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Induction of penetration gland emptying by cercariae of the bird schistosomes Trichobilharzia szidati and T. regenti employing linoleic acid, linolenic acid, praziquantel and calcium ionophore A23187 showed that both postacetabular and circumacetabular cells released their content at chosen stimulant concentrations. The gland secretions consisted of soluble and insoluble parts. The former one adhering to the ground seemed to have different saccharide composition from the glands of Schistosoma mansoni. It bound labelled saccharides, thus exhibiting lectin-like activity. Protein profiles of the latter one were identical after stimulation by all four stimulants in T. szidati. The soluble secretions contained several proteolytic enzymes; 31 kDa and 33 kDa cysteine proteases were identified in E/S products of T. szidati and T. regenti, respectively. The circumacetabular glands contained a significant amount of calcium. Immunohistochemistry revealed that the origin of E/S products after in vitro stimulation is in both penetration glands and tegumental structures. No crossreactivity was observed between the bird schistosomes and a serum raised against S. mansoni elastase.

The karyotype of a European bird nasal schistosome, Trichobilharzia regenti Horák, Kolárová et Dvofák, 1998, was determined. The chromosome set comprises eight pairs, seven of which are autosomes and one sex pair consisting of ZZ in males and ZW in females. The Z is the largest chromosome of the set, having a submeta-metacentric location of centromere, while the W is a small metacentric chromosome. More than 60% of the analyzed cells originating from both male and female sporocysts contained one or two small submetacentric supernumerary B chromosomes, occurring more frequently in female than in male cells. The formula of the karyotype structure is 2n = 16 + (B), n=5m+1sm-m+ 1sm+Zsm-m/Wm+(Bsm). The C-banding method revealed small pericentric heterochromatin blocks in the sex chromosome Z and all autosomes, except pairs 5 and 6. Satellited pair 5 showed a distinct terminal band representing the nucleolar organizer. Both the W and B chromosomes contained conspicuous heterochromatin blocks at the long arm adjacent to the centromere. The currently described supernumerary chromosomes in T. regenti represent the first record of B chromosomes within the family Schistosomatidae.

Wild rodent species are naturally infected by Schistosoma mansoni; however, the genetic characterization of the parasite, its parasitological features, and its role in human schistosomiasis are poorly understood. In this study, we isolated and characterized Schistosoma from naturally infected Holochilus sciureus, called HS strain, collected from a schistosomiasis endemic region in Maranhão State, Brazil. To isolate the parasite, miracidia obtained from the livers of H. sciureus were used to infect Biomphalaria glabrata of sympatric (called SB) and allopatric (called BH) strains, and the produced cercariae were subcutaneously inoculated into hamsters and/or BALB/c mice. Parasitological kinetics in experimentally infected hosts were evaluated, and the tRNACys-12S (referred to as 16S herein) and cox 1 regions of mtDNA from isolated worms were amplified and sequenced. Only miracidia obtained from infected mice, but not from hamsters, were capable of infecting B. glabrata, allowing maintenance of the isolated parasite. Cox1 and 16S mtDNA sequences showed 100% similarity with S. mansoni, and phylogenetic analysis showed that the HS strain of S. mansoni forms an assemblage with isolates from America and Kenya, confirming the conspecificity. Experimental infection of B. glabrata SB with S. mansoni HS resulted in two peaks of cercariae shedding at 45 and 70 days post-infection (dpi) and caused higher mortality than in B. glabrata BH. The worm recovery rate in mice was approximately 13%, and the peak of egg elimination occurred at the 10th week post-infection. Therefore, S. mansoni obtained from H. sciureus was successfully isolated, genetically characterized, and maintained in mice, allowing further study of this schistosome strain. [ABSTRACT FROM AUTHOR]

Precise identification of avian schistosomes in the genus Trichobilharzia at the species level is difficult and requires both traditional morphological and molecular techniques. To obtain satisfactory results by traditional methods, the characteristics of the intact adults or large fragments of male and females are necessary. The present study aimed to introduce a more efficient method for collecting eggs and both fragments and intact worms for morphological identification of visceral Trichobilharzia spp. Thirty-eight domestic ducks (twenty-eight fresh and ten frozen) were studied. For fresh samples, warm saline (40–45 °C) was injected into the portal vein or liver tissue, followed by slicing of the liver to small pieces in a large Petri dish. All materials were then transferred into the laboratory sieves arranged from the largest to the smallest mesh size and while crushed with the hand, washed, and filtered using a trigger water sprayer. The collected materials were studied under a stereomicroscope for parasite eggs, fragments, and full-length worms. Out of 28 freshly killed ducks, 19 (67.9%) and of 10 frozen ducks 6 (60%) were positive for visceral Trichobilharzia spp. The full-length worms and large fragments of male worms were mostly recovered with the mesh no. 150 (diameter of 106 μm) and small fragments, especially of females, and eggs with the mesh no. 270 (diameter of 53 μm). In addition to large numbers of fragments, 15 full-length adults were obtained from fresh and 2 from frozen ducks. The number of collected full-length adults was related to the worm burden. Since morphological description of different species of the genus Trichobilharzia is primarily based on the availability of adult worms, the application of methods that provide a higher number of intact males and females will result in better characterization of the species and deposition of appropriate voucher specimens. These results show the present method as a suitable tool for the collection of quality adults of visceral Trichobilharzia spp. in ducks. [ABSTRACT FROM AUTHOR]

Swimmer's itch has historically been controlled by applying copper sulfate (CuSO4) to lakes as a way to eliminate snails that serve as the intermediate hosts for swimmer's itch-causing parasites. CuSO4 is still sometimes applied specifically to areas of lakes where swimmer's itch severity is high. It is unclear whether targeted application of chemical molluscicides like CuSO4 is effective for controlling swimmer's itch. Previous research has found that the larval stage of the parasites responsible for swimmer's itch are released from infected snails and are concentrated by onshore and alongshore winds, and thus, may not be affected by such focused applications. In this study, we evaluated the impact of targeted CuSO4 application to a specific recreational swimming area in a lake in Michigan. We measured the effect on snail populations, as well as on the presence/abundance of swimmer's itch-causing parasites using qPCR. Ultimately, while CuSO4 was confirmed to significantly reduce populations of snails within the treatment area, it was found to have no significant impact on swimmer's itch-causing parasites in the water, likely due to the free-swimming larval stages (cercariae) moving into the treatment area from surrounding regions. [ABSTRACT FROM AUTHOR]

Cathepsin L-like protease is an important member of the papain-like cysteine protease and plays numerous indispensable roles in the biology of parasitic organisms. In a previous study, we identified a gene encoding a cathepsin L-like protease of Clonorchis sinensis (CsCPL) that was detected in the cercaria, metacercaria, and adult worm stages by immunolocalization, suggesting that this cysteine protease may be important and involved in the development of C. sinensis. In this study, the mature domain of CsCPL (CsCPL-m) was cloned and expressed in the form of inclusion bodies in Escherichia coli. After refolding, the recombinant CsCPL-m displayed optimal protease activity towards Z-Phe-Arg-AMC substrates but not towards Z-Arg-Arg-AMC, and the activity of the protease was inhibited completely by the cysteine protease-specific inhibitors E-64 and IAA, which further demonstrated that CsCPL belongs to the cathepsin L-like cysteine protease family. Recombinant CsCPL-m exhibited considerable activity at temperatures ranging from 28 to 42 °C, with the highest activity observed at 42 °C. Furthermore, recombinant CsCPL-m exhibited activity across a broad range of pH values (pH 4.0-8.0), with an optimal pH of 5.5. The Km and Vmax of the recombinant CsCPL-m towards Z-Phe-Arg-AMC were determined to be 5.71 × 10−6 M and 0.6 μM/min, respectively, at 37 °C and pH 5.5. The recombinant CsCPL-m could degrade BSA and gelatine, but could not degrade human hemoglobin and human immunoglobulin G. These results implied that CsCPL might participate in the catabolism of host proteins for nutrition during the parasitic life cycle of C. sinensis; thus, CsCPL could be used as a potential vaccine antigen and drug target against C. sinensis infection. [ABSTRACT FROM AUTHOR]

Trichobilharzia spp. have been identified as a causative agent of swimmers’ itch, a skin disease provoked by contact with these digenean trematodes in water. These parasites have developed a number of strategies to invade vertebrates. Since we have little understanding of the behavior of these parasites inside the human body, the monitoring of their invasion in snail host populations is highly recommended. In our research, lymnaeid snails were collected from several Polish lakes for two vegetation seasons. The prevalence of bird schistosomes in snail host populations was significantly lower than that of other digenean species. We were the first to detect the presence of the snails emitted Trichobilharzia regenti (potentially the most dangerous nasal schistosome) in Poland. In addition, by sequencing partial rDNA genes, we confirmed the presence of the snails positive with Trichobilharzia szidati in Polish water bodies, showing that swimmer’s itch is more frequent during summer months and that large snails are more often infected with bird schistosomes than small ones. [ABSTRACT FROM AUTHOR]

Nasal bird schistosomes can cause bilharziosis in birds and have the potential to cause swimmer's itch in humans. We determined the prevalence of bird schistosomes in 106 mallards ( Anas plathyrhynchos) from 11 water sources in Germany from 2014. Dissections were performed focusing on parasitic infections of the neural system. Infections with Trichobilharzia regenti (Horák et al. 1998) were found in 21% of the birds ( n = 22), whereas Bilharziella polonica (Kowalewski 1895) were found between the brain membranes (meninges) and the brain, in the spinal cord or in the intestine of 12% of the mallards ( n = 13). No significant influence of sex, age, and body condition between infected and non-infected animals was observed. Our study provides the first description of B. polonica from the neural system of birds and provides an epidemiological understanding of a parasite of human health concern. [ABSTRACT FROM AUTHOR]

After numerous reports the local press about the 'stinging water' in created on the Dzierżęcinka River-Water Valley reservoir and recognizing in bathers the symptoms of swimmers' itch, environmental study on the presence of bird schistosome larvae in snail hosts was conducted. Snails belonging to Lymnaeidae and Planorbidae were collected at two sites: (i) part of anthropogenic reservoir (192 individuals) and (ii) the river part (37 individuals). Higher prevalence of Digenea was observed in snail populations living in Water Valley (29.8 %) compared to Dzierżęcinka River (21.3 %). The larvae of bird schistosomes were recorded in both localities in 1.8 % of collected snails. The prevalence of bird schistosomes reached 2.9 % in Planorbarius corneus, 2.8 % in Radix auricularia, and 5.9 % in Radix balthica/labiata. Laboratory tests have shown that at 19 °C the number of bird schistosome cercariae released from snail hosts significantly exceeded the number of cercariae of other identified Digenea species. It is worth underlining that despite the low prevalence of bird schistosomes, the high number of released cercariae was sufficient to create a real threat of swimmers' itch in bathers. As indicated by the example presented, anthropogenic reservoirs create excellent conditions for Digenea species including bird schistosomes. In view of the real risk of people using the waters, tests on presence of the parasites in snail hosts should be included to the standard procedure of security control in bathing places. [ABSTRACT FROM AUTHOR]

Ten adult Quinqueserialis quinqueserialis specimens were removed from the intestine of a naturally infected muskrat, and scanning electron microscopy was used to study the morphological characteristics of the trematodes. The mature trematode, which was easy to recognize by the monostome holdfast organ, with no anterior cone, measured 2200-2500 μm in length by 900-1050 μm in width. The body was elongated and tapering at the anterior end, but the posterior end was rounded, and in some specimens was slightly truncated. The mouth opening lay at the anterior end and was surrounded by the oral sucker, which was round, small to medium in size, and subterminal. The tegument of the rim and inside of the oral sucker was smooth and had two types of papillae, domed and rosette papillae. Around the oral sucker, tegument was covered with sharp, pointed spines. The common genital pore was located on the median line of the body, posterior to the oral sucker. The cirrus had smooth tegument at the base and was armed with numerous conical spines throughout its length. The ventral surface was concave and provided with five distinct longitudinal rows of ventral papillae, which extended from the anterior to the posterior end of the body. Each row consisted of 15 to 20 papillae, making 81 to 88 papillae in all. These papillae were variable in size. In most specimens, the papillae were simple knob-like structures, but in some cases, they appeared to be bi- or trifurcate. The tegument at the base of each ventral papilla showed minute spiny pattern, but it was smooth or folded on top and had small rosette and ciliated papillae. Tegument at the edges of the worm was smooth in the mid-parts, spiny on lateral parts, and included rosette papillae. The dorsal surface of the worm was smooth and slightly convex, and the tegument was provided with two large domed papillae in one third of the anterior end of the dorsal part, few thick spines in the mid-part, and excretory pore at the level just posterior to the end. No spines or papillae were seen around the excretory pore. [ABSTRACT FROM AUTHOR]

Cathepsin Ls (CatLs), the major cysteine protease secreted by Fasciola spp., are important for parasite digestion and tissue invasion. Fasciola gigantica cathepsin L1H (FgCatL1H) is the isotype expressed in the early stages for migration and invasion. In the present study, a monoclonal antibody (MoAb) against recombinant F. gigantica cathepsin L1H (rFgCatL1H) was produced by hybridoma technique using spleen cells from BALB/c mice immunized with recombinant proFgCatL1H (rproFgCatL1H). This MoAb is an immunoglobulin (Ig)G1 with κ light chain isotype. The MoAb reacted specifically with rproFgCatL1H, the native FgCatL1H at a molecular weight (MW) 38 to 48 kDa in the extract of whole body (WB) of metacercariae and newly excysted juvenile (NEJ) and cross-reacted with rFgCatL1 and native FgCatLs at MW 25 to 28 kDa in WB of 2- and 4-week-old juveniles, adult, and adult excretory-secretory (ES) fractions by immunoblotting and indirect ELISA. It did not cross-react with antigens in WB fractions from other parasites, including Gigantocotyle explanatum, Paramphistomum cervi, Gastrothylax crumenifer, Eurytrema pancreaticum, Setaria labiato-papillosa, and Fischoederius cobboldi. By immunolocalization, MoAb against rFgCatL1H reacted with the native protein in the gut of metacercariae and NEJ and also cross-reacted with CatL1 in 2- and 4-week-old juveniles and adult F. gigantica. Therefore, FgCatL1H and its MoAb may be used for immunodiagnosis of both early and late fasciolosis in ruminants and humans. [ABSTRACT FROM AUTHOR]

Clonorchis sinensis excretory/secretory products (ESP) have gained high attentions because of their potential to be vaccine candidates and drug targets in C. sinensis prevention. In this study, we extensively profiled the characteristics of four C. sinensis cathepsin B cysteine proteases ( CsCB1, CsCB2, CsCB3, and CsCB4). Bioinformatics analysis showed all CsCBs contained signal peptides at the N-terminal. Functional domains and residues were found in CsCB sequences. We expressed four CsCBs and profiled immune responses followed by vaccine trials. Recombinant CsCBs could induce high IgG titers, indicating high immunogenicity of CsCB family. Additionally, ELISA results showed that both IgG1 and IgG2a levels apparently increased post-immunization with all four CsCBs, showing that combined Th1/Th2 immune responses were triggered by CsCB family. Both Real-time polymerase chain reaction (RT-PCR) and Western blotting confirmed that four CsCBs have distinct expression patterns in C. sinensis life stages. More importantly, we validated our hypothesis that CsCBs were C. sinensis excretory/secretory products. CsCBs could be recognized by C. sinensis-infected sera throughout the infection period, indicating that secreted CsCBs are immune triggers during C. sinensis infection. The protective effect was assessed by comparing the worm burden and egg per gram (EPG) between CsCB group and control group, showing that worm burden ( P < 0.01) and EPG ( P < 0.01) in CsCB2 and CsCB3 groups were significantly lower than in control group. In conclusion, we profiled secreted cathepsin B cysteine proteases family for the first time and demonstrated that all CsCB family were C. sinensis excretory/secretory products that may regulate host immune responses. [ABSTRACT FROM AUTHOR]

Thirty goats were randomly allocated in five groups of six animals each, for immunization with 1 × 10 phage particles of clones 11, 13, and 13 with Quil A adjuvant and wild-type M13KE phage at the beginning and 4 weeks later. The control group received phosphate-buffered saline. All groups were challenged with 200 metacercariae at week 6 and slaughtered 14 weeks later. The mean worm burdens after challenge were reduced by 46.91 % and 79.53 % in goats vaccinated with clones 13 and 13 with Quil A ( P < 0.05), respectively; no effect was observed in animals immunized with clone 11 and M13KE phage. Animals receiving clones 11, 13, and 13 with Quil A showed a significant reduction in eggs output. Vaccinated animals produced parasite-specific total IgG antibody which were boosted after challenge with metacercariae of F. hepatica. Furthermore, levels of anti-phage total IgG increased rapidly within 2 weeks of the first vaccination and were always significantly higher in all vaccinated goats than in the infected control group. The fluke burden of goats immunized with clones 13 and 13 with Quil A was significantly correlated with IgG2 and total IgG. Goats vaccinated with phage clones produced significantly high titres of IgG1 and IgG2 antibodies indicating a mixed Th1/Th2 response. These data indicate that cathepsin L1 mimotopes has a potential as a vaccine candidate against Fasciola hepatica, whose efficacy will be evaluated in other host species, including those of veterinary importance. [ABSTRACT FROM AUTHOR]

Cysteine protease plays a key role in host-parasite interactions. In this study, we identified a novel gene encoding a cathepsin B-like cysteine protease (AcCBL1) from the cDNA library of Angiostrongysus cantonensis fourth-stage larvae (L4) and characterized its biological role in the parasite. Sequence and phylogeny analysis showed that AcCBL1 is related to other cathepsin B family members with the conserved catalytic triad (Cys, His, Asn) and diagnostic occluding loop. In addition, the sequence contains a specific 'hemoglobinase motif' and might have a hemoglobinase (Hb)-degrading function. The recombinant AcCBL1 (rAcCBL1) exhibited the protease activity by gelation SDS/PAGE assay; rAcCBL1 can cleave the fluorogenic substrate Z-Arg-Arg-AMC, and the optimum pH was 5.5. The enzyme can hydrolyse several host proteins including Hb and human IgG in acidic pH, but low levels of hydrolysis were observed in neutral pH. Reverse transcription polymerase chain reaction revealed that AcCBL1 expression was detected throughout various developmental stages, L3, L4, adult male and female worms. Western blotting analysis indicated that AcCBL1 was an excretory/secretory product of L4 in mature form of protease. Immunolocalization demonstrated that AcCBL1 was mainly localized in the intestine of L4. These results suggest that rAcCBL1 may play an important role in the parasite nutrition uptake. [ABSTRACT FROM AUTHOR]