An epizootiological survey was conducted to investigate the prevalence of hepatozoonosis in a population of 924 apparently asymptomatic dogs from different regions of Croatia. DNA was isolated from ...canine blood and screening PCR on the 666bp fragment of 18S rRNA revealed that 108 (11.8%) of dogs were infected. Positive samples were confirmed by partial sequencing of the 18S rRNA gene. The consensus sequences, derived from various sequence data sets, were compared with sequences of 18S ssrRNA of Hepatozoon spp. available in GenBank. The alignments revealed 106 Hepatozoon canis and two Hepatozoon sp. sequences. Among H. canis isolates, we found a certain amount of heterogeneity, while both Hepatozoonsp. isolates were identical to the Spanish isolate (Accession No. AY600625) from Clethrionomys glareolus. On the basis of eight commonly mutated nucleotide positions in the partial 18S rRNA gene sequence, we divided the H. canis isolates into five groups. The results obtained indicate a higher prevalence and significance of hepatozoonosis in Croatia than previously believed and demonstrate that the organisms belonging to H. canis that infect European dogs are genetically very heterogeneous.
Babesiosis, the disease caused by tick-borne hematozoan parasites of the genus
Babesia, is particularly common in dogs, and is caused by several “large” species of
Babesia, as well as by an ...increasing number of “small” species of
Babesia, some of which appear to be more closely related to members of the genus
Theileria. In this work, blood samples were collected from 848 randomly selected, asymptomatic dogs and from 81 symptomatic dogs, microscopically positive for
Babesia, and characterised by PCR and sequence analysis of a fragment of the ssrRNA gene. A prevalence of 3.42% (29 of 848) was found in asymptomatic dogs and sequence analysis revealed the presence of
Babesia canis canis in 20 dogs (69%),
Babesia gibsoni in six dogs (21%),
Babesia canis vogeli in two dogs (7%) and
Theileria annae in one dog (3%). In the group of symptomatic dogs, which were all positive by PCR,
B. canis canis was the predominant species (78 dogs, or 96%), followed by single infections with
B. canis vogeli,
Babesia caballi and
Theileria equi. Our study has confirmed that dogs are infected with a wide range of both large and small piroplasm species and subspecies, including
B. caballi and
T. equi, two parasites usually found in horses. The detection of the pathogenic species
B. canis canis and
B. gibsoni in asymptomatic dogs indicates that the relationship between parasite species/subspecies and clinical signs of infection in dogs deserves further investigation. Finally, the identities of the tick vectors transmitting
T. annae and
B. caballi remain to be elucidated.
Trichinella spiralis and Trichinella britovi are species that are frequently found in domestic pigs and various sylvatic animals in Croatia. During routine trichinoscopy, non-encapsulated larvae were ...detected in the muscle tissue of a domestic pig. Artificial digestion revealed a larvae burden of 602 muscle larvae per gram of tissue. Tissue section analysis confirmed the presence of non-encapsulated larvae. Multiplex PCR identified the larvae as T. pseudospiralis. This observation is consistent with the reports of a local veterinary inspector who described the presence of non-encapsulated Trichinella in four individual cases over the last 2 years. This is the first report of T. pseudospiralis in Croatia and one of very few cases of T. pseudospiralis infection described in domestic pigs. The detection of non-encapsulated larvae stresses the need for implementation of artificial digestion instead of trichinoscopy for the detection and identification of Trichinella infections.
Because of its role in human disease, there are increasing global requirements for reliable diagnostic and control methods for Trichinella in food animals to ensure meat safety and to facilitate ...trade. Consequently, there is a need for standardization of methods, programs, and best practices used in the control of Trichinella and trichinellosis. This review article describes the biology and epidemiology of Trichinella, and describes recommended test methods as well as modified and optimized procedures that are used in meat inspection programs. The use of ELISA for monitoring animals for infection in various porcine and equine pre- and post-slaughter programs, including farm or herd certification programs is also discussed. A brief review of the effectiveness of meat processing methods, such as freezing, cooking and preserving is provided. The importance of proper quality assurance and its application in all aspects of a Trichinella diagnostic system is emphasized. It includes the use of international quality standards, test validation and standardization, critical control points, laboratory accreditation, certification of analysts and proficiency testing. Also described, are the roles and locations of international and regional reference laboratories for trichinellosis where expert advice and support on research and diagnostics are available.
Correspondence: 1 Corresponding Author: Caroline Frey, Vetsuisse Faculty, University of Bern, Institute of Parasitology, PO Box 8466, 3001 Bern, Switzerland. caroline.frey{at}ipa.unibe.ch
...Trichinellosis is a zoonotic disease that is caused by the nematode Trichinella spp. Both European Union regulations and guidelines from the World Organization for Animal Health foresee the possibility of conducting serological surveillance for Trichinella spp. A newly developed commercial enzyme-linked immunosorbent assay (ELISA) was evaluated against 2 existing diagnostic techniques: an in-house ELISA and an in-house Western blot. A total of 875 Trichinella larva–negative samples of pigs and 93 Trichinella larva–positive samples of both naturally and experimentally infected pigs were included in the study. Bayesian modeling techniques were used to correct for the absence of a perfect reference test. The sensitivity and specificity of the commercial ELISA was 97.1–97.8% and 99.5–99.8%, respectively. Sensitivity analysis demonstrated high stability in the models. In a serological surveillance system, ELISA-positive samples should be tested by a confirmatory test. The Western blot is a suitable test for this purpose. With the use of the results of the models, the sensitivity and specificity of a test protocol in both ELISA and Western blot were 95.9% and 99.9%, respectively. The high sensitivity and specificity were achieved with a lower limit of detection than that of the routine artificial digestion test, suggesting that serological surveillance is a valuable alternative in surveillance for Trichinella spp. in pig production.
Key Words: Bayesian techniques enzyme-linked immunosorbent assay evaluation pigs Trichinella
Milk contains about 70 indogene enzymes, while only twenty of them were investigated. One of the less explored is the milk enzyme cathepsin D, proteolytic enzyme located in the lysosomes, which are ...an integral part of the somatic cells whose number varies depending on the animal’s health. Unlike cow’s milk, in ewe’s milk the presence of cathepsin D was not determined, which could affect the production of dairy products, especially cheese, which are traditionally produced in Croatia and Mediterranean. This paper shows the presence of cathepsin D and its forms in ewe’s milk by modified Western Blotting method. The analysis confirmed the presence of procathepsin D, mature and heavy chain of cathepsin D. Pseudocathepsin D and light chain of cathepsin D were not detected.
Faecal and vomit samples were collected from 12 griffon vultures (
Gyps fulvus
) in May 2002 and examined for bacteria, fungi and parasites. Bacteria present included
Escherichia coli
,
Aeromonas ...salmonicidia
,
Proteus
sp.,
Escherichia fergusonii
,
Providencia alcalifaciens
,
Micrococcus
sp. and
Streptococcus
sp. Mycological examination revealed the presence of
Candida
sp., yeasts and in one case,
Rhodotorula rubra
. Strongyle eggs were found in several samples. In most faecal samples, ascarid eggs of
Toxocara
type were found. Tapeworm eggs of
Moniezia
type were present in one faecal sample. Both microbiologic and parasitic findings are most likely the result of the feeding habits of griffon vultures.
Complete sequences of ribosomal and mitochondrial genes of the giant liver fluke Fascioloides magna are presented. In particular, small subunit (18S) and internal transcribed spacers (ITS1 and ITS2) ...of the ribosomal gene (rDNA), as well as cytochrome c oxidase subunit I (cox1) and nicotinamide dehydrogenase subunit I (nad1) of the mitochondrial DNA (mtDNA), were analyzed. The 18S and ITS sequences were compared with previously published sequences of the liver fluke Fasciola hepatica. Fixed interspecific genetic differences were determined that allow molecular differentiation of F. magna and F. hepatica using either the PCR-RFLP method or PCR amplification of species-specific DNA regions. Additionally, intraspecific sequence polymorphism of the complete cox1 and nad1 mitochondrial genes in geographically distinct F. magna populations was determined. Based on the sequence divergences, short (<500 bp) variable regions suitable for broader biogeographical studies of giant liver fluke were designed.
Deer fascioloidosis is a serious and potentially fatal disease caused by the non-native trematode Fascioloides magna. Infections of red deer with F. magna in Croatia have been reported for the first ...time in 2000 in the Baranja region. Subsequently, the disease spread throughout the Eastern parts of the country, involving all 3 deer species (red, roe, and fallow) and mouflons. Within the disease control programme (DCP), livers from all shot deer were thoroughly analysed and all detected trematodes and gross lesions were counted and categorized. Prevalence of positive animals, in this study for Spačva region, in the period ranging from 2007 to 2012 was 36.42% (46.39% when fawns are not considered). Epidemiological analysis was applied to evaluate risk factors and disease patterns at the population level with the aim to understand factors with negative influence on therapeutic effect. Each demographic variable was tested at the seasonal, individual and location level. Model for pathological lesions suggested that the likelihood of lesions was dependent on age (p = 0.003). We did not find any locality or sex related significant differences. Finally, environmental characteristics and migratory patterns were analysed using Geographic Information System (GIS) and showed that Spačva region represents an epidemiological unit for red deer fascioloidosis.
Gastrointestinal nematodes (GIN) are a serious health problem and represent the most significant constraint in sheep grazing operations. Problems tend to be worse in organic sheep farming systems, as ...a consequence of a less restricted access of animals to outdoor environment with a higher exposure to infective larvae. In domestic animals, GIN are effectively controlled by an aggressive prophylactic administration of commercially available anthelmintics. As a consequence to a common overdose and misuse of readily available antiparasitic treatments, there is an inevitable development of populations of GIN resistant to all major classes of anthelmintics. Also, the control of GIN that is based entirely on the anthelmintic use, threatens sustainability of the sheep farming worldwide. The combination of the optimized use of anthelmintic drugs and alternative approaches seem to be a reasonable choice in sustainable parasitic control programs that offer a substantial reduction of anthelmintic treatments and conservation of anthelmintic efficacy. In that aspect, a "targeted selective treatment (TST)" directed towards animals clinically diagnosed with GIN, seems to be an effective approach to leave some parasite populations unexposed to anthelmintics (refugia) and to reduce development of anthelmintic resistance. Also, many current research efforts aim to find and validate sustainable non-chemotherapeutic approaches to GIN control, including changes in grazing management, optimized nutrition, dietary supplementation, consumption of plants with anthelmintic properties, biological control by nematophagous fungi, copper oxide wire particles (COWP), and homeopathic treatments. This manuscript outlines (outlines) and discusses relevant alternative approaches for GIN control in modern sheep farming systems.
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