Molecular phylogeny and ultrastructure of Myxobolus cf. cuneus, a parasite of patinga hybrid and Henneguya pseudoplatystoma, a parasite of pintado hybrid.

Through morphological, histopathological and ultrastructural analysis of Myxobolus cuneus Adriano, Arana et Cordeiro, 2006 and Henneguya pseudoplatystoma Naldoni, Arana, Maia, Ceccarelli, Tavares, Borges, Pozo et Adriano, 2009 were identified infecting pacu respectively (Piaractus mesopotamicus) and hybrid pintado (Pseudoplatystoma corruscans x Pseudoplatystoma reticulatum) taken from Brazilian fish farms. The present study describes 18S rDNA sequencing of Myxobolus cf. cuneus infecting the spleen of farmed patinga, a hybrid fish resulting from the crossing of P. mesopotamicus x Piaractus brachypomus, and H. pseudoplatystoma found in farmed hybrid pintado from the state of Sao Paulo, Brazil. The study also provides new details of the host-parasite interface of M. cf. cuneus, which reveal that the plasmodial wall is composed of a single membrane connected to the plasmodium ectoplasm by numerous pinocytic canals. The plasmodia also displayed asynchronous development but had disporic pansporoblasts at different developmental stages; immature and mature spores were found at different depth levels of the plasmodium. Maximum likelihood phylogenetic analysis showed that M. cf. cuneus appeared as a sister species of Henneguya pellucida Adriano, Arana et Cordeiro, 2005 in a sub-clade composed mainly of myxosporean parasites of characiforms, and that H. pseudoplatystoma clustered in a sub-clade composed of Henneguya/Myxobolus spp. parasites of siluriform fish.

Supplementary data of Henneguya leporinicola (Myxozoa, Myxosporea) a parasite of Leporinus macrocephalus from fish farms in the state of São Paulo, Brazil.

Henneguya leporinicola is a parasite of the gill filament of Leporinus macrocephalus, a characiform fish belonging to the Anostomidae family, which is of major economic importance. Despite the damage it causes in fish, little is known about this parasite. Therefore, a study was undertaken with fourteen specimens of L. macrocephalus taken from fish farms in the state of Sao Paulo. The fish were collected and examined searching for lesions and/or myxosporean plasmodia. One of the specimens (7.14%) contained white elongated plasmodia in the gill filament. The mature spores had elongated bodies with polar capsules of equal size and a caudal length greater than body length. Morphological characteristics identified the parasite as H. leporinicola. Molecular analysis of the 18S rDNA sequence resulted in a 1954 bp, demonstrating significant genetic differences with previously described species of Henneguya/Myxobolus. Phylogenetic analysis comparing the 18S rDNA sequence of H. leporinicola with other species, previously described in South America, and the 20 closest species as indicated by BLASTn Max Score showed H. leporinicola as a basal branch of a subclade composed by Henneguya spp. parasite of characiform hosts.

Prevalence, intensity, and phylogenetic analysis of Henneguya piaractus and Myxobolus cf. colossomatis from farmed Piaractus mesopotamicus in Brazil.

Henneguya piaractus and Myxobolus colossomatis (Myxosporea: Myxobolidae) are commonly found in the characid Piaractus mesopotamicus, an important fish farm species in Brazil. This paper describes the prevalence, mean intensity, molecular phylogeny, ultrastructure, and histology of H. piaractus and M. cf. colossomatis found infecting specimens of P. mesopotamicus collected from fish farms in the state of São Paulo, Brazil. A total of 278 fish were collected from 3 fish farms between February 2008 and July 2010. Parasite prevalence and mean intensity varied throughout the study period, and according to location and year. A phylogenetic tree, placing South American species in a global context, showed a clear tendency among myxosporean species to cluster according to host families. Ultrastructural analysis for M. cf. colossomatis showed the plasmodial wall with numerous projections toward host cells and phagocytic activity. Histopathological data showed hyperplasia caused by H. piaractus in highly infected fish. Histological and ultrastructural analysis of H. piaractus showed results similar to those that have previously been reported.

Haplotypes of the bovine IgG2 heavy gamma chain in tick-resistant and tick-susceptible breeds of cattle.

Bovines present contrasting, heritable phenotypes of infestations with the cattle tick, Rhipicephalus (Boophilus) microplus. Tick salivary glands produce IgG-binding proteins (IGBPs) as a mechanism for escaping from host antibodies that these ectoparasites ingest during blood meals. Allotypes that occur in the constant region of IgG may differ in their capacity to bind with tick IGBPs; this may be reflected by the distribution of distinct allotypes according to phenotypes of tick infestations. In order to test this hypothesis, we investigated the frequency of haplotypes of bovine IgG2 among tick-resistant and tick-susceptible breeds of bovines. Sequencing of the gene coding for the heavy chain of IgG2 from 114 tick-resistant (Bos taurus indicus, Nelore breed) and tick-susceptible (B. t. taurus, Holstein breed) bovines revealed SNPs that generated 13 different haplotypes, of which 11 were novel and 5 were exclusive of Holstein and 3 of Nelore breeds. Alignment and modeling of coded haplotypes for hinge regions of the bovine IgG2 showed that they differ in the distribution of polar and hydrophobic amino acids and in shape according to the distribution of these amino acids. We also found that there was an association between genotypes of the constant region of the IgG2 heavy chain with phenotypes of tick infestations. These findings open the possibility of investigating if certain IgG allotypes hinder the function of tick IGBPs. If so, they may be markers for breeding for resistance against tick infestations.

Recombinant expression of Taenia solium TS14 antigen and its utilization for immunodiagnosis of neurocysticercosis.

In order to evaluate the potential use of TS14 antigen in an enzyme-linked immunosorbent assay (ELISA) for immunodiagnosis of neurocysticercosis (NC), its open reading frame (ORF) was amplified by RT-PCR from mRNA isolated from Taenia solium cysticerci. The ORF was subcloned into the expression vector pET-28a, and was used to transform Escherichia coli BL21 (DE3) cells to produce TS14 antigen. The His-tagged expressed protein was purified on a nickel affinity column. Using the HISTS14 as antigen, ELISA was positive for 100% of cerebrospinal fluid (CSF) and 97% of serum samples from NC patients. No positive results were observed with sera and CSF samples from control groups. Cross-reactivity with sera from patients with schistosomiasis and Chagas' disease was not observed. Serum samples from patients with taeniasis were evaluated and 2 of 13 cases showed reactivity in this assay. Our data indicate the usefulness of HISTS14 in ELISA for an accurate and rapid assay for diagnosis of NC and seroepidemiological studies.

Effect of Cysticercus cellulosae fractions on the respiratory burst of pig neutrophils.

Neutrophils, eosinophils and macrophages are cells that interact with invading parasites and naive hosts have been shown to have anti-parasitic activity. The initial reaction of these leukocytes is the generation of reactive oxygen species (ROS) to play in parasite expulsion. The present work was carried out to study the effect of total extract, scolex and membrane fractions from Cysticercus cellulosae on respiratory burst by pig neutrophils. Hydrogen peroxide (H2O2) production by neutrophils incubated with metacestode fractions from C. cellulosae showed an increase of: 190% (total extract), 120% (scolex) and 44% (membrane). High antioxidant catalatic activity (33%, 28%, 28% by total extract, scolex and membrane, respectively) was observed in neutrophils incubated with metacestode fractions, which could be an attempt at self-protection. Scolex and membrane fractions increased the phagocytic capacity of neutrophils (44% and 28%, respectively). On the other hand, total cysticerci did not alter the phagocytosis, possibly due to modifications in membrane function, caused by high ROS production from neutrophils in the presence of total cysticerci. Total fraction from C. cellulosae is toxic for neutrophils as shown by the decrease in phagocytic capacity, probably caused by high levels of ROS formation. The difference in toxicity of total extract, scolex and membrane fractions on neutrophils can be explained by the presence of an antigenic effect of the vesicular fluid in the total extract of C. cellulosae.

Effect of Cysticercus cellulosae fractions on the respiratory burst of pig neutrophils

Neutrófilos, eosinófilos e macrófagos são células que interagem com os parasitas no corpo do hospedeiro desenvolvendo atividade antiparasitária. A reação inicial destes leucócitos é a geração de espécies reativas de oxigênio (ERO) a fim de expulsar os parasitas. No presente trabalho estudou-se o efeito da fração total, de escolex e de membrana de Cysticercus cellulosae sobre a explosão respiratória de neutrófilos de suínos. A produção de peróxido de hidrogênio (H2O2) pelos neutrófilos incubados com as frações de C. cellulosae apresentou acréscimo de 190% (extrato total), 120% (escolex) e 44% (membrana). Alta atividade de catalase (33%, 28% e 28% para extrato total, escolex e membrana respectivamente) foi observada nos neutrófilos incubados com as frações de metacestodeo, podendo representar a própria proteção celular do neutrófilo. Frações de escolex e de membrana aumentaram a capacidade fagocitária dos neutrófilos (44% e 28%, respectivamente). Por outro lado, a fração total do cisticerco não alterou a capacidade fagocitária dos neutrófilos, o que pode estar relacionada com modificações na função da membrana celular causadas pela alta produção de ERO na presença da fração total. O extrato total de C. cellulosae é tóxico para os neutrófilos, indicada pela diminuição da capacidade fagocitária, provavelmente pela indução de alto nível de ERO. A diferença de toxicidade do extrato total, de escolex e de membrana para os neutrófilos pode ocorrer pelo efeito antigênico presente no fluido vesicular no extrato total de C. cellulosae.