Amoebic gill disease and host response in Atlantic salmon (Salmo salar L.): A review.

Gill health is one of the main health challenges for Atlantic salmon (Salmo salar L.) mariculture worldwide, and amoebic gill disease (AGD), caused by the marine ectoprotozoan Neoparamoeba perurans, is currently one of the most significant diseases in terms of prevalence and economic impact. This review describes the host response of Atlantic salmon to the disease, focusing on the pathological changes, immune response and mechanisms underlying the prominent epithelial proliferation and mucus hypersecretion occurring in affected fish. Health management strategies and risk factors are also discussed.

Repeat-enriched proteins are related to host cell invasion and immune evasion in parasitic protozoa.

Proteins containing repetitive amino acid domains are widespread in all life forms. In parasitic organisms, proteins containing repeats play important roles such as cell adhesion and invasion and immune evasion. Therefore, extracellular and intracellular parasites are expected to be under different selective pressures regarding the repetitive content in their genomes. Here, we investigated whether there is a bias in the repetitive content found in the predicted proteomes of 6 exclusively extracellular and 17 obligate intracellular protozoan parasites, as well as 4 free-living protists. We also attempted to correlate the results with the distinct ecological niches they occupy and with distinct protein functions. We found that intracellular parasites have higher repetitive content in their proteomes than do extracellular parasites and free-living protists. In intracellular parasites, these repetitive proteins are located mainly at the parasite surface or are secreted and are enriched in amino acids known to be part of N- and O-glycosylation sites. Furthermore, in intracellular parasites, the developmental stages that are able to invade host cells express a higher proportion of proteins with perfect repeats relative to other life cycle stages, and these proteins have molecular functions associated with cell invasion. In contrast, in extracellular parasites, degenerate repetitive motifs are enriched in proteins that are likely to play roles in evading host immune response. Altogether, our results support the hypothesis that both the ability to invade host cells and to escape the host immune response may have shaped the expansion and maintenance of perfect and degenerate repeats in the genomes of intra- and extracellular parasites.

Cloning and functional characterisation of a peroxiredoxin 1 (NKEF A) cDNA from Atlantic salmon (Salmo salar) and its expression in fish infected with Neoparamoeba perurans.

Peroxiredoxin 1 (Prx 1), also known as natural killer enhancing factor A (NKEF A), has been implicated in the immune response of both mammals and fish. Amoebic gill disease (AGD), caused by Neoparamoeba perurans, is a significant problem for the Atlantic salmon (Salmo salar L.) aquaculture industry based in Tasmania, Australia. Here we have cloned and functionally characterized a Prx 1 open reading frame (ORF) from Atlantic salmon liver and shown that Prx 1 gene expression was down-regulated in the gills of Atlantic salmon displaying symptoms of AGD. The Prx 1 ORF encoded all of the residues and motifs characteristic of typical 2-Cys Prx proteins from eukaryotes and the recombinant protein expressed in Escherichia coli catalyzed thioredoxin (Trx)-dependent reduction of H(2)O(2), cumene hydroperoxide (CuOOH) and t-butyl hydroperoxide (t-bOOH) with K(m) values of 122, 77 and 91 µM, respectively, confirming that it was a genuine 2-Cys Prx. The recombinant protein also displayed a double displacement reaction mechanism and a catalytic efficiency (k(cat)/K(m)) with H(2)O(2) of 1.5 × 10(5) M(-1) s(-1) which was consistent with previous reports for the 2-Cys Prx family of proteins. This is the first time that a Prx 1 protein has been functionally characterized from any fish species and it paves the way for further investigation of this important 2-Cys Prx family member in fish.

Identification of antigenic targets for immunodetection of Balamuthia mandrillaris infection.

The free-living amoeba Balamuthia mandrillaris causes granulomatous amoebic encephalitis (GAE) in humans. Rapid identification of balamuthiasis is critical for effective therapeutic intervention and case management. In the present study we identified target antigens for the development of a serological assay for B. mandrillaris infection. We demonstrated by silver staining that protein profiles for all eight isolates of B. mandrillaris, independent of human or animal origin or geographic origin, appeared to be similar except for some minor differences, indicating the molecular homogeneity of these strains. The profiles of all isolates, which ranged from 200 to 10 kDa, were similar, with a prominent protein visible around 30 kDa; all appeared considerably different from protein profiles of the control E6 cells and Acanthamoeba castellanii and Naegleria fowleri isolates. Western blot analysis with rabbit hyperimmune serum identified the major immunodominant antigens of 25, 50, 75, and 80 kDa; positive human sera reacted strongly with proteins around 25, 40, 50, and 75 kDa. Proteins around 40 kDa detected by human serum were not recognized by hyperimmune rabbit serum. None of the target proteins were detected by uninfected control sera. Reactivities of five patients' sera with 4 different isolates of B. mandrillaris (2 strains of human and 2 strains of animal origins) revealed that patients' sera reacted slightly differently with different B. mandrillaris isolates, although major proteins of approximately 25, 50, and 75 kDa were present in all extracts.

Notes from the field: transplant-transmitted Balamuthia mandrillaris --- Arizona, 2010.

On August 23, 2010, CDC was notified regarding two organ transplant recipients in Arizona who had encephalitis with multiple ring-enhancing lesions revealed by cerebral magnetic resonance imaging. The common organ donor, a Hispanic male landscaper aged 27 years, had died in Arizona from a presumed stroke on July 21. He had a large skin lesion for approximately 6 months on his back that he had attributed to an insect bite. The ill recipients, a male liver recipient aged 56 years, and a male recipient of a kidney and pancreas aged 24 years, received organ transplants on July 22. In addition, two other recipients received organs from this donor: an adult male heart recipient received his transplant in California on July 22, and an adult male kidney recipient received his transplant in Utah on July 23.

Characterisation of an immunodominant, high molecular weight glycoprotein on the surface of infectious Neoparamoeba spp., causative agent of amoebic gill disease (AGD) in Atlantic salmon.

Amoebic gill disease can be experimentally induced by the exposure of salmonids to Neoparamoeba spp. freshly isolated from infected fish, while cultured amoebae are non-infective. Results from our previous work suggested that one key difference between infectious and non-infectious Neoparamoeba were the highly glycosylated molecules in the glycocalyx. To characterise these surface glycans or glycoproteins we used a monoclonal antibody (mAb 44C12) specific to a surface molecule unique to infective parasites. This mAb recognised a carbohydrate epitope on a high molecular weight antigen (HMWA) that make up 15-19% of the total protein in a soluble extract of infectious parasites. The HMWA consisted of at least four glycoprotein subunits of molecular weight (MW) greater than 150 kDa that form disulfide-linked complexes of MW greater than 600 kDa. Chemical deglycosylation yielded at least four protein bands of approximate MW 46, 34, 28 and 18 kDA. While a similar HMWA complex was present in non-infective parasites, the glycoprotein subunits were of lower MW and exhibited differences in glycosylation. The four glycoproteins subunits recognised by mAb 44C12 were resistant to degradation by PNGase F, PNGase A, O-glycosidase plus ß-1, 4-galactosidase, ß-N-acetylglucosaminidase and neuraminidase. The major monosaccharides in the HMWA from infectious parasites were rhamnose, fucose, galactose, and mannose while sialic acids were absent. The carbohydrate portion constituted more than 90% of the total weight of the HMWA from infectious Neoparamoeba spp. Preliminary results indicate that immunisation of salmon with HMWA does not lead to protection against challenge infection; rather it may even have an immunosuppressive effect.

Balamuthia and Acanthamoeba-binding antibodies in West African human sera.

Little is known about the prevalence of Balamuthia mandrillaris amoebae and Balamuthia amoebic encephalitis in Africa. As an approach, relative concentrations of amoebae-binding serum antibodies (Ab) were assessed by flow cytometry using formaldehyde-fixed B. mandrillaris, Acanthamoeba lenticulata 72-2 and Acanthamoeba castellanii 1BU amoebae for specific Ab capture (B.m.-Ab, A.l.-Ab, A.c.-Ab). One hundred and ninety-two sera from West African (Côte d'Ivoire) donors aged 11-95years (mean 38 a; 51% males), and living in villages surrounded by rainforest near the Liberian border, were tested and related to reference sera from Berlin. While B.m.-Ab tended to increase with donor age, A.l.-Ab and A.c.-Ab did not. Accordingly, B.m.-Ab correlated only weakly with A.l.-Ab or A.c.-Ab. Of the nine individuals with the highest B.m.-Ab concentrations, most were elderly (mean 58 a), male (78%), and professed intensive outdoor activity (hunting, farming). Only three of these sera also showed elevated A.l.-Ab, and none elevated A.c.-Ab.