Rumen Fluke Research Articles

Helminth co-infections have no additive detrimental impact on milk yield and milk quality compared to mono-infections in German dairy cows

BackgroundInfections with (tricho-)strongyles, Dictyocaulus viviparus or Fasciola hepatica have been shown to reduce milk production in dairy cows. However, the current published studies focused on one single helminth infection by neglecting helminth co-infections and their possible (additive) effects on host performance. Hence, for the first time, we investigated differences in the impact of patent helminth co-infections versus mono-infections on milk production parameters in individual cows.MethodsA total of 1583 dairy cows from 27 herds were included in this study. Faecal samples were examined in 2015 and 2021/2022 to determine the number of eggs/larvae per gram faeces for (tricho-)strongyles, D. viviparus, F. hepatica and rumen flukes. The cows were classified as non-infected, mono-infected and co-infected. Linear mixed models were applied to analyse the association between infection status (non-infected vs. mono-infected vs. co-infected) with milk yield, milk protein and milk fat content by including potential confounders.ResultsInfections with (tricho-)strongyles, D. viviparus, F. hepatica and rumen flukes were detected in 100%, 28.6%, 50.0% and 21.4% of herds, and 27.4%, 2.6%, 10.8% and 0.8% of faecal samples in 2015, while 100%, 0.0%, 86.7% and 60.0% of herds and 52.3%, 0.0%, 13.3% and 26.8% of faecal samples were positive in 2021/2022. Co-infections with two or more helminth taxa were detected in 74.4% of herds and 5.0% of faecal samples in 2015, and in 93.3% of herds and 21.7% of faecal samples in 2021/2022. The correlations between strongyle EPG, D. viviparus LPG and F. hepatica EPG were significantly positive in 2015. Significantly higher mean EPGs were identified in 2015 in faecal samples presenting co-infections with F. hepatica and one or two other helminth taxa than in faecal samples presenting F. hepatica mono-infections (P = 0.013). Although expected, the infection status (mono- or co-infected) had no significant impact on milk yield, milk protein and milk fat content in the linear mixed model analyses based on individual faecal examinations.ConclusionsPatent helminth co-infections had no additive detrimental impact on milk production parameters in the present study. This might be a result of presumably low worm burdens, but should be confirmed in future studies.Graphical

Multi‐Environment Quantification of Parasite and Intermediate Host DNA on Pasture for Fine‐Scale Disease Risk Assessment

ABSTRACTParasite transmission occurs in complex environments comprising multiple matrices. Trematode parasites of ruminant livestock such as the liver fluke, Fasciola hepatica and the rumen fluke, Calicophoron daubneyi, show affinity with freshwater environments shared with their amphibious snail intermediate host, Galba truncatula. Isolation of environmental DNA (eDNA) from these parasites and their snail hosts in water draining from grazing land provides opportunities for improved molecular diagnostic detection and can help identify infection risks at farm level. The detection and quantification of eDNA from other environmental matrices has received less attention but would improve the understanding of parasite dynamics on pasture. Our study has considerably extended eDNA sampling methods for the detection of parasitic trematodes of ruminant livestock and their snail intermediate host by including for the first time the analysis of soil and herbage environmental samples alongside water collections. A droplet digital PCR (ddPCR) workflow was developed to detect parasite and snail eDNA from soil, herbage, and water collected from livestock farms. For the first time, C. daubneyi eDNA was isolated from agricultural soil alongside water samples and G. truncatula eDNA was detected in water, soil, and herbage samples. No environmental samples were positive for F. hepatica eDNA. Assessing multiple environmental matrices increased the number of positive sites. Future implementation of eDNA detection methods alongside traditional parasite diagnostics can underpin more holistic evaluations of the environmental components of parasite epidemiology and facilitate adaptation to changing disease patterns.

Morphological and molecular data confirm the occurrence of Paramphistomum leydeni (Trematoda: Paramphistomidae) in ruminants from Southern Brazil

Species belonging to the family Paramphistomidae Fischoeder, 1901, commonly known as “rumen flukes”, are a group of parasites frequently related to Brazilian livestock production. They inhabit the digestive tract of ruminants and have recognized pathogenicity during the early stages of infection, which can be responsible for economic losses. These trematodes are often associated with Southern Brazil, a region heavily focused on animal farming, which also makes it ideal for the life cycle of paramphistomes. Despite their aforementioned importance, studies regarding their distribution, molecular taxonomy and biology are still scarce in the country. In the present study, rumen flukes collected from cattle (n = 22) and sheep (n = 3) from 9 batches of ruminants from the cities of Jaguarão, Pelotas and Rio Grande, state of Rio Grande do Sul, Brazil, between May and July 2022, were subjected to morphological and molecular study. The microscopic analysis of histological and manual cuts revealed diagnostical traits compatible with Paramphistomum leydeni Näsmark, 1937, including the presence of tegumental papillae, pharynx of the liorchis type and acetabulum of the leydeni type. Molecular data corroborated the morphological identification, with ITS-2 and cox-1 sequences here obtained presenting 100% and 96.8–99.8% similarity, respectively, to P. leydeni samples previously characterized in different countries from Asia, Europe, and South America. Intensity of infection ranged from 5 to 458 and 1 to3 specimens of P. leydeni in sampled cattle and sheep, respectively. The present study contributes to a better understanding of the taxonomy of the flukes involved in cattle and sheep paramphistomosis in Brazil, suggesting that P. leydeni could be the main paramphistome species found in ruminants in the studied region.

Epidemiology of rumen fluke infection in selected buffalo farms in perak, malaysia: prevalence, molecular species identification, and associated risk factors.

Rumen flukes cause heavy economic losses in the ruminant industry worldwide, especially in tropical and subtropical countries. This study estimated the prevalence of rumen flukes in buffaloes, identified the species diversity, and determined risk factors associated with rumen fluke prevalence in Perak, Peninsular Malaysia. A cross-sectional study was conducted, and 321 faecal samples were collected from six buffalo farms. A structured questionnaire was developed, and farmers were interviewed to obtain information regarding risk factors associated with rumen fluke infection. The faecal samples were examined using sedimentation and Flukefinder® techniques. Genomic DNA was extracted from the fluke eggs recovered using the Flukefinder® method, and the internal transcribed spacer 2 (ITS2) fragment was amplified and sequenced to facilitate species identification. The results showed that the overall prevalence of rumen fluke across the sampled farms was 40.2% (129/321). Three rumen fluke species were identified, namely, Fischoederius elongatus, F. cobboldi, and Orthocoelium streptocoelium. Several management factors had a significant association (P < 0.05) with rumen fluke prevalence, including production type, cleaning of the stable, drinking water system, flooding around the farm, grazing system, pasture sharing with other livestock, and deworming program. This work constitutes the first attempt to understand the epidemiology of rumen fluke infection in the region and suggests that good farm management, pasture management, choosing appropriate drugs, and proper husbandry practices may improve buffalo health and production in areas where rumen flukes are prevalent.

Calicophoron daubneyi (Paramphistomidae) in deer of the Šumava National Park, Czech Republic – Consequence of prevalent rumen fluke infection in cattle

A substantial parallel increase in prevalence and geographical spread of the rumen fluke, Calicophoron daubneyi, in livestock in western and central Europe has been recognized in the recent past.In the course of the examination of rectum feces of 471 red deer (Cervus elaphus) and one sika deer (Cervus nippon) from the Fascioloides magna endemic Šumava National Park in the years 2021 and 2022, rumen fluke eggs were detected in four red deer (0.8%) and the sika deer and identified as eggs of C. daubneyi by molecular analysis. Subsequent examination of rectal fecal samples of 247 beef cattle from 22 herds of 14 farms located in or nearby the national park revealed rumen fluke eggs in 53 samples (21.5%) originating from 16 herds of 11 farms, molecularly identified as C. daubneyi eggs as well. One C. daubneyi egg positive red deer and three C. daubneyi egg positive cattle samples also contained fasciolid eggs, respectively, which were detected in 9.5% or 3.6% of the total samples from red deer or cattle, respectively.Results of this investigation reveal the first finding of C. daubneyi in sika deer worldwide and in red deer in mainland Europe and add to the growing number of reports on C. daubneyi in livestock in Europe. Considering that the ratio of cattle excreting rumen fluke eggs exceeded that of deer substantially, it can reasonably be assumed that the C. daubneyi infections in deer are a consequence of the prevalent infection in cattle, illustrating a pathogen spillover event from livestock into wildlife.

Tegumental surface change in Paramphistomum epiclitum caused by Bombax ceiba flowers and black pepper seed extract

Rumen flukes, parasites of the superfamily Paramphistomoidea, are found in cattle rumen. Heavy infections can cause symptoms such as diarrhea, weight loss, and poor body condition, resulting in a decrease in milk and meat production. This study compares the tegumental surface change of Paramphistomum epiclitum as a response to ethanolic extracts of Bombax ceiba flowers and black pepper seeds. Adult flukes were subjected to various concentrations of crude extracts, including 12.5, 25, 50, 100, and 200 μg/mL for 12, 18, and 24 h incubation. Scanning electron microscopy (SEM) exhibited that the ethanolic extracts of both Bombax ceiba flowers and black pepper seeds caused tegumental surface changes in adult P. epiclitum. Based on the results, Bombax ceiba flower extract has anthelmintic activity, compared with black pepper seed extract, towards adult P. epiclitum due to the deformation of the tegument at lower concentrations than black pepper extract.

Visualization and development of colorimetric loop-mediated isothermal amplification for the rapid detection and diagnosis of paramphistome infection: colorimetric PAR-LAMP.

Colorimetric detection can be applied to differentiate between positive and negative conditions. It can be coupled with loop-mediated isothermal amplification to diagnose rumen fluke or paramphistome infection, also called colorimetric PAR-LAMP. This study conducted LAMP using three candidate indicator dyes, namely malachite green (MLG), methyl green (MTG), and neutral red (NTR), and the results were observed by the naked eye. The dye concentration was optimized to obtain the most pronounced positive-negative result discrimination. Subsequently, we conducted target sensitivity tests using the DNA of Fischoederius elongatus at different concentrations. To validate the detection accuracy, the result was confirmed by gel electrophoresis. The sensitivity test presented the lowest detectable DNA concentration or limit of detection (LOD), with 1pg for MLG, 0.5ng for MTG, and 50pg for NTR. Different LODs revealed inhibition of LAMP reaction and reduced efficiency of result presentation for colorimetric-based detection, particularly NTR and MTG. For MLG-LAMP, we observed no cross-reaction of non-target DNA and improved reaction with the DNA of Fischoederius cobboldi and Calicophoron sp., with multi-detection. In addition, naked eye observation and agarose gel electrophoresis (AGE) evaluation of the MLG-LAMP results showed a moderate and strong agreement with LAMP-AGE and microscopic examinations. Based on our results, colorimetric PAR-LAMP is a rapid, comfortable, and point-of-care procedure for the diagnosis of paramphistome infection.

Assessing the microbiota of the snail intermediate host of trematodes, Galba truncatula

BackgroundThe microbiome is known to play key roles in health and disease, including host susceptibility to parasite infections. The freshwater snail Galba truncatula is the intermediate host for many trematode species, including the liver and rumen flukes Fasciola hepatica and Calicophoron daubneyi, respectively. The snail-parasite system has previously been investigated. However, the specific interaction between the snail-associated microbiota and intra-snail developmental stages of trematodes has yet to be explored.MethodsGalba truncatula snails were collected from farms in Northern Ireland and trematode infection was diagnosed using PCR. High-throughput sequencing analysis of the bacterial 16S ribosomal DNA V3-V4 hypervariable regions was subsequently applied to characterise the microbiota of both uninfected and infected snails.Results We first showed that the snail harboured microbiota that was distinct for its environment. The microbiota of infected snails was found to differ significantly from that of uninfected snails. In particular, the bacterial genera Mycoplasma and Methylotenera were significantly more abundant in infected snails, while genera Sphingomonas and Nocardioides were predominantly associated with uninfected snails. ConclusionThese findings pave the way to future studies on the functional roles of bacteria in host-parasite relationships.Graphical

Prevalence of Gastrointestinal Parasites in Free-Ranging Bantengs (Bos javanicus) and Domestic Cattle at a Wildlife and Livestock Interface in Thailand

Impact of human activities on conservation of endangered species is significant due to the creation of wildlife-livestock interfaces (WLI), which increases the likelihood of disease transmissions. Protected wildlife populations may suffer from diseases caused by gastrointestinal (GI) parasites, potentially impacting conservation efforts. A population of ~100 wild bantengs inhabits the Lam-Pao non-hunting area, which is adjacent to human communities, animal farms and the Pao River, making this WLI suitable for a study of prevalence of GI parasites in the bantengs and domestic cattle. This research examined the prevalence and diversity of GI parasites and explored the associations between the parasitic species identified in the 2 animal groups. Formalin ethyl - acetate concentration technique (FECT) identified overall prevalence of 26.2, 17.4, 53.3 % for both animal groups, bantengs and domestic cattle respectively. The following prevalence of parasitic eggs were identified in the bantengs (N = 46) and the domestic cattle (N = 15) respectively: 1) 8.7 and 46 % of strongyle-type; 2) 4.3 and 6.7 % of Moniezia spp.; 3) 2.2 and 0 % of Fasciola spp.; 4) 6.5 and 6.7 % of Rumen flukes; 5) 0 and 13.3 % of Toxocara spp.; 6) 0 and 6.7 % of Capillaria spp.; 7) 0 and 6.7 % of Strongyloides spp.; and 8) 0 and 6.7 % of Trichuris spp. The combination of coproculture and polymerase chain reactions (PCR) was used to identified common strongyle-type species. We determined Oesophagostomum radiatum in 2 cattle (13.3 %) and 3 (6.5 %) banteng samples, Cooperia onchophora in 1 (6.7 %) cattle sample and Ostertagia ostertagi in 1 (6.7 %) cattle sample. The results present in this study would enable management to proactively strategize and prepare for parasitic diseases that can endanger such a population. HIGHLIGHTS FECT indicated that strongyle-type eggs were the most observed in bantengs and domestic cattle. Overall prevalence of 26.2, 17.4 and 53.3 % for bantengs, domestic cattle and both groups combined were reported, which were relatively low. The prevalence of strongyle-type and Toxocara eggs differed statistically between the 2 groups, potentially due to farming, leading to re-infections and high prevalence in domestic animals. GRAPHICAL ABSTRACT

In vitro anthelmintic activity of fresh fruit juice of Solanum verbascifolium Linn. against digenetic trematodes

Abstract Background: The substantial costs of conventional anthelmintics, the presence of drug residues in animal products, and the increasing prevalence of anthelmintic-resistant strains have spurred research on medicinal plants as alternative anthelmintic sources. This study evaluates the anthelmintic activity of Solanum verbascifolium Linn. fruit juice against digenetic trematodes. Materials and Methods: This study involves testing the efficacy of S. verbascifolium fruit juice at varying concentrations (10, 50, and 100 mg/mL) on five groups of rumen fluke worms. Albendazole (10 mg/mL) was used as the positive control, while distilled water was used as the negative control. Rumen flukes, digenetic trematodes found in the cow’s gut, were selected to simulate worms commonly found in the human gut. Results: The results indicated a statistically significant association between the different concentrations of the fruit juice and their anthelmintic activity. All concentrations of S. verbascifolium fruit juice demonstrated anthelmintic properties. Notably, the 50 and 100 mg/mL concentrations exhibited a significant effect on the worms compared to the standard drug albendazole. Conclusion: This study concludes that the fruit juice of S. verbascifolium possesses promising anthelmintic properties in vitro. These findings indicate the potential use of the botanical as an anthelmintic drug, warranting further investigation and development.

Relationship between number of adult rumen fluke (Calicophoron microbothrioides) and eggs per gram of feces in culled dairy cattle in Peru: A pilot study

Background: Paramphistomosis, a parasitic condition caused by Calicophoron microbothrioides in domestic ruminants, has garnered limited attention among cattle breeders in the Cajamarca Valley, Peru. Despite its status as an endemic affliction with considerable risk, scant investigations have been conducted on this parasite within this region. Objective: This study aimed to assess the correlation between the population of adult parasites (referred to as “number of adult parasites” or NAP) in rumen and reticulum of naturally infected cows and the number of eggs per gram of feces (EPG). Methods: A sample of twenty-two cattle harboring adult parasites in their rumen and reticulum was selected for analysis. Fecal samples were collected from these animals to establish a correlation between NAP and EPG. Data analysis included linear regression, the Shapiro-Wilk normality test, and the nonparametric Mann-Whitney test. Results: The study revealed a remarkably strong and directly proportional association between the number of adult parasites in rumen and reticulum and the coproparazitologic EPG (r = 0.971). Conclusion: According to the linear model (NAP = 60.381 ± 3.37; EPG - 22.979) increasing EPG by one lead to an increase by 60 NAP in rumen and reticulum (p&lt;0.01).

First Molecular Identification of Calicophoron daubneyi (Dinnik, 1962) and Paramphistomum leydeni (Nasmark, 1937) in Wild Ruminants from Romania.

Rumen flukes are geographically widespread trematodes affecting wild and domestic ruminants. The juvenile forms, which are found in the small intestine, are more pathogenic compared to the adults. Severe diarrhoea and weight loss are the major clinical signs, and the disease might be fatal in severely infested individuals. In the last decade, paramphistomosis has been described as an emerging parasitic disease in Europe. This study aimed to identify the rumen fluke species in wild ruminants from western Romania. Fifty-two pre-stomachs obtained from roe deer (Capreolus capreolus) that were hunted on 14 hunting grounds from Timiș and Arad counties were examined for the presence of paramphistomes. Three (9.09%) out of 33 samples were positive in Timiș County, and one (5.26%) out of 19 samples was positive in Arad County. Subsequent PCR testing revealed that three samples were positive for Calicophoron daubneyi and one for Paramphistomum leydeni. The presence of C. daubneyi and P. leydeni in roe deer has not been previously reported in Romania. Two Paramphistomum species-C. daubneyi and P. leydeni-were revealed as the main species of rumen flukes in roe deer from forests in Romania.

Species identity and phylogeny of Paramphistomoidea Fischoeder, 1901 occurring in cattle and sheep in North Cameroon.

Paramphistomidae and Gastrothylacidae are parasitic flatworms occurring in wild and domestic ruminants in different parts of the world especially in Asia and Africa. In Central Africa, few studies have been done using molecular techniques to resolve taxonomical groupings and understand the epizootiology of these parasites. In this study, we molecularly characterized two hundred adult flukes collected from the fore stomachs of cattle and sheep in the Adamawa region of the northern Cameroon. PCR and sequencing of the nuclear ITS-2 of the ribosomal DNA gene and a portion of the mitochondrial cox-1 locus revealed the presence of at least nine species belonging to the genera of Cotylophoron, Calicophorn, Orthocoelium and Carmyerius. In Zebu cattle, we identified Ca. microbothrium, Ca. clavula, Ca. phillerouxi, Co. cotylophorum, Co. fuelleborni, O. scoliocoelium, Car. gregarius, Car. graberi and Car. mancupatus and one yet unknown Paramphistomoidea sp, whereas in sheep, only Ca. microbothrium was found. The present study also strongly suggests cross-hybridization between the two Cotylophoron species coexisting in cattle. These results have implications for the diagnosis and control of rumen flukes in the region and point to the need for accurate species identification to understand parasite distribution and population genetics.

Treatment of rumen fluke in cattle and sheep.

Treatment of rumen fluke in cattle and sheep.

Comparison of Mini-FLOTAC, Flukefinder® and sedimentation techniques for detection and quantification of Fasciola hepatica and Calicophoron daubneyi eggs using spiked and naturally infected bovine faecal samples

BackgroundFasciolosis (Fasciola hepatica) and paramphistomosis (Calicophoron daubneyi) are two important infections of livestock. Calicophoron daubneyi is the predominant Paramphistomidae species in Europe, and its prevalence has increased in the last 10–15 years. In Italy, evidence suggests that the prevalence of F. hepatica in ruminants is low in the southern part, but C. daubneyi has been recently reported at high prevalence in the same area. Given the importance of reliable tools for liver and rumen fluke diagnosis in ruminants, this study evaluated the diagnostic performance of the Mini-FLOTAC (MF), Flukefinder® (FF) and sedimentation (SED) techniques to detect and quantify F. hepatica and C. daubneyi eggs using spiked and naturally infected cattle faecal samples.MethodsBriefly, negative bovine faecal samples were artificially spiked with either F. hepatica or C. daubneyi eggs to achieve different egg count levels: 10, 50 and 100 eggs per gram (EPG) of faeces. Moreover, ten naturally infected cattle farms from southern Italy with either F. hepatica and/or C. daubneyi were selected. For each farm, the samples were analysed individually only with MF technique and as pools using MF, FF and SED techniques. Bayesian latent class analysis (LCA) was used to estimate sensitivity and accuracy of the predicted intensity of infection as well as the infection rate in the naturally infected farms.ResultsThe outcome of this study showed that the highest number of eggs (F. hepatica and C. daubneyi) recovered was obtained with MF, followed by FF and SED in spiked infected samples at 50 and 100 EPG, while at lower infection levels of 10 EPG, FF gave the best results. Moreover, the sensitivity for all the techniques included in the study was estimated at > 90% at infection levels > 20 EPG for both F. hepatica and C. daubneyi eggs. However, MF was the most accurate of the three techniques evaluated to estimate fluke infection intensity. Nevertheless, all three techniques can potentially estimate infection rate at farm level accurately.ConclusionsOptimization and standardization of techniques are needed to improve the FEC of fluke eggs.Graphical

Identification of Cotylophoron cotylophorum (Fischoeder, 1901) in cattle on St. Kitts, West Indies and its relationship with African and Asian populations

There is limited information about the species of rumen fluke (Family Paramphistomidae) in the Caribbean. However, knowledge of species distribution is needed to better understand disease risk and epidemiology. Morphological identification is challenging with more recent DNA sequencing enabling a better understanding of rumen fluke distribution. In this study, rumen fluke specimens, collected between 2015 and 2016 from cattle on the island of St. Kitts, West Indies, were analysed. The ribosomal internal transcribed spacer 2 (ITS-2) region of rDNA was amplified using generic trematode primers. Results from Sanger sequencing were compared to reference sequences in GenBank and indicated the species was Cotylophoron cotylophorum with 100% sequence identity and 91% query cover. The ITS2 sequences were then compared to previously published ITS2 sequences for the Cotylophoron genus. When all the St. Kitts C. cotylophorum ITS2 sequences were compared with all other Cotylophoron sequences from India, Kenya, and Zimbabwe, three variable nucleotide sites, resulting in five unique haplotypes, were identified. Nine ITS2 sequences shared haplotype 1, which included all those from St. Kitts and single representatives from India and Kenya, potentially indicating global movement of this species.

Prevalence of flukes (Fasciola hepatica and paramphistomids) in cattle in south-eastern Mexico.

The objective of this study was to determine the risk factors and prevalence of trematodes in south-eastern Mexico. The prevalence of trematodes was determined in 1010 bovines. The study was carried out from October 2018 (n=291) to December 2019 (n=719). Only in 2019 rumen and liver fluke eggs were differentiated. Faecal samples (n=311) were obtained from farms in southeast Mexico located in Tabasco, Chiapas and Campeche. In addition, the presence of flukes in liver and rumen from slaughtered cattle in abattoirs was recorded with a total of 408 samples. A logistic procedure was used to obtain the prevalence and the effect of main risk factors such as land physiography (flooded areas and hills), year, sex, animals' age and type of sample obtained (eggs in faeces and flukes). The general prevalence of flukes in cattle was 32.3 % in 2018 and 41.7 % in 2019. Prevalence of F. hepatica (liver fluke) was 18.6 % (134/719) and that of paramphistomids (rumen fluke) was 33.4 % (240/719). The infected cattle from the slaughterhouse indicated a lower prevalence of F. hepatica (1 %) and rumen fluke (26.7 %) than in farms detected by egg in faeces (41.8 % and 42.1 %, respectively). The physiographic zone was decisive in the presence of F. hepatica and rumen fluke, while sex did not represent a risk factor (P > 0.05). The environmental conditions of the Mexican southeast favour the presence of both liver and rumen fluke.

Identification and prevalence of fluke infection in yak and Tibetan sheep around Qinghai Lake, China.

Liver flukes (Fasciola spp.) and rumen flukes (Paramphistomum spp.) are significant parasites in livestock worldwide, and Fasciola spp. are considered an important zoonotic parasite. To our knowledge, there are no reports on fluke species identification and epidemiological prevalence in yak and Tibetan sheep around Qinghai Lake, China. Therefore, this study aimed to identify the major fluke species and determine the prevalence of fluke infections among yak and Tibetan sheep in this area. A total of 307 fecal samples were collected and fluke eggs identified using morphology and molecular methods. Our study is the first to display that the predominant fluke species were F. hepatica and P. leydeni in yak and Tibetan sheep around Qinghai Lake. The overall prevalence of fluke infections in yak and Tibetan sheep was 57.7% (177/307). Specifically, the prevalences of F. hepatica and P. leydeni were 15.0% (46/307) and 31.6% (97/307), respectively, and the co-infection of both species was 11.1% (34/307). No significant difference existed in the prevalence of overall fluke infection between yak and Tibetan sheep (p < 0.05). However, F. hepatica prevalence was significantly different in yak and Tibetan sheep (p < 0.05) but not P. leydeni. The findings of this study provide useful information about the current status of natural fluke invasion in yak and Tibetan sheep around Qinghai Lake, which could be important for monitoring and controlling these parasites in the region.

Multispecies helminth parasitism of grazing dairy cows in Germany and Austria, examined in the housing period

Helminth composition and burden data for dairy cows have not been reported for >40years for Germany and even less information is available for Austria. In the context of two recent studies, helminth parasitism was studied in 32 cows (23 from six farms in Bavaria and Tyrol; 9 from one farm in Saxony) from pasture-based dairy farms necropsied during the housing period. Helminths were enumerated and identified based on morphological characters (all helminths but rumen flukes) or molecular techniques (rumen flukes). Thirteen species of gastrointestinal nematodes and two species each of liver flukes (Fasciola hepatica, Dicrocoelium dendriticum) and rumen flukes (Calicophoron daubneyi, Paramphistomum leydeni) were recorded; no lungworms were recovered from any cow. Early fourth-stage (inhibited) larval Ostertagia species nematodes (210 to 140,600) were recovered from all cows, 31 each had adult Ostertagia ostertagi/Ostertagia lyrata (40 to 2020) and Trichostrongylus axei (10 to 53,400), 23 Oesophagostomum radiatum (1 to 242) and 20 Cooperia punctata (10 to 3330). Other nematodes present in descending order of prevalence were: Cooperia oncophora/Cooperia surnabada, Ostertagia leptospicularis/Ostertagia kolchida, Oesophagostomum venulosum, Bunostomum phlebotomum, Chabertia ovina, Nematodirus helvetianus, Trichostrongylus longispicularis, Haemonchus contortus and Aonchotheca bilobata. The cows from Bavaria and Tyrol harbored more total gastrointestinal nematodes than that from Saxony (geometric mean adult plus inhibited larval nematodes, 6510 vs. 2051, respectively). However, in both cohorts of cows abomasal nematodes accounted for ∼97% of the total nematode burden with inhibited larval Ostertagia species nematodes contributing over 70% of the total gastrointestinal nematode burden and∼96% of the Ostertagia species burden. Approximately 44%, 37% and 19% of the cows harbored <5000, 5000 to 10,000 or>10,000 total gastrointestinal nematodes, respectively. Fecal nematode egg and coproculture nematode larval counts significantly correlated with the cows' total adult nematode burden (rs=0.354, p<0.05, and rs=0.608, p<0.001, respectively). Although the magnitude of nematode burden to exert production effects on dairy cows is not well defined and may vary relative to several factors including nutritional supplementation, the level of mixed parasitism found in this investigation supports consideration of grazing dairy cows in helminth control measures, especially at the time of housing in autumn.

Multi-detection for paramphistomes using novel manually designed PAR-LAMP primers and its application in a lateral flow dipstick (LAMP-LFD) system

Loop-mediated isothermal amplification (LAMP) has been applied for the detection of various parasites, and its application in lateral flow dipstick (LFD) can improve the convenience of point-of-care diagnosis. A novel PAR-LAMP probe and primers were designed by manual selection from a region of low variation in the ITS-2 DNA sequence. Up to six species of rumen fluke were detected by LAMP and LAMP-LFD in this study. Target specificity and sensitivity were tested, revealing a high target specificity (accuracy) and a low limit of detection (sensitivity). Different target sensitivities of paramphistome were presented, including 5 pg for Gastrothylax crumenifer and Carmyerius sp.; 1 pg for Fischoederius elongatus, Orthocoelium parvipapillatum, and O. dicranocoelium; and 0.1 pg for Paramphistomum epiclitum. LAMP-LFD can detect a paramphistome egg even in contaminated in feces that was spiked with the egg under laboratory conditions. In addition, natural paramphistome infection in cattle from Surat Thani and Khon Kaen provinces, Thailand, was evaluated by detection of egg contamination in fecal specimens using PAR-LAMP primers. The PAR-LAMP detection result was also statistically evaluated by microscopic examination of feces. This study presents the application of novel manually designed primers in a LAMP-LFD system for improving performance in detection and diagnosis assays for paramphistomosis.