A natural infection by the red sea bream iridovirus-type Megalocytivirus in the golden mandarin fish Siniperca scherzeri.

An outbreak of a Megalocytivirus infection was found in the golden mandarin fish Siniperca scherzeri during September and October 2016, in Korea. Phylogeny and genetic diversity based on the major capsid protein (MCP) and adenosine triphosphatase (ATPase) genes showed a new strain. Designated as GMIV, this strain derived from the golden mandarin fish was suggested to belong to the red sea bream iridovirus (RSIV)-subgroup I. Additionally, this train clustered with the ehime-1 strain from red sea bream Pagrus major in Japan and was distinguished from circulating isolates (RSIV-type subgroup II and turbot reddish body iridovirus [TRBIV] type) in Korea. The infection level, evaluated by qPCR, ranged from 8.18 × 102 to 7.95 × 106  copies/mg of tissue individually, suggesting that the infected fish were in the disease-transmitting stage. The diseased fish showed degenerative changes associated with cytomegaly in the spleen as general sign of Megalocytivirus infection. The results confirm that the RSIV-type Megalocytivirus might have crossed the environmental and species barriers to cause widespread infection in freshwater fish.

Contribution of the CD5+ B cell to D-proximal VH family expression early in ontogeny.

In this study, the contribution of the CD5+ B cell to the preferential expression of VH 7183 and Q52 observed early in development was determined. CD5+ and CD5- B cells from BALB/c mice were isolated by fluorescence-activated cell sorter and the expression of particular VH gene families was determined directly by in situ hybridization. The results indicate that CD5+ B cells obtained from both adult and neonatal animals express Q52 at increased levels compared with CD5- B cells. Preferential expression of VH 7183 was observed only in the neonatal CD5- B cell subset. Thus, the increased expression of VH 7183 early in development is caused by the CD5- subset whereas increased Q52 expression is caused by the CD5+ subset. These results indicate that the fetal/neonatal conventional B cell is distinct from conventional adult B cells in terms of Ig gene repertoire expression.

Comparison of the fetal and adult functional B cell repertoires by analysis of VH gene family expression.

The functional B cell repertoire in BALB/c mice was assessed at various stages in ontogeny. This was done by analyzing VH gene family expression using the sensitive technique of in situ hybridization. The B cell repertoire was probed with the mitogen, LPS, and the antigen DNP. DNP was chosen because B cells responsive to this hapten appear very early in ontogeny. The APCs that developed after stimulation with LPS or DNP were analyzed for VH gene expression by in situ hybridization of individual cells using radiolabeled VH gene family probes. The results indicated that VH gene expression in fetal B cells after stimulation was distinct from adult B cells in that there was a biased expression of D proximal families. The results indicated that this bias was associated with developmental age and not a given differentiation stage in the B cell lineage. In addition, stimulation of fetal B cells with DNP resulted in a large increase in expression of member(s) of VH 36-60, suggesting that the early appearance of DNP-responsive B cells is not strictly correlated with preferential rearrangement of D proximal families, VH 7183 and VH Q52. However, the results suggested that a large proportion of pre-B cells that preferentially rearrange D proximal families early in ontogeny become part of the functional developing repertoire.

Strain-dependent expression of VH gene families.

The tremendous diversity of the antibody specificity repertoire stems from the ability of each developing B cell to select one out of many possible variable, diversity, and joining gene segments by specific rearrangement of the DNA. The mechanism by which V region gene segments is selected is not known. Moreover, evidence for both random and nonrandom expression of VH genes in mature B cells has been presented previously. In this report, the technique of in situ hybridization is used to accurately measure at the single cell level VH gene family expression in LPS-induced cells from several strains. In this way, at least one-third of the B cells are stimulated and a large sampling of activated splenocytes from each strain analyzed. The use of in situ hybridization eliminates any potential biases resulting from transformation protocols. In addition, because all populations of cells are analyzed by both in situ hybridization and immunocytochemical staining with anti-IgM, the proportion of cells detected by in situ hybridization could be compared with the proportion of B cells, blasts, and plasma cells in the population. It was concluded from these comparisons that the cells being detected by in situ hybridization under the conditions described are plasmablasts and plasma cells. Therefore, an accurate measure of the functional and expressed VH gene repertoire could be made. The results clearly demonstrate strain-dependent variation in VH gene family expression, particularly VH 7183 and VH J558 with up to three-fold differences observed. Thus, either there is considerable strain variation in the number of functional VH gene family segments or the expression of VH genes is not entirely random.