Biochemical Liver Functions and Molecular Identification of Fasciola hepatica from Experimentally Infected Rat Model

The current study was performed to evaluate the liver function status as well as molecular characterization of the recovered worms in rats experimentally infected with F. hepatica. Sixteen male Wister rats aged 30 days were randomly allocated into two groups (n = 8). The first group was infected orally with 15 viable encysted metacercaria of F. hepatica per animal. The other group was kept non-infected (control group). At zero time (before infection), the 2nd, 4th, 6th, 8th, 10th, 12th and 14th weeks post-infection (WPI), blood and serum samples were collected via puncture of retro-orbital plexus of veins from each rat. Serum enzyme level (AST and ALT) and total protein were measured, and the serum protein profile was carried out using agarose gel electrophoresis. During the period of the experiment, serum ALT and AST activities and serum total globulins significantly increased while serum total proteins and albumin markedly decreased in the infected group. On the 14th WPI, the data of the electropherogram showed that globulin fractions (α1-, β- and γ-globulin) levels were significantly increased while α2-globulin was markedly decreased in the infected group. The molecular analysis confirmed the amplification of the ITS1, ITS2 and NDI genes of F. hepatica recovered from the infected liver of rats with amplicon sizes of 630, 510 and 560 bp, respectively. Sequencing of the amplified ITS gene resulted in the determination of 3 strains (PP108836, PP108837, and PP108838). Also, analysis of the ITS2 gene resulted in obtaining 3 isolates under the accession numbers (PP109065, PP109066, and PP109067). In conclusion, fasciolosis in the rat model is suitable for routine experimental infections and caused a pronounced liver dysfunction with discharging of the Fasciola eggs in the faeces and the development of adult stages in the bile ducts. Furthermore, molecular techniques are a sensitive tool for the identification and characterisation of the Fasciola parasite.

Capillaria hepatica in China

Capillaria hepatica(C.hepatica) is a parasitic nematode causing hepatic capillariasis in numerous mammals.Ecologic studies showed that the first hosts of C.hepatica were rodents,among which rats had relatively high infection rates,which explains why C.hepatica spreads globally.Anatomical studies showed that the liver was the principal site of colonization by these parasites and physical damage tended to occur.Although C.hepatica might lead to serious liver disorders,relevant clinical reports were rare,because of the non-specific nature of clinical symptoms,leading to misdiagnosis.This review mainly focuses on the biological characteristics and epidemiology of C.hepatica in China and histopathologic changes in the liver,with expectation of gaining a better understanding of the disease and seeking more effective treatment.

Changes of the Levels of Blood NO and TNF-αConcentrations in Water Buffaloes and Goats Infected with Fasciola hepatica

Changes of concentrations were studied in water buffaloes and goats infected with Fasciola he-patica on blood NO(nitric oxide) and TNF-α(tumor necrosis factor-α). Twenty healthy male castrated water buffaloes of 2 - 3 years old and weighing 300 - 500 kg as well as six goats were confirmed free of fasciolosis by fecal examination and Dot-ELISA. Two studies were conducted using the water buffaloes. In the first experiment, 8 water buffaloes were randomly divided into control group (n = 3) and infection group (n = 5). Each buffalo in the infected group received orally 60 metacercariae of F. hepatica per day for 20 days (total 1 200 metacercariae) to produce a chronic infection. In the second experiment, 12 water buffaloes were randomly divided into infected (n=9) and control group (n = 3). Each buffalo in the infected group was given a single oral dose of 1 600 metacercariae to produce an acute infection. The 6 goats were randomly divided into two infected groups and a control group. The sheep in two infections received a single oral dose of 200 and 500 metacercariae respectively, the control group remained uninfected. Blood NO and TNF-a concentrations of the test animals were measured by a reductive enzyme assay and RIA, respectively. Blood NO concentration in both acutely and chronically infected water buffaloes progressively increased from week 3 post-infection and was significantly greater than that of the control group (P<0. 05) at the 5th week (acute infection) and 7th week (chronic infection), and remaining at higher concentration for the remaining period of the studies. Blood TNF-αconcentrations in both chronically and acutely infected water buffaloes also increased after infection. In the goat experiment, plasma NO concentrations in both infection groups increased from week 3 after infection, and remained higher than that of the control group until the end of the experiment. TNF-α concentrations in goats in infection group 1 and 2 gradually increased after infection and were significantly greater than those of the control group from the 9th to 11th week and from the 11th to 15th week respectively.

Fascioliasis presenting as colon cancer liver metastasis on 18Ffluorodeoxyglucose positron emission tomography/computed tomography: A case report

BACKGROUND Fascioliasis is caused by watercress and similar freshwater plants or drinking water or beverages contaminated with metacercariae. Fascioliasis can radiologically mimic many primary or metastatic liver tumors. Herein, we aimed to present the treatment process of a patient with fascioliasis mimicking colon cancer liver metastasis. CASE SUMMARY A 35-year-old woman who underwent right hemicolectomy due to cecum cancer was referred to our clinic for management of colon cancer liver metastasis. Both computed tomography and 18F-fluorodeoxyglucose positron emission tomography revealed several tumoral lesions localized in the right lobe of the liver. After a 6-course FOLFOX (folinic acid, fluorouracil, oxaliplatin) and bevacizumab regimen, the hypermetabolic state on both liver and abdominal lymph nodes continued, and chemotherapy was extended to a 12-course regimen. The patient was referred to our institute when the liver lesions were detected to be larger on dynamic liver magnetic resonance imaging 6 weeks after completion of chemotherapy. Right hepatectomy was performed, and histopathological examination was compatible with fascioliasis. Fasciola hepatica IgG enzyme-linked immunosorbent assay was positive. The patient was administered two doses of triclabendazole (10 mg/kg/dose) 24 h apart. During the follow-up period, dilatation was detected in the common bile duct, and Fasciola parasites were extracted from the common bile duct by endoscopic retrograde cholangiopancreatography (ERCP). Triclabendazole was administered to the patient after ERCP. CONCLUSION Parasitic diseases, such as those caused by Fasciola hepatica, should be kept in mind in the differential diagnosis of primary or metastatic liver tumors, such as colorectal cancer liver metastasis, in patients living in endemic areas.

<i>In Vitro</i>Evaluation of Fasciolicide Activity with Hexane, Methanol and Ethyl Acetate with Extracts Processed and Obtained from Some Mexican Plants Used in Traditional Medicine Based on Ethno Botanical Studies

Fasciolosis is a parasitic disease of world distribution affecting mainly domestic ruminants. The control of this disease is carried out using chemical fasciolicides which, in some cases, have been observed to have environmental problem such us pollution resistance. An alternative is to investigate extracts from plants with anti-Fasciola hepatica effects, taking advantage of the great diversity of flora of our country. The aim of this paper is to identify, assess and elucidate the anti-Fasciola hepatica effect in vitro using antiparasitic extracts of some plants used in Mexican ethno botany. We collected, dried, processed and tested in vitro about 20 plants with their respective chemical elements (hexane, ethyl acetate and methanol), obtaining results of a total of 60 extracts tested. The in vitro evaluations were carried out for 3 days, and the efficacy of each extract was compared with an untreated control group. Each test was repeated 6 times with 13 extracts that showed greater fasciolocide activity. Results from these 13 extracts tested ranged from 80% to 100% activity and the plants tested were: Castela tortuosa (chaparro amargo), Achillea millefolium (plumajillo), Thymus vulgaris (thyme), Justicia spicigera (muicle), Limpia critridora (cedron), Populus alba (Alamo), Mentha piperita (mint), Chenopodium graveolens (epazote de zorrillo), Lippia graveolens (oregano), Artemisia mexicana (estafiate) and Artemisia absinthium (wormwood), which is the hexane variable which showed higher fasciolicide capacity;using a dose of 500 gr/Lt in all the trials. Further in vitro studies should be conducted to obtain the LD50 of each extract to be able to isolate the main active element found in the hexane variable.

Study on Blood Cell Immune Response in Water Buffaloes Infected Acutely with F. hepatica

Action mechanism of blood cell immune response in water buffaloes against acute infection with F. Hepatica was studied. The results showed that after water buffaloes were infected, the total levels of WBC surpassed control group during whole infection period; Eosinophiles (%) of DC were higher than control group at the 2nd week until 19th week, and then dropped and was close to control group; Neutrophiles(%) was low or significantly lower than control group within the 5 - 16th weeks; The total levels of lymphocytes (%) was lower than control group during the whole infection period; T-lymphocytes (%) dropped significantly, but B-lymphocytes(%) had opposite changes from the first week of infection, and they were close to the control group after 11 weeks; RBC-CR] and RBC-IC rosette rates dropped and rose during 2 - 16 and 2-18 weeks, respectively, and then approached the same between both groups. It was suggested that the violent changes of specific and nonspecific immune responses in water buffaloes with acute F. hepatica infection are related with the mechanism against infection with F. hepatica together.

Obtaining the Minimum Lethal Dose against <i>Fasciola hepatica in Vitro</i>Using Plant Extract Hexanes with Fasciolicide Activity and Toxicity Evaluation on CD1 Male Mice

Fascioliasis is a parasitic disease of worldwide distribution affecting mainly cattle and sheep. Its importance lies in the economic losses it produces in the livestock industry. Its control is carried out by using a chemical fasciolicide showing resistance problems and environmental contamination. Looking for an alternative control for this disease the present study was aimed at determining the hexane anti-Fasciola hepatica in the in vitro effect of some plant extracts and the minimum lethal dose of the mentioned extracts. All selected plants were tested in vitro at concentrations of 500, 250, 125 and 50 mg/L):Achilleamillefolium (plumajillo), Artemisiaabsinthium (wormwood), Artemisia mexicana (estafiate), Castelatortuousa (chaparroamargo), Chenopodiumgraveolens (epazote de zorrillo), Gymnospermaglutinosum (popote) Justicia spicigera (muicle), Limpia critridora (cedron), Lippiagraveolens (oregano), Menthapiperita (Mint), Populus alba (alamo) and Thymusvulgaris (thyme). Subsequently proceeded to perform a toxicity study with these fractions in CD1 male mice 10-13 weeks of age, forming groups of 3-5 animals they were administered a single oral dose being (5 mg/kg, 50 mg/kg, 500 mg/kg, 2500 mg/kg and 5000 mg/kg) and were kept under observation 20 days, later were sacrificed and a kidney and liver histology was performed, finding the safety of the extracts. To perform the toxicity study with these fractions, groups of five CD1 male-mice were formed, they were treated with oral doses of 5, 50, 500, 2500 and 5000 mg/kg, administered with a cannule. All mice were kept under observation for 20 days. Finally they were sacrificed to perform histology of the kidney and liver in search of possible side effects. Results show that none of the extracts exhibited that fasciolocide activity for mice CD1 even at the highest dose thereforefinding the safety of the extracts.

Evaluation of the Fasciolicidal/Nematicidal Efficacy of an Intramuscular Combination of Clorsulon/Levamisole in Naturally Infected Dairy Cattle

The aim of the present study was to assess the fasciolicidal and nematicidal efficacy of a new combination of clorsulon/levamisole, which was formulated for intramuscular administration, in dairy cattle. For the study, sixteen Holstein-Freisian dairy cows were selected for inclusion in the trial based on finding Fasciola hepatica eggs or gastrointestinal nematodes (GIN) in the feces. Ten of these cows were naturally infected with F. hepatica, and 6 were naturally infected with GIN. On Day 0 (zero), all animals were treated with a combination of clorsulon (2.5 mg/kg/IM and levamisole (2.5 mg/kg/IM) administered at the dose recommended by the manufacturer (1 ml/20kg/bw). Subsequently, their feces were analyzed on days 0, 7, 14 and 21 post treatments by the Flukefinder sedimentation method and McMaster technique to determine the percentages of egg reduction in the animals, with reference to the egg load count on day 0. The results obtained in the current work indicated an efficacy of 100% that was determined by the percentage reduction of eggs of F. hepatica and/or GIN. This new fasciolicide/nematicide formulation exerted a remarkable effect and can be considered a new alternative to control F. hepatica and gastrointestinal worms in dairy cattle.

Experimentelle Embolisation unterschiedlicher hepatischer Gefaβe beim Hund mit Bletilla striata

Diese vergleichende Untersuchung uber Embolisation differenter hepatischer GefaBe bei Hunden mit Bletilla striata und Gelfoam ergab,d aB letilla striala als Embolisationsmaterial bessere Erfolge zeigt als Gelfoam.Die Mechanismen der Embolisation mit Bletilla striata sind folgende :1)Bletilla striata wird nicht leicht von Geweben absorbiert;2)Bletilla striata bewirkt mechanische Blockierung;3)Bletilla striata beeinflt das Gerinnungssystem und dasAntikoagulationssystem;4)Bletilla siriata kann die Geafwandschadigen und dadurch sekundare Blockierung auslosen.

Effectiveness of the Experimental Fosfatriclaben in Comparison with Two Commercial Fasciolicides in Cattle

The aim of the present study was to compare the fasciolicidal efficacy in cattle of an experimental water-soluble compound, named fosfatriclaben, a prodrug of triclabendazole, with two commercial fasciolicides. For this, twenty-four cross mixed milking cows, aged between 1 to 3 years, naturally infected with Fasciola hepatica, were used. To apply the treatment, animals were divided into 4 groups of 6 animals each being ranked according to their fecal fluke-egg count from the highest to the lowest number of eggs. The compounds were administered in the groups with a single dose. Group 1 (G1) was treated with 6 mg/kg/IM of fosfatriclaben, G2 was treated with triclabendazole at 12 mg/kg/PO;G3 was treated with 5% closantel at 10 mg/kg/SC and G4 was administered with 5 mL/IM of deionized water and served as untreated control. Fecal samples were analyzed on days—8, 0 (treatment day), 7, 14, 21 and 28 to count the number of fasciola eggs before and after the treatment using the sedimentation test. The efficacy was assessed as the percentage of fluke-egg reduction on treated groups relative to the untreated control. The resulting data were analyzed by the Kruskal-Wallis non-parametric test, using the statistical IBM SPSS 24.0 (2016) program. Results indicated a fluke-egg reduction of 100% for fosfatriclaben along the study;triclabendazole showed an efficacy of 99.7% - 100% from day 7 to day 28. In the case of closantel, values from 98.8% to 99.6% were recorded from day 7 to day 28. No statistical differences between treatments were observed (P < 0.05). It was concluded that the experimental fosfatriclaben administered at 6 mg/kg/IM to naturally infected cattle exerted a fasciolicidal efficacy similar to the drugs of choice for the treatment of fasciolosis (triclabendazole) when administered at 12 mg/kg/ PO, and closantel injected at 10 mg/kg/SC.

Trait plasticity, not values, best corresponds with woodland plant success in novel and manipulated habitats

Aims The clustering of plants with similar leaf traits along environmental gradients may arise from adaptation as well as acclimation to het-erogeneous habitat conditions.Determining the forces that shape plant leaf traits requires both linking variation in trait morphology with abiotic gradients and linking that trait variation with plant per-formance under varying abiotic conditions.across the spectrum of plant types,shade-tolerant evergreen herbs are relatively low in trait plasticity,compared to deciduous and sun-adapted species.These plants employ stress-tolerant strategies for survival,which coincide with relatively static trait morphologies,slow growth and hence a lower ability to adjust to changing environmental conditions.Methods We investigate how the survival of two ecologically similar under-story evergreen species,Asarum arifolium and Hepatica nobilis,cor-responds with variation in six commonly measured functional traits(leaf area,specific leaf area,plant height,leaf number,leaf length and shoot mass)along natural and experimental abiotic gradients.We examine temporal(the period 2007-9)and spatial(100 km)variations in these traits after(i)translocating 576 plants across a span from the southerm Appalachian Mountains in NC,USA,to the Piedmont,GA,USA,which includes north-and south-facing slope habitats and(ii)the experimental manipulation of diffuse light and soil moisture.Important findings We find that when translocated into a novel habitats,with novel environmental conditions that often are more extreme than the source habitat,both species appear capable of considerable mor-phological acclimation and generally converge to similar trait values.Hepatica nobilis does not exhibit mean trait values particu-larly different from those of A.arifolium,but it demonstrates much greater phenotypic plasticity.These results indicate that relatively conservative plant species nonetheless acclimate and survive across heterogeneous environmental conditions.

Fasciola hepatica in a country of low incidence: a tricky diagnosis

Background:Fasciola hepatica is a foodborne trematode present worldwide.Definitive hosts are mostly ruminants such as cattle and sheep,as well as humans.In Switzerland,Fasciola infection in humans is rare.Unfortunately,many patients are likely to undergo multiple unnecessary investigations before the parasite is suspected and fascioliasis diagnosed,especially if symptoms are unspecific.Methods:Retrospective analysis of all patients diagnosed with Fasciola hepatica at the University Hospital of Bern between 2005 and 2018.Diagnosis was positive if a positive serology and/or eggs in stool samples correlated with clinical presentation(symptoms and/or imaging).Patients were excluded if serology was weakly positive and another diagnosis more likely.Personal data,laboratory results,imaging,proposed treatment and outcome were collected from patient files.Results:Sixty patients had a positive serology during this time period.Forty-seven of them had a more plausible alternative diagnosis and were not included in the study,leaving 13 patients for analyses;46.2%(6/13)were male,mean age was 45.8 years old(range,17-80 years old).Four patients(4/13,30.8%)were asymptomatic,nine(9/13,69.2%)presented with symptoms ranging from right upper quadrant abdominal pain(44.4%)and generalized pruritus(33.3%)to weight loss and night sweats(33.3%).The mean duration of symptoms until correct diagnosis was 8.9 months(range,1-48 months).Five patients(5/13,38.5%)had documented eosinophilia,four(4/13,30.8%)elevated liver enzymes and seven(7/13,53.8%)elevated cholestasis parameters.Mean antibody level on serology was 88 AU/mL(range,3-134 AU/mL).Ultrasound was used most frequently(7/13,53.8%),followed by magnetic resonance imaging(4/13,30.8%),computed tomography and endoscopic retrograde cholangiopancreatography(3/13,23.1%).The most common findings were bile duct dilatation,followed by hepatic lesions.Treatment consisted of Triclabendazole 10 mg/Kg.One patient needed a second treatment course for persistent disease.There were no recurrences.Conclusions:With a low incidence of Fasciola hepatica in Switzerland,correct diagnosis is often substantially delayed.Raising awareness among Swiss physicians is paramount,and a higher level of suspicion necessary when confronted with unspecific symptoms or liver imaging,thus avoiding a long delay in diagnosis,as well as unnecessary tests.