b>A</b>) Distribution and (<b>B</b>) the number of clinical parasitic cases by searching the literature released online and published in local journals in both English and Chinese in different geographical areas along with different cities/counties in Taiwan from 2001 to 2018.</p>
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8 pages, 437 KiB  
Brief Report
Infection with Clonorchis sinensis (Cobbold, 1875) Metacercariae in Fish from the East Lake of Wuhan: Freshwater Fish in Urban Lakes May Act as Infection Sources of Liver Fluke
by Jia-Nan Jiang, Hui-Fen Dong, Hou-Da Cheng, Hong Zou, Ming Li, Wen-Xiang Li and Gui-Tang Wang
Microorganisms 2024, 12(5), 898; https://doi.org/10.3390/microorganisms12050898 - 30 Apr 2024
Viewed by 695
Abstract
The liver fluke disease caused by Clonorchis sinensis is one of the most serious food-borne parasitic diseases in China. Many freshwater fish and shrimps can be infected with C. sinensis metacercariae as the second intermediate hosts in endemic regions. Owing to the lack [...] Read more.
The liver fluke disease caused by Clonorchis sinensis is one of the most serious food-borne parasitic diseases in China. Many freshwater fish and shrimps can be infected with C. sinensis metacercariae as the second intermediate hosts in endemic regions. Owing to the lack of infected humans and the good administration of pet dogs and cats in cities of non-endemic regions, few fish are expected to be infected with C. sinensis metacercariae in urban lakes. To determine the infection of C. sinensis metacercariae in freshwater fish and shrimps in urban lakes, a total of 18 fish species and one shrimp species were investigated in the East Lake of Wuhan City. Metacercariae were isolated by artificial digestive juice and identified using morphology and rDNA-ITS2 sequences. Five species of fish, Pseudorasbora parva, Ctenogobius giurinus, Squalidus argentatus, Hemiculter leuciclus, and Rhodeus spp., were infected with C. sinensis metacercariae. The overall prevalence of C. sinensis was 32.5%. The highest prevalence was found in P. parva with 57.9%, while S. argentatus exhibited the highest mean abundance (13.9). Apart from the C. sinensis metacercariae, four species of other trematode metacercariae were also identified across twelve fish species in total. Owing to the consumption of undercooked fish and feeding cats with small fish caught by anglers, there is a potential risk that the small fish infected with C. sinensis metacercariae may act as an infection source to spread liver fluke. Given the complete life cycle of C. sinensis, stray cats and rats were inferred to act as the important final hosts of C. sinensis in urban lakes in non-endemic areas. Full article
(This article belongs to the Section Parasitology)
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<p>Morphological characteristics of <span class="html-italic">Clonorchis sinensis</span> (<b>A</b>) and four forms of other trematode metacercariae ((<b>B</b>), Form 1; (<b>C</b>), Form 2; (<b>D</b>), Form 3; (<b>E</b>), Form 4) collected from freshwater fish in the East Lake of Wuhan, China.</p>
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23 pages, 6306 KiB  
Article
Long Non-Coding RNA Levels Are Modulated in Schistosoma mansoni following In Vivo Praziquantel Exposure
by Pedro Jardim Poli, Agatha Fischer-Carvalho, Ana Carolina Tahira, John D. Chan, Sergio Verjovski-Almeida and Murilo Sena Amaral
Non-Coding RNA 2024, 10(2), 27; https://doi.org/10.3390/ncrna10020027 - 19 Apr 2024
Cited by 1 | Viewed by 1384
Abstract
Schistosomiasis is a disease caused by trematodes of the genus Schistosoma that affects over 200 million people worldwide. For decades, praziquantel (PZQ) has been the only available drug to treat the disease. Despite recent discoveries that identified a transient receptor ion channel as [...] Read more.
Schistosomiasis is a disease caused by trematodes of the genus Schistosoma that affects over 200 million people worldwide. For decades, praziquantel (PZQ) has been the only available drug to treat the disease. Despite recent discoveries that identified a transient receptor ion channel as the target of PZQ, schistosome response to this drug remains incompletely understood, since effectiveness relies on other factors that may trigger a complex regulation of parasite gene expression. Long non-coding RNAs (lncRNAs) are transcripts longer than 200 nucleotides with low or no protein-coding potential that play important roles in S. mansoni homeostasis, reproduction, and fertility. Here, we show that in vivo PZQ treatment modulates lncRNA levels in S. mansoni. We re-analyzed public RNA-Seq data from mature and immature S. mansoni worms treated in vivo with PZQ and detected hundreds of lncRNAs differentially expressed following drug exposure, many of which are shared among mature and immature worms. Through RT-qPCR, seven out of ten selected lncRNAs were validated as differentially expressed; interestingly, we show that these lncRNAs are not adult worm stage-specific and are co-expressed with PZQ-modulated protein-coding genes. By demonstrating that parasite lncRNA expression levels alter in response to PZQ, this study unravels an important step toward elucidating the complex mechanisms of S. mansoni response to PZQ. Full article
(This article belongs to the Section Long Non-Coding RNA)
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<p>Heatmap of differentially expressed long non-coding RNAs (lncRNAs) detected by RNA-Seq of adult worm couples obtained from mice across ten time points at 0, 0.25, 1, 3, 6, 9, 12, 24, 48, and 96 h after treatment (Treat) with a single, curative dose of praziquantel (400 mg/kg) delivered by oral gavage at 7 weeks post-infection. Hierarchical clustering of differentially expressed lncRNAs (lines) from adult worm samples (columns) harvested at various time points following PZQ treatment of infected mice, as indicated by the color bar at the top and the blue color scale at right. These results were obtained by re-analyses of the RNA-Seq data from McCusker et al. [<a href="#B11-ncrna-10-00027" class="html-bibr">11</a>] using the <span class="html-italic">S. mansoni</span> lncRNA reference transcriptome published by Maciel et al. [<a href="#B26-ncrna-10-00027" class="html-bibr">26</a>]. Gene expression levels were measured by RNA-Seq and are shown as Z-scores, which are the number of standard deviations below (blue, downregulated) or above (red, upregulated) the mean expression value among treated and control samples for each gene, as indicated in the scale at right. 189 lncRNAs were considered significantly differentially expressed, being 127 long intergenic ncRNAs, 58 antisense lncRNAs and 4 sense lncRNAs (FDR &lt; 0.05).</p>
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<p>Heatmaps of differentially expressed long non-coding RNAs (lncRNAs) detected by RNA-Seq of adult worm couples obtained from mice 14 h after treatment with a single, sub-lethal dose of praziquantel (100 mg/kg) delivered by intraperitoneal injection at 4 (<b>a</b>) or 7 (<b>b</b>) weeks post-infection. Hierarchical clustering of differentially expressed lncRNAs (lines) from five adult worm sample replicates (columns) harvested 14 h following praziquantel (PZQ) treatment of infected mice or from five control (CTL) replicates, as indicated by the color bars at the top and the color legend at right. These results were obtained by re-analysis of the RNA-Seq data from McCusker et al. [<a href="#B11-ncrna-10-00027" class="html-bibr">11</a>] using the <span class="html-italic">S. mansoni</span> lncRNA reference transcriptome published by Maciel et al. [<a href="#B26-ncrna-10-00027" class="html-bibr">26</a>]. Gene expression levels were measured by RNA-Seq and are shown as Z-scores, which are the number of standard deviations below (downregulated, light blue) or above (upregulated, dark blue) the mean expression value among treated and control samples for each gene, as indicated in the scale at right. (<b>a</b>) In the 4-weeks experiment, 181 lncRNAs were considered significantly differentially expressed, being 108 long intergenic ncRNAs, 57 antisense lncRNAs and 16 sense lncRNAs (FDR &lt; 0.05). (<b>b</b>) In the 7-weeks experiment, 348 lncRNAs were considered significantly differentially expressed, being 236 long intergenic ncRNAs, 95 antisense lncRNAs and 17 sense lncRNAs (FDR &lt; 0.05).</p>
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<p>Analysis of long non-coding RNAs (lncRNAs) differentially expressed upon distinct praziquantel treatment regimens. The Venn diagram shows the number of lncRNAs that were detected as differentially expressed in one experiment alone, or at the same time in three different experiments: RNA-Seq from adult worm couples obtained from mice across ten time points at 0, 0.25, 1, 3, 6, 9, 12, 24, 48, and 96 h after treatment with a single, curative dose of PZQ (400 mg/kg) delivered by oral gavage at 7 weeks post-infection (“Time-course” experiment, pink circle); (ii) RNA-Seq from adult worm couples obtained from mice 14 h after treatment with a single, sub-lethal dose of PZQ (100 mg/kg) delivered by intraperitoneal injection at 4 weeks post-infection (“4-weeks” experiment, blue circle); (iii) RNA-Seq from adult worm couples obtained from mice 14 h after treatment with a single, sub-lethal dose of PZQ (100 mg/kg) delivered by intraperitoneal injection at 7 weeks post-infection (“7-weeks” experiment, green circle).</p>
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<p>Validation by RT-qPCR of lincRNAs detected as differentially expressed upon praziquantel treatment <span class="html-italic">in vivo</span>. Ten lincRNAs detected by RNA-Seq as differentially expressed were selected for in vitro RT-qPCR assays, namely: (<b>a</b>) SmLINC101519-IBu, (<b>b</b>) SmLINC105115-IBu, (<b>c</b>) SmLINC110492-IBu, (<b>d</b>) SmLINC121232-IBu, (<b>e</b>) SmLINC133371-IBu, (<b>f</b>) SmLINC142502-IBu, (<b>g</b>) SmLINC159037-IBu, (<b>h</b>) SmLINC161393-IBu, (<b>i</b>) SmLINC163938-IBu, and (<b>j</b>) SmLINC172840-IBu. For each of the ten selected lincRNAs, the expression profiles obtained with RNA-Seq are shown on the right (blue line) as TMM (trimmed mean of M values), whereas the RT-qPCR results are shown on the left (black line). For the RT-qPCR data, mean ± SEM from three biological replicates is shown, and a linear mixed-effects statistical model was applied. * <span class="html-italic">p</span>  &lt;  0.05, ** <span class="html-italic">p</span>  &lt;  0.01, *** <span class="html-italic">p</span>  &lt;  0.001, and **** <span class="html-italic">p</span>  &lt;  0.0001.</p>
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<p>RNA-Seq expression profiles of selected lincRNAs measured in control assays at different <span class="html-italic">S. mansoni</span> life-cycle stages. The expression levels of the ten selected lincRNAs are shown, namely: (<b>a</b>) SmLINC101519-IBu, (<b>b</b>) SmLINC105115-IBu, (<b>c</b>) SmLINC110492-IBu, (<b>d</b>) SmLINC121232-IBu, (<b>e</b>) SmLINC133371-IBu, (<b>f</b>) SmLINC142502-IBu, (<b>g</b>) SmLINC159037-IBu, (<b>h</b>) SmLINC161393-IBu, (<b>i</b>) SmLINC163938-IBu, and (<b>j</b>) SmLINC172840-IBu. These <span class="html-italic">S. mansoni</span> lincRNAs were selected after re-analysis of RNA-Seq public datasets of parasites collected from mice treated with PZQ in the “time-course experiment” [<a href="#B11-ncrna-10-00027" class="html-bibr">11</a>]. The y-axis shows the expression level (shown as TMM—trimmed mean of M values) for each lincRNA in control RNA-Seq assays, as compiled by Silveira et al. [<a href="#B31-ncrna-10-00027" class="html-bibr">31</a>], at the stage indicated in the x-axis as follows: miracidia/sporocysts (M/S), cercariae (C), schistosomula (S), adult males (M), and adult females (F).</p>
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<p>Analysis of the co-expression modules identified in WGCNA. (<b>a</b>) This plot shows the proportion of differentially expressed genes (DEGs) inside each of the 15 modules. The total number of genes inside each module is proportional to the point size; blue points (representing “brown2”, “floralwhite”, “plum” and “plum2”) indicate modules that contain the differentially expressed lncRNAs tested by RT-qPCR. The most significantly enriched and relevant gene ontology (GO) terms, including biological process and molecular function terms, are shown for “brown2” (<b>b</b>) and “floralwhite” (<b>c</b>) modules. The size of each point is proportional to the precision, also known as gene ratio (i.e., the proportion of genes in the input list that are annotated to the function). The colors show the statistical significance of the enrichment.</p>
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<p>Single-cell clusters expression profiles of three RT-qPCR validated differentially expressed lncRNAs matching the expression profile of three differentially expressed protein-coding genes modulated under praziquantel exposure [<a href="#B11-ncrna-10-00027" class="html-bibr">11</a>]. UMAP plots show the expression enrichment of genes and are colored by gene expression (blue = low, red = high). The scale represents log10(UMIs+1). The genes with expression enrichment in tegument are (<b>a</b>) SmLINC121232-IBu and (<b>b</b>) Smp_086480 (SmTAL2); in vitellocytes (<b>c</b>) SmLINC110492-IBu and (<b>d</b>) Smp_076320 (Myb/SANT-like DNA-binding domain-containing protein); and in neurons and muscles (<b>e</b>) SmLINC101519-IBu and (<b>f</b>) Smp_105220 (Lymphocyte antigen 6B).</p>
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16 pages, 29694 KiB  
Article
Histopathological Lesions Caused by a Digenean Trematode in a Pest Apple Snail, Pomacea canaliculata, in Its Native Geographic Distribution Area
by Lorena Evangelina Martinez, Carmen Gilardoni, Cintia Débora Medina, Florencia Cremonte and Jorge Alejandro Etchegoin
Animals 2024, 14(8), 1191; https://doi.org/10.3390/ani14081191 - 16 Apr 2024
Viewed by 818
Abstract
Pomacea canaliculata is one of the most dangerous invasive species. Morphological and molecular analyses have revealed that a digenean species belonging to the family Echinostomatidae parasitizes this snail at two sites in Buenos Aires Province, Argentina, South America. Molecular results confirmed that the [...] Read more.
Pomacea canaliculata is one of the most dangerous invasive species. Morphological and molecular analyses have revealed that a digenean species belonging to the family Echinostomatidae parasitizes this snail at two sites in Buenos Aires Province, Argentina, South America. Molecular results confirmed that the species belongs to a genus closely related to Patagifer. Analysis of the 28S rDNA showed that the sequences of the rediae and metacercariae are identical, indicating that the apple snail acts as the first and second intermediate host. The cercariae may encyst as metacercaria inside the redia and also emerge and re-infect the same snail or another snail. The prevalence of digeneans was higher in one of the sampling locations (15.1% vs. 0.72%), probably because the bird species that acts as the definitive host is more abundant in that area. Histopathological examination showed that the parasite quickly invades multiple host organs (gills, intestines, albumen gland, lung, kidney, and mantle border) besides the gonad and digestive gland, as is usual in digeneans. In addition, the partial or total castration of snails was observed in cases of moderate and high infection intensity. In males, there was loss of integrity in testicular tubules, while in females, the replacement of ovarian tissue by rediae was found. Full article
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<p>Developmental stages of Echinostomatidae gen. et. sp. (<b>A</b>) Redia, in vivo, showing pharynx (ph), caecum (dc), cercaria (Ce), podium (p); (<b>B</b>) cercarial body, line drawing; (<b>C</b>) tail ventral view, line drawing; (<b>D</b>) detail of head collar spines; (<b>E</b>) metacercaria from pericardial cavity, in vivo, showing the excretory granules (eg) and spines (s). Bars = 50 µm (<b>D</b>), 100 µm (<b>B</b>,<b>E</b>), 150 µm (<b>C</b>), and 200 µm (<b>A</b>).</p>
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<p>Phylogram resulting from using Bayesian inference (BI) and maximum likelihood (ML) on the partial 28S rDNA gene sequences of Echinostomatidae rooted in <span class="html-italic">Fasciola hepatica</span>. Posterior probability values (BI) and bootstrap values (ML) associated with the branches are shown as BI/ML; support values lower than 0.70 (BI) and 50 (ML) are not shown. The scale bar indicates the number of substitutions per site. Newly generated sequences are highlighted in bold.</p>
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<p>Histological sections (H&amp;E) of unparasitized and parasitized snails. (<b>A</b>) Unparasitized testis. (<b>B</b>) Testis showing hemocytic encapsulation (black arrow) and tissue lysis (black asterisk). (<b>C</b>) Redia between the sheath groove of the penis (arrowhead). (<b>D</b>) Unparasitized ovary. (<b>E</b>) Atrophied ovarian tubule and gonadal connective tissue replaced by rediae in different stages of development. (<b>F</b>) Rediae with cercariae inside the albumen gland. Abbreviations: Ce (cercariae), dc (developing cercariae), dgt (digestive gland tubule), gb (germinal ball), hl (hemolymphatic space), oo (oocyte), ot (ovarian tubules), r (rediae), sc (secretory cells of the albumen gland), s (spermatocyte), sz (spermatozoa), and tt (testis tubules). Scale bars: 100 µm (<b>A</b>); 200 µm (<b>B</b>–<b>F</b>).</p>
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<p>Histological sections (H&amp;E) of unparasitized and parasitized snails. (<b>A</b>) Unparasitized digestive gland. (<b>B</b>) Rediae invading the connective tissue of the digestive gland. (<b>C</b>) Compressed digestive gland tube (black arrow) caused by rediae and metacercariae. (<b>D</b>) Normal structure of the gill filament. (<b>E</b>) Deformation of the gill filament caused by the presence of numerous rediae (black triangle). (<b>F</b>) Hemocytic encapsulation and granulocytomas (black asterisks) in the gill. (<b>G</b>) Enlargement of the intestinal wall caused by rediae in the connective tissue. (<b>H</b>) Rupture of the muscle fibers in the foot caused by the presence of rediae (white arrow). (<b>I</b>) Metacercariae between renal tubules. Abbreviations: cw (cyst wall), dgt (digestive gland tubules), gf (gill filament), hl (hemolymphatic space), L (lumen of digestive gland tube), li (light of the intestine), mly (muscle layer), ms (muscle fibers), ome (outer mantle epithelium), ph (pharynx), mc (metacercariae), r (rediae), c (cercariae), rhi (renal hemocyte islet), and rt (renal tubule). Scale bars: 100 µm (<b>A</b>,<b>G</b>,<b>H</b>,<b>I</b>); 200 µm (<b>B</b>–<b>F</b>).</p>
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14 pages, 1609 KiB  
Article
New Data on the Larval Stages of Leptophallus nigrovenosus (Digenea, Plagiorchiata)
by Srisupaph Poonlaphdecha, Alexis Ribas, Albert Martínez-Silvestre and Mercedes Villa
Animals 2024, 14(8), 1154; https://doi.org/10.3390/ani14081154 - 10 Apr 2024
Viewed by 1117
Abstract
(1) Background: Leptophallus nigrovenosus, an esophageal parasite that primarily affects water snakes of the genus Natrix, has a known life cycle that involves snail and amphibian hosts. However, the biological aspects, chaetotaxic patterns, and pathogeny of this parasite in its hosts [...] Read more.
(1) Background: Leptophallus nigrovenosus, an esophageal parasite that primarily affects water snakes of the genus Natrix, has a known life cycle that involves snail and amphibian hosts. However, the biological aspects, chaetotaxic patterns, and pathogeny of this parasite in its hosts have not been fully elucidated. (2) Methods: Snails (Planorbarius metidjensis) were collected in Spain and examined for cercaria emergence. The larvae were used to experimentally infect Salamandra salamandra, and metacercariae were isolated. Their chaetotaxy was studied using microscopy and scanning electron microscopy. The eye histology was also examined. (3) Results: The cercariae displayed distinctive morphological characteristics. The results of this study revealed three types of ciliated sensory papillae on the cercarial teguments, suggesting an adaptation for host detection and orientation. The metacercariae isolated from subcutaneous tissues showed oval bodies covered in spines. The chaetotaxy patterns matched those of Leptophallinae species. This is the first report of the presence of L. nigrovenosus in the snail P. metidjensis. Additionally, this study detected metacercariae in the eyes of S. salamandra, emphasizing the need for further research on trematode infections in amphibian eyes. (4) Conclusions: Members of the genus Salamandra can serve as secondary intermediate hosts for L. nigrovenosus, and the presence of metacercariae in amphibian eyes may have implications for the survival and habitat management of these amphibians. Understanding this parasite’s prevalence, transmission dynamics, and impacts on host populations is crucial for conservation strategies. Full article
(This article belongs to the Section Herpetology)
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<p>The life cycle of <span class="html-italic">Leptophallus nigrovenosus</span> according to a literature compilation and our findings. (<b>a</b>) Definitive host: Ophidians are the main definitive hosts of the adult trematodes; they acquire the parasite by feeding on amphibians. (<b>b</b>) Eggs expelled in the feces of reptiles reach a body of water. (<b>c</b>) The first intermediate host: After the eggs are ingested, the miracidium leaves the egg shell in the snail intestine, develops into a sporocyst, and starts to produce cercariae (asexual multiplicative reproduction). (<b>d</b>) The cercariae are shed from the snail and actively search for a second intermediate host (amphibians). (<b>e</b>) The first intermediate host: Anurans and urodeles (larvae and adults) in contact with water harbor the metacercariae as a result of cercariae penetration in different parts of their bodies.</p>
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<p>The distribution of the integumentary ventral sucker papillae on a cercaria of <span class="html-italic">Leptophallus nigrovenosus</span>.</p>
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<p>The distribution of the integumentary papillae on a cercaria of <span class="html-italic">Leptophallus nigrovenosus</span>: ventral surface, dorsal surface, and tail, from left to right.</p>
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<p>The patterns of argentophilic papillae on the scanning electron micrographs of <span class="html-italic">L. nigrovenosus</span> cercariae: (<b>a</b>) the morphology of the integumental cephalic ring papillae; (<b>b</b>) the anterior ventral surface and (<b>c</b>,<b>d</b>) posterior lateral view of a cercaria, showing medium sensilla; (<b>e</b>) the stylet’s integumentary surface; (<b>f</b>,<b>g</b>) the details of small dome sensilla on a ventral sucker; (<b>h</b>) the ventral topographic surface of a cercaria; (<b>i</b>) the tail region; (<b>j</b>) the secretory penetration glands; and (<b>k</b>) the dorsal anterior integumental long sensilla.</p>
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<p>(<b>a</b>) An encysted mature metacercaria from a salamander eye (direct observation in saline solution), at 25 days post-infection; (<b>b</b>–<b>d</b>) metacercariae observed using histology, at 25 days post-infection; (<b>e</b>) an excysted metacercaria at 25 days post-infection (direct observation in saline solution); (<b>f</b>) an excysted metacercaria at 25 days post-infection, stained with Semichon’s carmine; and (<b>g</b>) a metacercaria at 25 days post-infection, located in the retrobulbar choroidal space, close to the sclerotia and surrounded by fibrous tissue and a low inflammatory reaction.</p>
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