Characterization of a Panel of Cross-Reactive Hantavirus Nucleocapsid Protein-Specific Monoclonal Antibodies.

Hantaviruses are emerging pathogens with a worldwide distribution that can cause life-threatening diseases in humans. Monoclonal antibodies (MAbs) against hantavirus nucleocapsid (N) proteins are important tools in virus diagnostics, epidemiological studies and basic research studies on virus replication and pathogenesis. Here, we extend the collection of previously generated MAbs raised against a segment of Puumala orthohantavirus (PUUV) N protein harbored on virus-like particles (VLPs) and MAbs against N proteins of Sin Nombre orthohantavirus/Andes orthohantavirus by generating nine novel MAbs against N proteins of Dobrava-Belgrade orthohantavirus (DOBV), Tula orthohantavirus (TULV), Thottapalayam thottimvirus (TPMV) and PUUV. In order to have a wide collection of well-described hantavirus-specific MAbs, the cross-reactivity of novel and previously generated MAbs was determined against N proteins of 15 rodent- and shrew-borne hantaviruses by different immunological methods. We found that all MAbs, excluding TPMV-specific MAbs, demonstrated different cross-reactivity patterns with N proteins of hantaviruses and recognized native viral antigens in infected mammalian cells. This well-characterized collection of cross-reactive hantavirus-specific MAbs has a potential application in various fields of hantavirus research, diagnostics and therapy.

Influence of chronic exposure to treated sewage effluent on the distribution of white blood cell populations in rainbow trout (Oncorhynchus mykiss) spleen.

Impairment of immune function in aquatic animals has been proposed as a possible consequence of low-level contamination of surface waters with anthropogenic substances such as through the discharge of wastewater into rivers, lakes, and oceans. The study at hand investigated the effects of chronic (32 weeks) exposure to sewage treatment plant (STP) effluent on the prevalence and distribution of different leucocyte populations in spleen samples of rainbow trout (Oncorhynchus mykiss). To simulate an infection, fish were injected intraperitoneally (ip) with inactivated Aeromonas salmonicida salmonicida, 6 weeks prior to the termination of the experiment. Immunohistological analysis revealed a marked decrease in thrombocyte numbers, an increase of monocytes, altered distribution of B-cells, and higher surface immunoglobulin expression, as well as activation of MHC class II expression in the spleen after exposure to 15% (v/v) effluent. The most prominent finding of the present study, however, was the occurrence of intraplasmatic deposits or inclusions with strong autofluorescence in spleen sections from effluent-exposed trout. In addition to effluent effects, injection of trout with A. salmonicida stimulated infiltration of monocytes, increased staining intensity on thrombocytes, and enhanced MHC class I expression in larger leucocytes surrounding melanomacrophage centres. In general, the current study demonstrates a marked, potentially adverse effect of STP effluent on spleen leucocytes and on the integrity of spleen tissue. The observed response suggests a constant unspecific stimulation of different leucocyte populations and is reminiscent of chronic inflammation.

Contrasted TCRß diversity of CD8+ and CD8- T cells in rainbow trout.

Teleost fish express highly diverse naive TCRß (TRB) repertoires and mount strong public and private clonal responses upon infection with pathogens. Fish T cells express typical markers such as CD8, CD4-1 and CD4-2, CD3, CD28 and CTLA4. Fish CD8(+) T cells have been shown to be responsible for antigen-specific cell-mediated cytotoxicity in in vitro systems using histo-compatible effector and target cells. We compare here the complexity of TRB repertoires between FACS sorted CD8(+) and CD8(-) T cells from spleen and pronephros of rainbow trout. In contrast to human, while the TRB repertoire is highly diverse and polyclonal in CD8(+) T cells of naïve fish, it appeared very different in CD8(-) lymphocytes with irregular CDR3 length distributions suggesting a dominance of activated clones already in naïve fish or the presence of non conventional T cells. After infection with a systemic virus, CD8(+) T cells mount a typical response with significant skewing of CDR3 length profiles. The infection also induces significant modifications of the TRB repertoire expressed by the CD8(-) fraction, but for a different set of V/J combinations. In this fraction, the antiviral response results in an increase of the peak diversity of spectratypes. This unusual observation reflects the presence of a number of T cell expansions that rise the relative importance of minor peaks of the highly skewed distributions observed in unchallenged animals. These results suggest that the diversity of TRB expressed by CD8(+) and CD8(-) αß T cells may be subjected to different regulatory patterns in fish and in mammals.