Els Goossens

Occurrence and Molecular Characterization of Cryptosporidium and Giardia in Captive Wild Ruminants in Belgium

Abstract Both Cryptosporidium and Giardia are frequently found in the stool of domestic ruminants, especially young animals. Wild ruminants are also host to these protozoa, but the prevalence of these parasites in both free-ranging and captive nondomesticated ruminants needs to be further investigated. Moreover, the role of wild ruminants serving as reservoirs for these zoonotic parasites remains unclear. Therefore, a cross-sectional survey was conducted to estimate the occurrence of Cryptosporidium and Giardia in captive wild ruminants younger than 6 mo and to determine the potential of these animals to serve as reservoirs for these zoonotic parasites. A total of 67 captive wild ruminants belonging to 21 different animal species at the Antwerp Zoo (Belgium), along with 82 American bison (Bison bison) on a commercial breeding farm, were sampled for the detection of Cryptosporidium and Giardia, using a commercial immunofluoresence assay (Merifluor Cryptosporidium/Giardia IFA). The Cryptosporidium prevalence was 7.5% in the Antwerp Zoo animals and 3.7% in the bison from the breeding farm. All but two of the Cryptosporidium-positive animals were younger than 1 mo of age. Molecular characterization by amplification of the 70-kDa heat-shock protein and the 18S ribosomal DNA gene identified Cryptosporidium parvum in four animals of the Antwerp Zoo. The prevalence of Giardia was 8.9% in the Antwerp Zoo animals and 23.2% in the bison calves. Most Giardia-positive animals were older than 1 mo of age. Molecular characterization on the β-giardin gene and the triose phosphate isomerase gene identified Giardia duodenalis assemblage A in the Antwerp Zoo and both G. duodenalis assemblage A and assemblage E in the bison calves. These findings indicate that both protozoan parasites are prevalent in captive wild ruminants and that these animals can serve as a potential reservoir for zoonotic transmission.

The influence of peptide impurity profiles on functional tissue-organ bath response: the 11-mer peptide INSL6[151-161] case.

Bioactive peptides have great pharmaceutical potential as nutraceuticals, diagnostics, and therapeutic drugs in several clinical areas. Thus, the search for novel lead peptides with a biological function has attracted renewed interest. Crude peptide material (i.e., ~70% purity) of INSL6[151-161] (NH₂-FRSLFWGNHSQ-COOH) was found to trigger a contractile response in guinea pig ileum longitudinal smooth muscle preparations using tissue-organ baths. However, the purified peptide (i.e., ≥ 95% purity) had no effect on this model. Further investigation with crude materials from other suppliers, with purities ranging between 50% and 80%, indicated that the crude products gave a false-positive functional tissue-organ bath conclusion. These observations question the functionality conclusions when using crude-purity peptide materials; during the initial research or discovery phase, peptide quality is generally neglected, possibly leading to misinterpretation of biological results due to by-products from peptide synthesis and, thus, wrong fail/pass decisions. Therefore, we strongly recommend appropriate quality control testing before using any peptides for initial biomedical research or discovery purposes.

Antiserum against the conserved nine amino acid N-terminal peptide of influenza A virus matrix protein 2 is not immunoprotective

The recent emergence and rapid spread of the pandemic H1N1 swine influenza virus reminded us once again of the need for a universal influenza vaccine that can elicit heterosubtypic protection. Here, we show the superior immunogenicity and immunoprotective capacity of the full-length matrix protein 2 ectodomain (M2e) peptide coupled to keyhole limpet haemocyanin (KLH) compared with the N-terminal 9 aa residues of M2e (SP1). Immunization with M2e-KLH protected mice against a lethal challenge with influenza A virus and significantly reduced weight loss and lung virus titres. In addition, passive transfer of serum raised in rabbits against M2e-KLH protected mice against a lethal influenza virus challenge, whereas serum from rabbits immunized with SP1-KLH did not. Nevertheless, immunofluorescence staining revealed that rabbit serum raised against SP1-KLH bound specifically to infected Madin-Darby canine kidney cells. We conclude that the peptide SP1 contains an immunogenic epitope that is not sufficient for immunoprotection.

Serologic screening for 13 infectious agents in roe deer ( Capreolus capreolus ) in Flanders

Introduction In order to investigate the role of roe deer in the maintenance and transmission of infectious animal and human diseases in Flanders, we conducted a serologic screening in 12 hunting areas. Materials and methods Roe deer sera collected between 2008 and 2013 (n=190) were examined for antibodies against 13 infectious agents, using indirect enzyme-linked immunosorbent assay, virus neutralisation, immunofluorescence, or microagglutination test, depending on the agent. Results and discussion High numbers of seropositives were found for Anaplasma phagocytophilum (45.8%), Toxoplasma gondii (43.2%) and Schmallenberg virus (27.9%), the latter with a distinct temporal distribution pattern following the outbreak in domestic ruminants. Lower antibody prevalence was found for Chlamydia abortus (6.7%), tick-borne encephalitis virus (5.1%), Neospora caninum (4.8%), and Mycobacterium avium subsp paratuberculosis (4.1%). The lowest prevalences were found for Leptospira (1.7%), bovine viral diarrhoea virus 1 (1.3%), and Coxiella burnetii (1.2%). No antibodies were found against Brucella sp., bovine herpesvirus 1, and bluetongue virus. A significant difference in seroprevalence between ages (higher in adults >1 year) was found for N. caninum. Four doubtful reacting sera accounted for a significant difference in seroprevalence between sexes for C. abortus (higher in females). Conclusions Despite the more intensive landscape use in Flanders, the results are consistent with other European studies. Apart from maintaining C. abortus and MAP, roe deer do not seem to play an important role in the epidemiology of the examined zoonotic and domestic animal pathogens. Nevertheless, their meaning as sentinels should not be neglected in the absence of other wild cervid species.

EVALUATION OF THREE STRATEGIC PARASITE CONTROL PROGRAMS IN CAPTIVE WILD RUMINANTS

Abstract Parasite control in wild ruminants is based mainly on anthelmintics. The objective of the present study was to evaluate three parasite control programs in seven herds of captive wild ruminants in 3 consecutive years. In the first year, a biannual spring–summer treatment regime with fenbendazole at 7.5 mg/kg body weight p.o. for 3 days was applied. The next year, an early-season treatment program with three administrations of fenbendazole at the same dosage at 3-wk intervals was used. In the third year, an early-season treatment program with ivermectin (0.2 mg/kg p.o. for 3 days), applied three times at 5-wk intervals, was evaluated. Effectiveness of these control programs was assessed by fecal egg counts and by scores of body condition and fecal consistency at weekly intervals. Involved animal species were Arabian oryx (Oryx leucoryx), scimitar-horned oryx (Oryx dammah), slender-horned gazelle (Gazella leptoceros), Soay sheep (Ovis aries soay), ibex (Capra ibex), red deer (Cervus elaphus hippelaphus), and Nelsons elk (Cervus elaphus nelsoni). With the spring–summer regime, fecal egg counts remained low during the first 5 mo, but from September onward, they slowly increased to significant levels in all seven herds. The early-season program with fenbendazole resulted in fecal egg counts near zero during the entire year in four herds but a significant egg shedding from August onward was noted in Arabian oryx, scimitar-horned oryx, and Soay sheep. The early-season program with ivermectin resulted in very low to zero egg shedding in gazelle, adult Soay sheep, ibex, red deer, and Nelsons elk during the entire grazing season, but failed to prevent high shedding in October in Arabian oryx and scimitar-horned oryx. High contamination of grassy pastures with infective larvae/eggs in the previous year or before the first treatment may explain failure of the treatment regimes in some herds.

A 12-MONTH SURVEY OF GASTROINTESTINAL HELMINTH INFECTIONS OF CERVIDS KEPT IN TWO ZOOS IN BELGIUM

Abstract Infections with helminths are a major health issue in captive and wild deer. In this study, fecal egg count patterns and clinical signs associated with gastrointestinal nematodes were assessed for 12 mo in nine cervid herds kept under different husbandry conditions at two sites. At site 1, an urban zoo, fecal egg counts remained low and no clinical signs of parasitic gastroenteritis were seen in the herds of fallow deer (Dama dama), Dybowskis deer (Cervus nippon dybowski), pudu (Pudu pudu), and reindeer (Rangifer tarandus tarandus). Helminth infection at this site may have been successfully prevented by daily dung removal of the small sandy-soil enclosures, and applying routine anthelmintic treatment was not justified. At site 2, a wild animal park, involved species were red deer (Cervus elaphus hippelaphus), Nelsons elk (Cervus elaphus nelsoni), Père Davids deer (Elaphurus davidianus), European elk (Alces alces alces), and reindeer (Rangifer tarandus tarandus). Nematode eggs were frequently encountered in herds of red deer, Nelsons elk, and European elk, which were kept on larger, grassy enclosures that were irregularly cleaned. The trimodal pattern of fecal egg counts in herds from the wild animal park, consisting of a small spring rise in June, a peak in October, and a small rise in February, indicates that infective larvae on pastures are the main source of infection. In addition, routine anthelmintic treatment with fenbendazole in April and July limited egg shedding, but reinfection rapidly occurred. In two European elk and one reindeer, increasing fecal egg counts were associated with loss of fecal consistency and reduced appetite. Three genera and three species of nematodes were recovered at necropsy of one red deer and three Nelsons elk: Spiculopteragia spiculoptera, Trichostrongylus spp., Nematodirus filicollis, Capillaria spp., Oesophagostomum radiatum, and Trichuris spp., with total worm counts between 950 and 8,700.

Field evaluation of the efficacy of fenbendazole in captive wild ruminants

The efficacy of in-feed fenbendazole at a dose rate of 7·5 mg/kg bodyweight for three consecutive days was assessed in five Arabian oryx (Oryx leucoryx), six scimitar-horned oryx (Oryx dammah), 14 slender-horned gazelles (Gazella leptoceros), eight Soay sheep (Ovis aries aries soay), 13 alpine ibex (Capra ibex ibex), six red deer (Cervus elaphus hippelaphus) and 11 Nelson’s elk (Cervus elaphus nelsoni) kept in five herds in a zoo. The efficacy was assessed by means of repeated faecal egg count reduction (FECR) tests and in vitro egg hatch assays. Fenbendazole was highly effective against nematodes in five of the seven species, consistently reducing egg shedding by more than 90 per cent. In the egg hatch assays of the five herds, 50 per cent inhibition of hatching (LD50) was observed at a concentration of thiabendazole below 0·1 μg/ml. In the Arabian oryx and alpine ibex the efficacy of fenbendazole was less than 90 per cent, and the LD50 in the egg hatch assays was between 0·1 and 0·2 μg/ml thiabendazole.