Clinical forms of peritoneal larval cestodiasis by Mesocestoides spp. in dogs: diagnosis, treatment and long term follow-up.

Canine peritoneal larval cestodiasis (CPLC) is a little-known parasitological infestation of the peritoneal cavity of wild and domestic carnivores with Mesocestoides spp. larvae. While adult Mesocestoides tapeworms reside within the small intestine, the larvae occasionally penetrate the host's intestinal wall, causing a potentially life-threatening peritonitis. Severity of infection as well as the host response influences the prognosis significantly, and early diagnosis and treatment are essential. However, due to the lack of specific symptoms, this condition is underdiagnosed and, furthermore, no clear effective treatment has yet been described. The aim of this study is therefore to report two clinical cases of CPLC in dogs and to illustrate their clinical presentation and follow-up to serve as a reference for clinicians and researchers alike. Both animals were presented with abdominal distention as their main complaint. They underwent clinical examination, abdominal ultrasonography, abdominocentesis, and laparotomy followed by biochemical, cytological, parasitological, and molecular examination of the collected samples. After surgical lavage, the dogs received anthelmintic treatment with either fenbendazole (FBZ) or praziquantel (PZQ). Overall, timely and prolonged administration of high doses of FBZ seems to be the most effective treatment method. Irrespective, to date, no treatment capable of complete eradication of the infection and prevention of recurrence of disease has been found. In conclusion, further investigation into appropriate treatment plans as well as diagnostic development is needed.

The potential for histone deacetylase (HDAC) inhibitors as cestocidal drugs.

BACKGROUND: Echinococcosis and cysticercosis are neglected tropical diseases caused by cestode parasites (family Taeniidae). Not only there is a small number of approved anthelmintics for the treatment of these cestodiases, but also some of them are not highly effective against larval stages, such that identifying novel drug targets and their associated compounds is critical. Histone deacetylase (HDAC) enzymes are validated drug targets in cancers and other diseases, and have been gaining relevance for developing new potential anti-parasitic treatments in the last years. Here, we present the anthelmintic profile for a panel of recently developed HDAC inhibitors against the model cestode Mesocestoides vogae (syn. M. corti). METHODOLOGY/PRINCIPAL FINDINGS: Phenotypic screening was performed on M. vogae by motility measurements and optical microscopic observations. Some HDAC inhibitors showed potent anthelmintic activities; three of them -entinostat, TH65, and TH92- had pronounced anthelmintic effects, reducing parasite viability by ~100% at concentrations of ≤ 20 µM. These compounds were selected for further characterization and showed anthelmintic effects in the micromolar range and in a time- and dose-dependent manner. Moreover, these compounds induced major alterations on the morphology and ultrastructural features of M. vogae. The potencies of these compounds were higher than albendazole and the anthelmintic effects were irreversible. Additionally, we evaluated pairwise drug combinations of these HDAC inhibitors and albendazole. The results suggested a positive interaction in the anthelmintic effect for individual pairs of compounds. Due to the maximum dose approved for entinostat, adjustments in the dose regime and/or combinations with currently-used anthelmintic drugs are needed, and the selectivity of TH65 and TH92 towards parasite targets should be assessed. CONCLUSION, SIGNIFICANCE: The results presented here suggest that HDAC inhibitors represent novel and potent drug candidates against cestodes and pave the way to understanding the roles of HDACs in these parasites.

Co-administration of silymarin elevates the therapeutic effect of praziquantel through modulation of specific antibody profiles, Th1/Th2/Tregs cytokines and down-regulation of fibrogenesis in mice with Mesocestoides vogae (Cestoda) infection.

Silymarin (SIL) represents a natural mixture of polyphenols showing an array of health benefits. The present study, carried out on a model cestode infection induced by Mesocestoides vogae tetrathyridia in the ICR strain of mice, was aimed at investigating the impact of SIL as adjunct therapy on the activity of praziquantel (PZQ) in relation to parasite burden, immunity and liver fibrosis within 20 days post-therapy. In comparison with PZQ alone, co-administration of SIL and PZQ stimulated production of total IgG antibodies to somatic and excretory-secretory antigens of metacestodes and modified the expression patterns of immunogenic molecules in both antigenic preparations. The combined therapy resulted in the elevation of IFN-γ and a decline of TNF-α and TGF-ß1 in serum as compared to untreated group; however, SIL attenuated significantly the effect of PZQ on IL-4 and stimulated PZQ-suppressed phagocytosis of peritoneal macrophages. In the liver, SIL boosted the effect of PZQ on gene expression of the same cytokines in a similar way as was found in serum, except for down-regulation of PZQ-stimulated TNF-α. Compared to PZQ therapy, the infiltration of mast cells into liver after SIL co-administration was nearly abolished and correlated with suppressed activities of genes for collagen I, collagen III and α-SMA. In conclusion, co-administration of SIL modified the effects of PZQ therapy on antigenic stimulation of the immune system and modulated Th1/Th2/Tregs cytokines. In liver this was accompanied by reduced fibrosis, which correlated with significantly higher reduction of total numbers of tetrathyridia after combined therapy as compared with PZQ treatment.

Differential Effects of the Flavonolignans Silybin, Silychristin and 2,3-Dehydrosilybin on Mesocestoides vogae Larvae (Cestoda) under Hypoxic and Aerobic In Vitro Conditions.

Mesocestoides vogae larvae represent a suitable model for evaluating the larvicidal potential of various compounds. In this study we investigated the in vitro effects of three natural flavonolignans-silybin (SB), 2,3-dehydrosilybin (DHSB) and silychristin (SCH)-on M. vogae larvae at concentrations of 5 and 50 µM under aerobic and hypoxic conditions for 72 h. With both kinds of treatment, the viability and motility of larvae remained unchanged, metabolic activity, neutral red uptake and concentrations of neutral lipids were reduced, in contrast with a significantly elevated glucose content. Incubation conditions modified the effects of individual FLs depending on their concentration. Under both sets of conditions, SB and SCH suppressed metabolic activity, the concentration of glucose, lipids and partially motility more at 50 µM, but neutral red uptake was elevated. DHSB exerted larvicidal activity and affected motility and neutral lipid concentrations differently depending on the cultivation conditions, whereas it decreased glucose concentration. DHSB at the 50 µM concentration caused irreversible morphological alterations along with damage to the microvillus surface of larvae, which was accompanied by unregulated neutral red uptake. In conclusion, SB and SCH suppressed mitochondrial functions and energy stores, inducing a physiological misbalance, whereas DHSB exhibited a direct larvicidal effect due to damage to the tegument and complete disruption of larval physiology and metabolism.

Reduction of oxidative stress and liver injury following silymarin and praziquantel treatment in mice with Mesocestoides vogae (Cestoda) infection.

Oxidative stress is a common mechanism contributing to hepatic damage and fibrogenesis in a variety of liver disorders. The liver is the target organ for many parasitic infections, hence there is a great demand for the development of novel treatment strategies. In the present study conducted on mice infected with larval stage of Mesocestoides vogae, we investigated effects of therapy with praziquantel (PZQ) alone and in combination with silymarin on liver GSH content, lipid peroxidation and larval reduction. Proliferation of liver cells by means of BrdU incorporation into DNA and production of superoxide anions by peritoneal adherent cells was measured to assess the antioxidant activity of silymarin. Drug administration was carried on from day 15 post infection (p.i.) for ten consecutive days and examination was performed during 20 days of follow-up the therapy. Larval M. vogae infection caused liver damage and triggered extensive oxidative stress, resulting in the abolishment of GSH redox balance and ROS-induced lipid peroxidation. PZQ administration caused short-term decline of GSH levels in healthy mice. Low GSH levels in infected mice were elevated gradually in response to the drug, but respiratory burst in cells was not reduced. Silymarin in combination with PZQ showed strong direct antioxidant capacity and stimulated the larvicidal effect of praziquantel. Treatment with PZQ and silymarin downregulated the generation of superoxide anions, prevented lipid peroxidation, stimulated GSH synthesis and proliferation of hepatocytes in infected livers. These findings demonstrated that silymarin can markedly decrease the liver injury and its co-administration with PZQ potentiate effect of therapy, probably due to the down-regulation of fibrogenesis.

Impact of treatment with praziquantel, silymarin and/or beta-glucan on pathophysiological markers of liver damage and fibrosis in mice infected with Mesocestoides vogae (Cestoda) tetrathyridia.

Mesocestoides vogae tetrathyridia infection in mice causes hepatocyte injury, hepatic granulomatous inflammmation, liver fibrosis and chronic peritonitis manifested with portal hypertension. To reduce the detrimental effect of parasites on the host liver, the effect of the anthelmintic drug praziquantel (PZQ) in combination with natural products silymarin (an antioxidant) and beta-glucan (an immunomodulator) was investigated. The therapeutic effect of drugs was assessed by means of aminotransferase (alanine aminotransferase (ALT) and aspartate aminotransferase (AST)) activities, content of albumin, total proteins and hyaluronic acid (HA) in sera of ICR mice infected with M. vogae larvae. Animals were treated with PZQ suspended in oil emulsion (Group 1), PZQ combined with silymarin incorporated into lipid microspheres (LMS) (Group 2), PZQ combined with beta-glucan incorporated into liposomes (LG) (Group 3), PZQ co-administered with LMS and LG (Group 4). Untreated animals (Group 5) served as the control. Treatment of animals started at the early chronic phase of infection (day 14 p.i.) and lasted 10 days; serum samples were collected on days 0, 7, 14, 25, 28, 31, 35 and 45 p.i. ALT and AST activities were significantly (P < 0.05) decreased in Groups 2, 3 and 4. HA content was significantly (P < 0.05 and 0.01) lower in Groups 2 and 4. Albumin levels were decreased in Groups 2 and 4, total protein concentration decreased in Groups 1 and 3 (P < 0.05 and 0.01). These results showed that combined treatment of PZQ with silymarin and/or beta-glucan was able to ameliorate or suppress fibrogenesis in the liver, protect liver cells from oxidative damage and, possibly, stimulate regeneration of the parenchyma.

Antibody response in mice infected with Mesocestoides vogae (syn. Mesocestoides corti) tetrathyridia after treatment with praziquantel and liposomised glucan.

The therapeutic effect of praziquantel (PZQ) involves synergy with the humoral immune response during helminth infections, which is modulated by parasitic antigens. During experimental murine infections with the larval stage of cestoda Mesocestoides vogae (syn. M. corti), dynamic changes in the IgG and IgM antibody serum levels to both soluble somatic and secretory larval antigens were investigated after administration of PZQ alone and after its co-administration with the immunomodulator (l-->3)-beta-D-glucan entrapped in liposomes (lip.glucan). During the 2 weeks of follow-up after termination of therapy, specific IgG and IgM serum levels to the somatic antigens (enzyme-linked immunosorbent assay test) significantly decreased, whereas concentrations of the antibodies to the secretory antigens moderately increased, both in comparison with the control. Moreover, the number of immunogenic larval antigens (analysed by Western blot) was higher after combined therapy in comparison with single drug administration, which correlated with the intensity of reduction of the larval counts in the liver and peritoneal cavity of mice. Our data showed that administration of PZQ alone and in combination with lip.glucan resulted in marked changes in the dynamics of IgG and IgM antibodies to the somatic larval antigens, which were probably induced by the newly exposed antigens. In this respect, glucan can enhance chemotherapeutic activity of PZQ against larval cestodes by means of stimulation of the macrophage/monocyte effector functions, which seemed to contribute to the more intense larval damage.

Praziquantel and liposomized glucan-treatment modulated liver fibrogenesis and mastocytosis in mice infected with Mesocestoides vogae (M. corti, Cestoda) tetrathyridia.

Beta-glucans are immunomodulators able to activate innate immunity and to potentiate acquired immune reactions. We investigated the impact of co-administration of liposomized beta-glucan on the larvicidal effect of the anthelmintic praziquantel (PZQ) in the livers and peritoneal cavities in mice infected with Mesocestoides vogae (M. corti). Also, within 2 weeks following therapy (up to day 29 p.i.) we examined collagen synthesis in the livers of mice by means of biochemical determination of hydroxyproline concentration, total mast cell counts and cell proliferative capacity using immunohistochemical and radiometrical methods. After co-administration of liposomized glucan (LG) and PZQ efficacy (%) was significantly higher than after treatment with either compound alone, particularly in the peritoneal cavity compared to the liver. In comparison with the control, more intense collagenesis was found in the B-liver parts (high intensity of infection) and lowering of collagen content in the A-parts (very weak infection). This effect was strongest after LG treatment and co-administration of PZQ abolished the pro-fibrotic effect of LG. In all groups, mast cell counts were higher in the B-liver parts than in the A-parts and the dynamics of mastocytosis was profoundly modulated following therapy. Whereas the effect of PZQ was only moderate, early and very strong onset was seen after LG treatment. Administration of PZQ suppressed LG induced-elevation of mast cells counts in both liver parts. Using DNA S-phase markers (BrdU and 3H-thymidine) the proliferative capacity was shown to be associated with several kinds of liver cells. Therapy significantly stimulated [3H]-thymidine incorporation (cell proliferation) only in the A-parts over that in control, the most after LG administration. In summary (i) the anthelmintic effect of PZQ could be enhanced after simultaneous administration of the immunomodulator beta-glucan entrapped in a liposomal carrier, (ii) intense mastocytosis seen after treatment with LG seems to have a direct role in the glucan's pro-fibrotic activity and can be abolished after co-administration of PZQ in a time-dependent manner, (iii) the pattern of cell proliferation indicates that in the case of PZQ treatment, the reparative processes of liver parenchyma are enhanced in an inverse correlation with the intensity of infection.

Praziquantel and albendazole damaging action on in vitro developing Mesocestoides corti (Platyhelminthes: Cestoda).

Parasitic flatworms present several steps of body architecture rearrangement during their fast transition from one developmental stage to another, which are, at least in part, responsible for their evasion from host immune response. Besides, different developmental stages present different degrees of susceptibility to drug action, and the identification of more susceptible stages is of importance for the definition of therapeutical approaches. Mesocestoides corti (syn. Mesocestoides vogae) is considered a good model to study cestode biology because it can be easily manipulated both in vivo and in vitro and due to its relatively close relationship to cestodes of medical relevance, such as those from genera Echinococcus or Taenia. We have analyzed the damaging action of two broad spectrum anthelmintic drugs (praziquantel and albendazole) throughout the in vitro strobilization process of M. corti in order to identify developmental stages or body structures more susceptible to these drugs. Tetrathyridia (larval stage) and segmented-induced worms were cultivated and treated with praziquantel and albendazole. Whole mounted samples, taken from different developmental stages, were fixed and stained with fluorophore-labeled WGA lectin and phalloidin for the analysis of tegument and muscles, respectively. Confocal laser scanning microscopy was used to identify anatomical changes and lesions caused by each anthelmintic drug in a 3D view. We demonstrated that both praziquantel and albendazole cause extensive tissue damage, especially on tegument, and that adult forms were the most susceptible to drug exposure.

Early post-larval development of the endoparasitic platyhelminth Mesocestoides corti: trypsin provokes reversible tegumental damage leading to serum-induced cell proliferation and growth.

Mesocestoides corti is a suitable in vitro model for studying the development of human endoparasitic platyhelminthes. Treatment with trypsin, supplemented with fetal bovine serum (FBS), induces M. corti development from larvae (tetrathyridia) to segmented adult worm; however, the role of this protease and of FBS in post-larval development induction remains unknown. To characterize the participation of trypsin enzymatic activity and of FBS in the induction of tetrathyridia growth and development, both stimuli were added to the larvae either together or sequentially. Additionally, specific inhibition of trypsin activity was also monitored. Finally, the effect of the enzyme on the parasite tegument as well as the proliferative activity and location of proliferating cells after induction of tetrathyridia development were also studied. We conclude that trypsin-induced tetrathyridia development to adult worm is FBS-dependent and that the effect of serum factors is dependent upon a previous trypsin-induced reversible damage to the larva tegument. In dividing and non-dividing tetrathyridia, proliferative activity of cells is mainly located within the apical massif in the anterior region and nerve cords of larvae, respectively. In tetrathyridia stimulated to develop to adult worms, an intense proliferative activity is evident along the nerve cords. Our results suggest that in natural infections the tetrathyridia tegument is temporally made permeable to growth factors by proteolytic enzyme activity in the intestine juice of the definitive host, thus leading to development to adult worms.

Pharmacological characterisation of neuropeptide F (NPF)-induced effects on the motility of Mesocestoides corti (syn. Mesocestoides vogae) larvae.

Neuropeptide F is the most abundant neuropeptide in parasitic flatworms and is analogous to vertebrate neuropeptide Y. This paper examines the effects of neuropeptide F on tetrathyridia of the cestode Mesocestoides vogae and provides preliminary data on the signalling mechanisms employed. Neuropeptide F (>/=10 microM) had profound excitatory effects on larval motility in vitro. The effects were insensitive to high concentrations (1 mM) of the anaesthetic procaine hydrochloride suggesting extraneuronal sites of action. Neuropeptide F activity was not significantly blocked by a FMRFamide-related peptide analog (GNFFRdFamide) that was found to inhibit GNFFRFamide-induced excitation indicating the occurrence of distinct neuropeptide F and FMRFamide-related peptide receptors. Larval treatment with guanosine 5'-O-(2-thiodiphosphate) trilithium salt prior to the addition of neuropeptide F completely abolished the excitatory effects indicating the involvement of G-proteins and a G-protein coupled receptor in neuropeptide F activity. Addition of guanosine 5'-O-(2-thiodiphosphate) following neuropeptide F had limited inhibitory effects consistent with the activation of a signalling cascade by the neuropeptide. With respect to Ca(2+) involvement in neuropeptide F-induced excitation of M. vogae larvae, the L-type Ca(2+)-channel blockers verapamil and nifedipine both abolished neuropeptide F activity as did high Mg(+) concentrations and drugs which blocked sarcoplasmic reticulum Ca(2+)-activated Ca(2+)-channels (ryanodine) and sarcoplasmic reticulum Ca(2+) pumps (cyclopiazonic acid). Therefore, both extracellular and intracellular Ca(2+) is important for neuropeptide F excitation in M. vogae. With respect to second messengers, the protein kinase C inhibitor chelerythrine chloride and the adenylate cyclase inhibitor MDL-2330A both abolished neuropeptide F-induced excitation. The involvement of a signalling pathway that involves protein kinase C was further supported by the fact that phorbol-12-myristate-13-acetate, known to directly activate protein kinase C, had direct excitatory effects on larval motility. Although neuropeptide F is structurally analogous to neuropeptide Y, its mode-of-action in flatworms appears quite distinct from the common signalling mechanism seen in vertebrates.

In vitro segmentation induction of Mesocestoides corti (Cestoda) tetrathyridia.

Mesocestoides corti is a suitable model for studying cestode development because of its ability to reproduce asexually and segment in vitro. The cultured parasite is also capable of sexual differentiation and, probably, reproduction. To establish conditions that increase the efficiency of in vitro M. corti larvae (tetrathyridia) segmentation, we tested the effects of an inducing agent and some physical parameters in cultures. We found that a 5% CO2-95% N2 gas phase, an incubation temperature of 39 C (instead of 37 C), and a 24-hr pretreatment with trypsin (10(5) BAEE/ml, BAEE = Na-benzoil-L-arginine ethyl ester unit of trypsin activity) in Roswell Park Memorial Institute (RPMI) 1640 medium supplemented with 20% fetal bovine serum (FBS) are able to increase individually or synergistically the segmentation rate of tetrathyridia. A segmentation rate of up to 100% was achieved on day 4 of culture, when all these conditions were used simultaneously, in comparison with an average rate of 40% obtained not before day 11 in cultures without any inducing treatment. Fetal bovine serum is essential for segmentation, and a concentration of 20% was established as the standard for induction.

The toxicity of praziquantel against Mesocestoides vogae (syn. corti) tetrathyridia can be assessed using a novel in vitro system.

We recently standardised Mesocestoides vogae (syn. corti) tetrathyridia cultures in the presence of sodium taurocholate. Parasite clustering and segmentation were observed as taurocholate-dependent effects in biphasic and monophasic media, respectively, and both were inhibited by a specific minimum inhibitory concentration (m.i.c.) of the cestocidal drugs albendazol and praziquantel. In the present study, we analysed the relationship between clustering inhibition and drug toxicity using praziquantel and a mouse experimental infection. In an "in vitro-in vivo" trial, a significant (ANOVA, P<0.05) reduction was observed in the infectivity of tetrathyridia previously cultured with praziquantel m.i.c. (0.06 micro g/ml) for 10 days. In an "in vivo-in vitro" trial, the clustering of tetrathyridia recovered from mice treated with praziquantel was found to be markedly reduced: 22%, compared with 83% cluster-containing wells of parasites from control mice. These results show that the outcome of infection and the suppression of taurocholate-induced clustering provide consistent indications of praziquantel toxicity against M. vogae, an observation confirmed by histological studies. The easily recorded clustering inhibition of M. vogae tetrathyridia in biphasic medium is a potentially useful system for the assessment of drug toxicity against cestode larvae.

Mesocestoides corti (syn. M. vogae): modulation of larval motility by neuropeptides, serotonin and acetylcholine.

The effect of platyhelminth FaRPs and selected classical neurotransmitters on the motility of intact Mesocestoides corti (syn. M. vogae) tetrathyridial larvae was studied in vitro using a micromotility meter. The effects of the test substances were temperature dependent and these were examined at 4, 23, 30 and 36 degrees C. At 36 degrees C all test substances had concentration-dependent excitatory effects, with thresholds for activity of: 100 nM (GNFFRFamide), 10 microm (YIRFamide), 30 microM (GYIRFamide), 100 nM (serotonin) and 100 microM (acetylcholine). At this temperature significant elevation of motility indices (MI) was recorded within 5 min of the addition of peptide or serotonin. The effect of acetylcholine was slower in onset and appeared 15-20 min post-addition. At 30 degrees C larval motility diminished more rapidly than that recorded at 36 degrees C, following the addition of 1 mM of each test substance. At 23 degrees C only serotonin (1 mM) significantly increased the MI, all other test substances having no apparent effect. Larval movement was completely arrested at 4 degrees C. The results demonstrate for the first time excitatory effects of platyhelminth neuropeptides and acetylcholine on muscle systems in cestode larvae. The fact that the only known cestode FaRP, GNFFRF amide, was more potent than any of the turbellarian FaRPs tested, suggests structural conservation of FaRPs and FaRP receptors within the cestodes.

In vitro culture of tetrathyridia of Mesocestoides corti using a gel based diphasic medium.

Tetrathyridia of Mesocestoides corti were cultured in vitro in a diphasic medium consisting of a liquid medium (CMRL Sigma) and a thixotropic nutrient gel (Oxoid). Tests demonstrated that a 50% medium/gel mixture produced optimum conditions for the survival and development of tetrathyridia. Established anthelminthic drugs were inoculated into the gel which demonstrated that this system can be used for preliminary anthelminthic drug screening. The development and survival of the tetrathyridia were influenced by the addition of pepsin, trypsin and liver peptone to the culture media. The development and maturation of proglottids were observed in addition to asexual reproduction by the process of budding. Tetrathyridia maintained in vitro and reinfected into both mouse and rat hosts retained their viability.

In vitro taurocholate-induced segmentation and clustering of Mesocestoides vogae (syn. corti) tetrathyridia (Cestoda)--inhibition byh cestocidal drugs.

Mesocestoides vogae (syn. M. corti) tetrathyridia were cultured in the presence of sodium taurocholate, for the purpose of exploring the suitability of this organism for the in vitro assay of cestocidal drugs. Parasite clustering and segmentation were observed as taurocholate-dependent effects in biphasic and monophasic media, respectively. Interestingly, representative members of two major classes of known cestocidal agents (namely, albendazole and praziquantel) blocked these effects. Furthermore, it was possible to determine a specific concentration of the drugs that inhibited clustering and segmentation (minimum inhibitory concentration). In contrast, no inhibition was obtained in the presence of anthelmintics without cestocidal activity. These observations open the way for further studies focused at understanding how the activity of the drugs is involved in the suppression of the taurocholate-induced effects.

Effect of praziquantel on the strobilar development of Mesocestoides corti in vitro.

The effect of praziquantel (PZQ) on the strobilar development of the cyclophyllidean cestode Mesocestoides corti was explored. Mesocestoides corti larvae were cultivated under conditions reported to favour their differentiation to the adult stage. Parasites were exposed to 0.1 microg ml(-1) PZQ for 16 h and subsequently transferred to drug-free medium. The ocurrence of segmentation--an early event of the larval somatic differentiation to the adult worm-- was considered as quantitative data. This phenomenon was evidenced earlier in worms transiently exposed to PZQ with respect to control cultures. Moreover, the rate of segmentation of drug-treated worms at the end of the experiment almost doubled that of control worms. To date, no similar effect on any cestode developmental process has been reported for an anthelmintic drug. In the light of the existing knowledge and understanding of PZQ mechanisms of action, the proposed experimental approach could contribute to the elucidation of pathways and mechanisms involved in cestode strobilar development.

An experimental, NADPH-diaphorase histochemical and immunocytochemical study of Mesocestoides vogae tetrathyridia.

In order to test the role of nitric oxide in flatworms, Mesocestoides vogae tetrathyridia were incubated together with L-arginine, which is the substrate for nitric oxide synthesis, or with NG-nitro-L-arginine, which is an irreversible inhibitor of nitric oxide synthase. Normally, tetrathyridia attach to each other with the aid of their suckers, forming clusters. The rate of cluster formation was followed during the incubations. L-Arginine stimulated, and NG-nitro-L-arginine clearly inhibited, the cluster formation. This is the first time that an effect of nitric oxide has been observed in a flatworm. In addition, the pattern of the NADPH-diaphorase histochemical reaction in the nervous system and the pattern of F-actin filaments in the musculature stained with TRITC-labelled phalloidin were studied. NADPH-d staining occurred in the brain and the main nerve cords but also followed the muscle fibres stained with phalloidin. The pattern of the NADPH-d reaction was compared with that of 5-HT immunoreactivity. The implications of the results are discussed in relation to the background of data on neuronal signal substances in M. vogae.

[The effect of anthelmintics on the serotonin content of cestodes]. / Effekt nekotorykh antigel'mintikov na soderzhanie serotonina u tsestod.

The effects of some anthelminthic agents, such as praziquantel, fenasal, albendazole, on the levels of the neurotransmitter serotonin in the tissues of Hymenolepis diminuta cestodes and Mesocestoides corti tetrathyridia were examined in vivo and in vitro. The finding showed lower serotonin levels in H.diminuta after praziquantel and albendazole and in M. corti after fenasal. It is suggested that the serotonin neurotransmitter system of helminths is involved in the mechanism of action of these agents.

Effects of free and liposomized praziquantel on the surface morphology and motility of Mesocestoides vogae tetrathyridia (syn. M. corti; Cestoda: Cyclophyllidea) in vitro.

The effects of in vitro exposure to praziquantel (PZQ), liposomized PZQ (lip.PZQ), and empty liposomes on the surface morphology and motility of Mesocestoides vogae tetrathyridia were investigated using scanning electron microscopy (SEM) and a motility apparatus. Examination of treated larvae showed an effect that was concentration- and time-dependent, involving morphological damage that was similar in character for all of the treated groups. The most marked effects were a flattening and elongation of the larval body accompanied by irregularities in the surface architecture involving the development of tegumental protuberances and depressions. Erosion of the surface microvillous layer occurred only after overnight incubation, being most pronounced after treatment with lip.PZQ. The motility index of treated tetrathyridia corresponded well to the SEM observations. The frequency of contractions was maximal in worms treated with free PZQ at 25 micrograms/ml in both regimens. However, after incubation with lip.PZQ the increase in motility was concentration-dependent and of a greater extent. Empty liposomes and lipid mixtures of the same concentration and composition resulted in increased motility in treated larvae as compared with controls. More extensive tegumental damage and higher motility of larvae occurred after lip.PZQ treatment, perhaps resulting from a synergistic action of the drug and its associated lipid.