Chromosome set and rDNA sequences of the larval digenean Cercariaeum crassum were analysed in order to clarify its systematic position and possible adult form. Parasites were obtained from the sphaeriid bivalve Pisidium amnicum, collected in Lithuanian and Finnish rivers. The karyotype is shown to consist of five pairs (2n = 10) of large, up to 14 μm, chromosomes. Complement, composed of a low diploid number of exclusively bi-armed elements, presumably arose through Robertsonian fusions of acrocentric chromosomes. Consistent with a Robertsonian-derived karyotype, one or two small, metacentric, mitotically stable B chromosomes were detected in the cells of parthenitae isolated from some host individuals. A phylogenetic analysis using rDNA internal transcribed spacer 2 (ITS2) and 28S sequences corroborates the allocation of C. crassum to the family Allocreadiidae. In neighbour-joining and maximum parsimony phylogenetic trees C. crassum clusters into one clade with Allocreadium spp., and is the closest sister group in relation to A. isoporum; the level of rDNA sequence divergence between them (2.67% for ITS2 and 1.16% for 28S) is consistent with the level expected for intrageneric variation. The present study adds significant information to a database for establishing species-specific characters for confident characterization of different developmental stages of allocreadiid species, clarification of their life cycles and evaluation of intra- and interspecific variability.

Very little is known about parasitic diseases of eastern box turtles (Terrapene carolina carolina). The objective of this study was to examine the parasitic fauna of eastern box turtles collected from southern Indiana, USA. Turtles (n = 40) were salvaged mostly as road kills from southern Indiana between May and October 2009. Seven species of helminths in total were found parasitizing the gastrointestinal tract, including two digenean trematodes (Brachycoelium salamandrae and Telorchis robustus) and five nematodes (Oswaldocruzia pipiens, Cosmocercoides dukae, Falcaustra affinis, F. chelydrae and Serpinema trispinosus). We report prevalence, abundance and mean intensity of infection for all helminths. Helminths were not found in any other organs examined (heart, gonads, liver, heart, kidney and urinary bladder) and no ectoparasites were found. Overall, mean intensity of infections was low (1–14 parasites/host), suggesting that these parasites are unlikely to be associated with negative health impacts. This constitutes the first study of this kind for Indiana.

Sporocysts of Prosorhynchoides borealis were obtained from the marine bivalves Abra prismatica and studied using transmission electron microscopy. The sporocyst body consists of a mass of branching and intertwining hollow tubules that ramify through the host's digestive gland and gonads. This study investigated the ultrastructure of the sporocyst branches which comprise alternate distended areas (brood chambers) with a relatively thin body wall, narrower portions with a thicker body wall (constricted areas) and terminal regions. Pronounced differences between these areas were revealed in the structure of their tegument and body cavity lining, as well as in the cellular composition of the subtegumental layers. Body wall composition in distended areas was consistent with the specialization for cercarial nurture in the brood chambers. The structure of the constrictions suggested a dual role of nutrient absorption and physical separation of adjacent brood chambers. Two types of terminal region were identified, one specialized for the investigation and penetration of host tissues and the other, in which the germinal cells are formed, for cercarial production. The overall structure of the sporocyst branches helps explain why this linear modular system, i.e. brood chambers and constrictions continuously growing into the host tissue, enables the sporocyst's long-term existence and can continuously produce cercariae in numbers comparable with those produced by rediae and/or daughter sporocyst infrapopulations in other digeneans. The origin of the nuclei in the outer tegumental layer of some branching bucephalid sporocysts is also discussed.

The pathogenesis of angiostrongyliasis, resulting from Angiostrongylus cantonensis invasion of the human central nervous system, remains elusive. Anthelmintics are usually used to kill worms, although dead worms in the brain may cause severe inflammation which will lead to central nervous system damage. Therefore, combination therapy with anthelmintics and anti-inflammatory drugs in the treatment of human angiostrongyliasis needs further study. To evaluate the efficacy of albendazole combined with a marine fungal extract (m2-9) in A. cantonensis infection, BALB/c mice infected by the third-stage larvae of A. cantonensis were divided into three groups: mice treated with albendazole or m2-9 alone or in combination from day 5 post-inoculation (PI). Several efficacy parameters were recorded, including weight change, worm recovery, neurological function, behavioural analysis, eosinophil and leucocyte counts. The results showed that combination therapy increased body weight, reduced worm burden, improved learning ability, memory and action, decreased neurological dysfunction and leucocyte response in these mice. The combination of albendazole and m2-9 treatment significantly decreased leucocyte response and increased the frequency of rearing, compared to infected mice treated with either drug alone. Therefore, m2-9 is a natural product with potentially significant therapeutic value for angiostrongyliasis and is worthy of further study.

Faecal samples (n = 1947) from captive wild impala (Aepyceros melampus melampus) were examined over a period of 14 months to determine quantitative seasonal helminth egg excretion patterns and qualitative protozoan oocyst excretion patterns. Geometric mean monthly faecal egg counts (FECs) ranged from 20 to 575 and coprocultures revealed three parasite genera, namely Trichostrongylus, Haemonchus and Strongyloides. Larvae of the Trichostrongylus spp. were most predominant from faecal cultures. No trematode eggs or lungworms were detected and eggs of the cestode Monezia were only seen in two samples during the entire study period. The nematode FECs showed a marked seasonal variation, being higher during the rainy season, moderate during the cool dry season and low during the hot dry season. The rainy season had significantly higher FECs than the dry season (P < 0.01). The percentage of helminth-egg positive faecal samples ranged from 90.6 to 100% in the rainy season and 72.4 to 85.6% in the dry season. Overall mean FECs in unpelleted faeces were significantly higher than in pelleted faeces (P < 0.01). However, the FECs were not significantly different among seasons in unpelleted faeces (P>0.05), but were significantly higher in pelleted faeces in the rainy season than the dry season (P < 0.05). Pellet size had a significant effect on FEC, with smaller pellets having higher FEC (P < 0.05). Strongyloides eggs and coccidia oocysts were only seen during the rainy season. This represents the first documentation of seasonal parasitic infestation in captive wild antelopes in Zambia. Treatment and control strategies for helminths in these captive wild impala are also suggested based on the findings from this study.

In the present study, 81 specimens of Capoeta capoeta gracilis (Keyserling, 1861) were collected from the Shiroud River, the Caspian drainage basin, Iran in June and October 2007. The fish were examined for infection with the nematode Rhabdochona fortunatowi (Dinnik, 1933). Nematodes were counted and sexed, then the sex ratio and intestinal site preference of the R. fortunatowi individuals and the intersexual interactions of the parasites were investigated. Significant differences were observed in the prevalence and intensity of infection in relation to host size and sampling season, but not host sex. Parasite burden (mean intensity) was higher in October than in June and the results revealed a decrease in female-to-male sex ratio (FMR) in the fish in October compared to the ones in June. In other words, as the mean intensity of infection of R. fortunatowi increases, the proportion of male worms increases, too. The female-biased sex ratio in June was replaced by a male-biased one in October. Nematode distribution rose to a significant peak in the mid-region of the fish intestine relative to the anterior and posterior parts. Some changes in the distribution of male and female worms in different parts of the host intestine were also observed during June and October. Density-dependent selection and intersexual competition seem to be the main factors driving such a shift in the sex ratio and its variation in different parts of the host intestine.

A survey of the occurrence of Dirofilaria immitis, Borrelia burgdorferi, Ehrlichia canis and Anaplasma phagocytophium in dogs was undertaken in the People's Republic of China between October 2008 and October 2009. A total of 600 blood samples were taken from dogs in four cities in China: 300 in Beijing, 150 in Shenzhen, 30 in Shanghai and 120 in Zhengzhou. All samples were tested for the heartworm antigen and antibodies of canine B. burgdorferi, E. canis and A. phagocytophium by using the canine SNAP® 4Dx® test kit. The occurrence of D. immitis, B. burgdorferi, E. canis and A. phagocytophium was 1.17% (7/600), 0.17% (1/600), 2.17% (13/600) and 0.5% (3/600), respectively. In Shenzhen city 2% (3/150), 8.67% (13/150) and 2% (3/150) of samples were positive for D. immitis, E. canis and A. phagocytophium, respectively. The occurrence of heartworm antigen was 0.33% (1/300) in Beijing, 2.00% (3/150) in Shenzhen, 3.33% (1/30) in Shanghai and 1.67% (2/120) in Zhengzhou. We found E. canis and A. phagocytophium only at one site, Shenzhen, while the only occurrence of B. burgdorferi was at Beijing. In conclusion, the dog population in China is at potential risk for D. immitis, B. burgdorferi, E. canis and A. phagocytophium infection, the risk being especially high in southern China.

The infection dynamics of the gill monogeneans Cichlidogyrus tilapiae and C. sclerosus on Oreochromis niloticus with respect to habitat type (reservoir, stream, ponds and cages), host sex, size and seasons was determined between January and November 2008. During the study period, 45.2% of the 650 fish examined were infected with Cichlidogyrus spp. The infected hosts harboured an average of 8.6 ± 3.4 parasites/fish. Across habitat types, the proportion of infected fish was not statistically different. In contrast, the number of parasites recorded on infected fish from different habitat types differed significantly. The highest parasite number was recorded in reservoir-dwelling fish and lowest in stream-dwelling hosts. Concerning sex, more female O. niloticus were infected and harboured a high number of parasites than male and sexually undifferentiated fish. A weak negative relationship was found between rainfall and monthly parasite infections. However, a higher number of parasites and proportion of infected hosts were found during dry than in wet seasons, except in ponds. Results of this study show that differential exposure due to changes in fish behaviour associated with habitat modification and sex may account for the infection difference across the sampled sites. Meanwhile, rainfall and the associated hydrological events are important factors regulating monogenean infections in tropical aquatic environments. The continuous presence of Cichlidogyrus spp. in fish provides evidence of possible parasite outbreaks, indicating the application of biosecurity measures as crucial for the success of intensive fish farming.

This paper describes the helminth community of the Pampean grassland mouse (Akodon azarae) inhabiting poultry farms in central Argentina. Winter diversity (season of high rodent abundance) has been compared to spring diversity (season of low rodent abundance). Species richness was seven in both seasons: five nematodes (Syphacia carlitosi, Stilestrongylus spp., Trichuris laevitestis, Pterygodermatites (Paucipectines) azarai and Protospirura numidica criceticola) and two cestodes (adults of Cyclophyllidea and Taenia taeniaeformis hepatic cysts). No difference in richness was detected between host sexes in each season or among host age classes. However, the helminth community showed 67% similarity between winter and spring, with diversity being significantly higher in spring (H = 0.873) than in winter (H = 0.546; P < 0.0005). This could be attributed to different factors, such as host abundance, host diet or environmental factors, that affect the transmission of each species differently. On the other hand, Stilestrongylus spp. and S. carlitosi showed higher dominance and intensity in both seasons compared to their cohabiting species, P. (P.) azarai and T. laevitestis, respectively. The lower values of the latter two species may be related to a crowding effect due to their large body sizes. This is the first report of cestodes in A. azarae. The finding of T. taeniaeformis strobilocerci could be important in the epidemiology of parasitosis in domestic animals of the farms.

The aims of this study were to investigate the prevalence of Moniezia spp. in domestic ruminants in central Vietnam and to develop a polymerase chain reaction (PCR) technique to distinguish M. expansa from M. benedeni. Among 2040 examined domestic animals (540 cattle, 800 goats, 700 sheep) Moniezia was recovered from 5.4% of cattle, 16.4% of sheep and 20.6% of goats. A set of primers for PCR was designed to classify M. expansa and M. benedeni based on the amplification of DNA corresponding to the internal transcribed spacer of 5.8S rRNA. The 457 specimens (75 from cattle, 162 from goats, 150 from sheep, 30 from horses, 30 from chickens and 10 from dogs) were subjected to PCR for classification of Moniezia spp. PCR products with the expected sizes were amplified from bovine, ovine and caprine specimens. No specific PCR products were found for specimens from horses, chickens and dogs. Of the 75 specimens from cattle, nine were classified as M. expansa and 66 were M. benedeni. Among 162 caprine specimens, 138 were M. expansa and 24 were M. benedeni. The distribution of M. expansa and M. benedeni in 150 ovine specimens was 132 and 18, respectively. These results show that M. expansa is dominant in goats and sheep, whereas M. benedeni is more common in cattle; PCR can be used for classification of these two species.

We investigated the effects of Fascioloides magna infection on the serum biochemistry values of the naturally infected red deer population in eastern Croatia. The investigation was performed on 47 red deer with F. magna infection confirmed patho-anatomically in 27 animals (57.4%). Fibrous capsules and migratory lesions were found in 14 deer while only fibrous capsules without migratory lesions were found in 13 deer. In 13 deer both immature and mature flukes were found, in 5 deer only immature flukes were found and in 9 deer only mature flukes were found. Fascioloides magna infected deer with fibrous capsules and migratory lesions had significantly higher values for lactate dehydrogenase (LDH), glutamate dehydrogenase (GLDH) and globulin, and lower values for albumin/globulin ratio and glucose compared to uninfected deer. Fascioloides magna infected deer with fibrous capsules without the presence of migratory lesions had higher values for alanine aminotransferase (ALT) and globulin, and lower values for albumin/globulin ratio and glucose, than the uninfected deer. The number of immature flukes was positively correlated with values of aspartate aminotransferase (AST), γ-glutamyltransferase (GGT), LDH, GLDH, urea and triglycerides. The number of migratory lesions was positively correlated with GGT, GLDH, globulin and urea values. The creatinine value was positively correlated with the number of mature flukes. The trial showed that F. magna infection causes significant changes in serum biochemistry. Moreover, these changes do not completely resemble changes following F. hepatica infection. Further investigation of changes in liver enzymes and other serum metabolites in controlled, experimentally induced fascioloidosis in red deer is needed to better understand the pathogenesis of F. magna.

In earlier studies of the anthelmintic activity of plant cysteine proteinases (CPs), a period of food deprivation was routinely employed before administration of CPs, but there has been no systematic evaluation as to whether this does actually benefit the anthelmintic efficacy. Therefore, we assessed the effect of fasting on the efficacy of CPs from papaya latex (PL) against Heligmosomoides bakeri in C3H mice. We used a refined, supernatant extract of papaya latex (PLS) with known active enzyme content. The animals were divided into three groups (fasted prior to treatment with PLS, not fasted but treated with PLS and fasted but given only water). The study demonstrated clearly that although food deprivation had been routinely employed in much of the earlier work on CPs in mice infected with nematodes, fasting has no beneficial effect on the efficacy of PLS against H. bakeri infections. Administration of CPs to fed animals will also reduce the stress associated with fasting.

Male and female Schistosoma japonicum worms have dissimilar appearances in their final host. In this study, a morphometric and morphological assessment of whole worms derived from unisexual and mixed infections in mice was conducted using confocal laser scanning microscopy. Worms from mixed infections showed significant morphological changes between 15 and 25 days post-infection (PI). On the fifteenth day PI, 33% of males had formed the conspicuous gynecophoric canal, but only 8% of them had testicular lobes containing a few germinative cells; 13% of females had incipient ovaries with a few immature ovarian cells inside. On the twentieth day PI, the testicular lobes contained more germinative cells in all male worms, while female worms presented vitelline glands. On the twenty-fifth day PI, more germinative cells were observed in the male testicular lobes, and differentiated cells were present in the female ovaries. All worms had fully developed reproductive organs from 30 days PI onwards. Morphometric analysis showed significant differences between mixed and unisexual infections at 35 days PI. Ovaries of worms from unisexual infections contained cells in one stage of maturation and vitelline glands had undifferentiated cells. Our study of S. japonicum provides a detailed comparison of different morphological traits from worms of mixed and unisexual infections throughout development.

A high prevalence (86.7%) of various species of nematodes was observed in the stomach of great cormorants living in Lake Biwa, Japan. There were varying numbers of adults belonging to two common genera, Eustrongylides Jagerskiold 1909 (Nematoda: Dioctophymatidae) and Contracaecum Railliet & Henry 1912 (Nematoda: Anisakidae). The first included common adenophorean nematodes comprising a single species, Eustrongylides tubifex and the second comprised ascaroid nematodes that contained four named species: Contracaecum rudolphii Hartwich, 1964, Contracaecum microcephalum Yamaguti, 1961, Contracaecum multipapillatum Drasche, 1882 and Contracaecum chubutensis Garbin, 2008. After the prevalence and intensity of the infection had been noted, both types of nematodes were frequently observed to penetrate the mucosa and intrude into the wall of the glandular stomach, where they caused gross haemorrhage and ulceration. The Eustrongylides sp. was predominantly found in a nodular lesion of the proventricular wall, while Contracaecum spp. were observed either free in the lumen of the proventriculus or, on occasion, deeply penetrating its wall. Of the Contracaecum spp., C. rudolphii was the most prevalent. Grossly, large numbers of nematodes were present in infected stomachs (for C. rudolphii intensity was 1–34 and 3–57 nematodes in male birds and 1–21 and 1–32 in females; for C. microcephalum 1–2 and 1 in male birds and 1–2 in females; for C. multipapillatum 2 in male cormorants and no infection in females; for C. chubutensis 1–2 and 1 in male birds and 1–5 and 1 in females and for E. tubifex 1–5 nematodes in male birds and 2–8 in females). Ulcerative inflammation and hyperaemia were the most common pathological presentations, especially in areas that had been invaded by parasites. Microscopically, varying degrees of granulomatous inflammatory reactions were seen, in addition to degenerated nematodes which appeared to have deeply penetrated mucosal surfaces and were surrounded by fibrous connective tissues.

Plasmids expressing macrophage migration inhibitory factor (MIF) of Trichinella spiralis (TsMIF), multi-cystatin-like domain protein (MCD-1) of T. spiralis (TsMCD-1), or co-expressing TsMIF and TsMCD-1 were constructed with a pVAX1 vector. Their ability to generate a protective immune response against T. spiralis infection was evaluated in BALB/c mice. Groups of mice were immunized twice at 2-week intervals with 100 μg of recombinant plasmids pVAX1-Tsmif, pVAX1-Tsmcd-1 or pVAX1-Tsmif-Tsmcd-1. Control animals were immunized with phosphate-buffered saline (PBS) or blank vector plasmid. Specific antibody levels (IgG, IgG1, IgG2a, IgG2b, IgM, IgA, IgE) against the recombinant protein TsMIF-TsMCD-1, serum cytokines (interferon (IFN)-γ, interleukin (IL)-4, IL-5, transforming growth factor (TGF)-β1 and IL-17) and CD4+/CD8+ T cells were monitored. Challenge infection was performed 2 weeks following the second immunization and worm burden was assayed at 35 days post-challenge. Vaccination with pVAX1-Tsmif induced moderate serum IFN-γ and increases of CD4+ and CD8+ T cells, but no specific immunoglobulin antibody response. Vaccination with pVAX1-Tsmcd-1 induced a predominant Th1 antibody (IgG2a and IgG2b) response and strong levels of serum IFN-γ, and increases of CD4+ T cells. Importantly, co-expression of TsMIF and TsMCD-1 in DNA immunization produced more serum IFN-γ and markedly enhanced CD4+ and CD8+ T cells than the single DNA vaccine of the two genes. Challenge infection demonstrated that immunization with pVAX1-Tsmif-Tsmcd-1 reduced worm burdens (by 23.17%; P < 0.05).

Previously, an in vitro effect was observed on the complement system not only of the excretory-secretory products but also of somatic antigens from L3 Anisakis simplex larvae. In the present work the effect of anti-A. simplex specific antibodies on C3 and C4 levels in human sera was investigated. Up to 309 samples of sera were tested to determine levels of C3 and C4 and anti-A. simplex antibodies, including immunoglobulins IgG, IgM, IgA and IgE. Significant differences were observed between levels of C3 and C4 and all immunoglobulins except for IgE. In the case of immunoglobulins, the probability that an anti-A. simplex positive subject has a C3 deficiency was 3.8 times higher than a subject without specific antibodies. In conclusion, an association between elevated levels of anti-A. simplex antibodies and C3 and C4 deficiency was demonstrated.

P-glycoproteins (P-gps) are proteins that function as efflux pumps, removing lipophilic xenobiotic compounds from cells. There is evidence that P-gps play a role in the resistance of parasitic nematodes to anthelmintic drugs such as benzimidazoles and macrocyclic lactones. As anthelmintic resistance becomes more common, it is important to identify candidate resistance genes with the aim of understanding the molecular basis of resistance, and of developing assays to detect these resistance-associated changes. We identified several sequences from the genome of the parasite Haemonchus contortus with convincing homology to the known P-gp coding genes of the model nematode Caenorhabditis elegans. Nine of these sequences were successfully amplified by polymerase chain reaction (PCR) and shown to be most similar to the C. elegans sequences for pgp-1, pgp-2, pgp-3, pgp-4, pgp-9, pgp-10, pgp-11, pgp-12 and pgp-14. These partial P-gp sequences from H. contortus were used to design and optimize a quantitative real-time PCR assay to investigate potential changes in the expression levels of P-gp transcripts associated with drug resistance. No significant changes in P-gp mRNA expression levels were found in a rapidly selected ivermectin-resistant parasite isolate compared to its drug-sensitive parent, but the assay has the potential to be used on other isolates in the future to further investigate resistance-associated changes in P-gp gene expression.

The objectives of this study were to evaluate the contamination by eggs of Toxocara in sandy areas or grass lawns of outdoor recreation areas that are used by children, and the frequency of seroprevalence in children, from three cities of fewer than 45,000 inhabitants in Paraná, Brazil. From May 2005 to December 2007, five samples were taken from each of 13 sandy sites and 18 grass lawns, all from plazas and public schools. Blood samples from children aged 0–12 years were analysed by immunoassay for anti-Toxocara IgG. The soil samples were processed by floatation and sedimentation. Eggs of Toxocara spp. were present in 44.7% (38/85) of the samples from grassed areas and in 21.4% (15/70) of the sand samples. The lawns were 2.16 times more contaminated than the sand (P = 0.0009). However, the epidemiological variables showed no statistically significant difference between seropositive (36.8%; 130/353), and seronegative children. The rate of seropositivity was higher in children aged 0–5 years (P = 0.03), who were 1.94 times more likely to develop persistent wheezing (P = 0.02).

There is an urgent need for information on schistosomiasis in preschool children, who are often excluded in mass treatment programmes. The prevalence and intensity of Schistosoma haematobium infection were determined in preschool children aged ≤ 6 years in two rural communities in Ijebu East, south-western Nigeria. Two urine samples each were collected from 83 preschool children from the two communities, tested for microhaematuria using reagent strips and then processed and examined with a microscope for S. haematobium eggs. Focus group discussions on perceptions of the disease and water contact practices were held in the communities with their guardians, caregivers and preschool children, using an interview guide. The prevalence of S. haematobium in the two communities was 14 (16.9%), with no significant differences (P = 0.661) in infection rate between boys (18.4%) and girls (14.7%). Both prevalence and intensity of infection did not increase significantly with age in both Korede and Obada community. However, there were significant differences in prevalence of infection between the two communities (P = 0.035). There was no association (P = 0.750) between intensity in boys (0.176 eggs/10 ml urine) and girls (0.110 eggs/10 ml urine). Focal group discussions with guardians and caregivers revealed that preschool children acquired infection early in their lives through exposure to infected stream water by their mothers, while the older children visit the stream for playing, bathing and swimming. It has therefore become imperative for preschool children to be included in the planning of schistosomiasis intervention programmes as a means of reducing transmission.

The ability of total homogenate (TH) of Fasciola hepatica conjugated with aluminium hydroxide (alum) or Freund's complete adjuvant (FCA) to protect cattle against experimental fasciolosis was evaluated. Compared with the infected group, the immunized animals with alum-TH and FCA-TH presented a significant reduction in fluke burden (85.9% and 96.8%, respectively), a higher percentage of short-sized worms, a marked reduction in the released eggs in faeces (89% and 57%, respectively), as well as an increased production of specific antibodies before infection. The alum-TH immunized group also showed a significant increase in the antigen-specific proliferation of peripheral blood mononuclear cells (PBMC) as early as 4 weeks before infection. Although both immunized groups (alum-TH and FCA-TH) were able to develop an efficient protective immune response to metacercarial challenge, an earlier PBMC response, lower hepatic damage and less effect on weight gain were found in alum-immunized animals. Therefore, alum is a good candidate for future immunization against bovine fasciolosis.