Comparative analysis of mycobacterial infections in wild striped bass Morone saxatilis from Chesapeake Bay

During an ongoing epizootic of mycobacteriosis, wild striped bass Morone saxatilis from Chesapeake Bay were analyzed using 3 methods for detection of either mycobacterial infection or associated granulomatous pathology. The specific detection techniques, which utilized aseptically collected splenic tissue, were histology, quantitative culture and nested PCR. Based on analysis of 118 samples, detection of infection differed significantly between the 3 methods (chi-square, p = 0.0007). Quantitative culture and nested PCR detected similar, higher rates of infection (69 and 75%, respectively) than the histological method (52%). Although primary PCR assays for a 924 to 940 bp segment of the mycobacterial 16S rRNA gene were positive for genomic DNA from mycobacterial cultures, a secondary, nested PCR reaction for an internal 300 bp gene segment was required in order to detect mycobacteria within splenic tissue. A similar rate of mycobacterial infection was present in fish collected from all sites tested. Although all detection methods found that striped bass age 4.0 to 4.9 yr had the highest positive incidence, nested PCR detected a higher frequency of mycobacterial infection in fish > or = 6.0 yr of age than the other 2 methods. Quantitative bacteriology was a more sensitive detection technique when the fish tissue contained < or = 10(3) mycobacteria g(-1).     

Two new Ig VH gene families in Oncorhynchus mykiss

Genes encoding the immunoglobulin heavy-chain variable region (Ig VH) in rainbow trout (Oncorhynchus mykiss) have been grouped into 11 families. While obtaining a baseline assessment of the various gene families utilized by trout in the production of secreted antibody, we discovered two new families. These proposed Ig VH families, Families XII and XIII, were rarely observed; only two VH sequence types were detected for each new family, suggesting that they may not be commonly used in response to antigens, or that the captive environment may not lead to typical exposures seen in the wild. Additionally, unlike preceding studies, we found at least one representative gene sequence for each of the 11 reported Ig VH gene families, possibly indicating that the repertoire of trout Ig VH gene families may be more universal among different stocks than previously realized. GenBank accession numbers: Family XII—DQ453185 and DQ453150; Family XIII—DQ453153 and DQ453146; others DQ453143, DQ453156, DQ831723, DQ831825.     

Cellular mechanisms of glucocorticoid immunosuppression in salmon

Limiting dilution analysis was employed to determine the effect of cortisol and conditioned media on precursor frequency and clone size of antibody-producing cells sensitive to trinitrophenylated-lipopolysaccharide (TNP-LPS). This analysis demonstrated that the restoration of the cortisol-suppressed antibody response by conditioned media occurs through the activation of previously inhibited B cell precursors to that antigen, and not by the recruitment of other antigen-insensitive B cell precursors.     

Analysis of the effects of Perkinsus marinus proteases on plasma proteins of the Eastern oyster (Crassostrea virginica) and the Pacific oyster (Crassostrea gigas).

We employed two in vitro buffer systems to determine the potential pathogenic effects of Perkinsus marinus serine proteases on the plasma proteins of the eastern oyster (Crassostrea virginica) and the Pacific oyster (Crassostrea gigas). Specifically, this study characterized the oyster plasma protein targets of P. marinus proteases. Additionally, protease-specific inhibitory activity was revealed upon comparison of artificial (PBS) and endogenous (plasma-based) diluents employed during protease digestions. It was found that a C. virginica plasma protein of approximately 35 kDa was eliminated when a standard buffer (PBS) was used as a diluent; however, this protein was preserved when a low-molecular-weight, plasma-based, diluent was used. The results strongly indicate that low-molecular-weight inhibitors of P. marinus proteases are present in oyster plasma. A control (nonparasitic) serine protease, alpha-chymotrypsin, was employed to ascertain the specificity of the protease inhibitors. Although alpha-chymotrypsin possesses ample proteolytic activity for C. virginica plasma proteins, the anti-proteases could specifically inhibit only P. marinus proteases. Such specificity of anti-protease activity is not uncommon among low-molecular-weight serine proteases. The hemolymph target protein was isolated by 2D electrophoresis and isoelectrically isolated for further characterization by N-terminal amino acid sequencing.     

The impacts of four potential bioenergy crops on soil carbon dynamics as shown by biomarker analyses and DRIFT spectroscopy

Perennial bioenergy crops accumulate carbon (C) in soils through minimally disturbing management practices and large root inputs, but the mechanisms of microbial control over C dynamics under bioenergy crops have not been clarified. Root‐derived C inputs affect both soil microbial contribution to and degradation of soil organic matter resulting in differing soil organic carbon (SOC) concentrations, storage, and stabilities under different vegetation regimes. Here, we measured biomarker amino sugars and neutral sugars and used diffuse reflectance mid‐infrared Fourier transform spectroscopy (DRIFTS) to explore microbial C contributions, degradation ability, and SOC stability, respectively, under four potential bioenergy crops, M.×giganteus (Miscanthus × giganteus), switchgrass (Panicum virgatum L.), a mixed prairie, and a maize (Zea mays L.)–maize–soybean (Glycine max(L.) Merr.) (MMS) rotation over six growing seasons. Our results showed that SOC concentration (g/kg) increased by 10.6% in mixed prairie over the duration of this experiment and SOC storage (Mg/ha) increased by 17.0% and 15.6% in switchgrass and mixed prairie, respectively. Conversion of row crops to perennial grasses maintained SOC stability and increased bacterial residue contribution to SOC in M.×giganteus and switchgrass by 20.0% and 15.0%, respectively, after 6 years. Degradation of microbe‐derived labile SOC was increased in M.×giganteus, and degradation of both labile and stable SOC increased in MMS rotation. These results demonstrate that microbial communities under perennial grasses maintained SOC quality, while SOC quantity increased under switchgrass and mixed prairie. Annual MMS rotation displayed decreases in aspects of SOC quality without changes in SOC quantity. These findings have implications for understanding microbial control over soil C quantity and quality under land‐use shift from annual to perennial bioenergy cropping systems.     

Antibody structural variation in rainbow trout fluids

Rainbow trout (Oncorhynchus mykiss) were immunized with trinitrophenylated-keyhole limpet hemocyanin (TNP-KLH) and the redox structure of induced anti-TNP antibodies from the serum, mucus, egg and ovarian fluid was examined. In conducting these studies it was determined that all TNP-specific antibody from each source possessed the mAb-specific H chain (1-14) epitopes, which facilitated the direct structural analysis of the induced antibodies. A protocol was developed which ensured complete adsorption of all specific anti-TNP antibody from each fluid. Together these protocols permitted the unbiased compositional analysis of all redox forms of the anti-TNP antibodies from each source. All antibodies, regardless of source, possessed the same molecular mass, characteristic of the trout tetramer (800 kDa). It was found that specific antibody titers were significantly higher in male than female trout, while the degree of disulfide polymerization was relatively invariant in male antibodies, while being highly variable in female antibodies. Within the females, no distinctively different redox ratios were between antibodies isolated from sera, ovarian fluid or eggs: however, mucus antibodies possessed a unique redox structure consisting of halfmeric constituents that were not observed in antibodies from other fluids.