Early, microbially driven follicular reactions in the neonatal piglet do not contribute to expansion of the immunoglobulin heavy chain V-D-J repertoire

Selective microbial colonisation of germ-free piglets is reported to result in expansion of immunoglobulin VH- and DH-segment usage from an initially limited repertoire. Here, the response of the palatine tonsil to microbial colonisation was compared in age-matched conventionally reared and germ-free piglets. At 3 and 5 days after birth an expansion in the B-cell follicle area was observed in the conventional, microbially colonised animals, which was not seen in the germ-free piglets. Consistent with this observation, sequencing of re-arranged heavy chain V-D-J units demonstrated accumulation of point mutations indicating somatic hypermutation in the conventional, microbially colonised piglets but not in the germ-free animals. However, VH- and DH-segment usage and CDR3 length did not differ between the groups. The results suggest that the follicle reaction observed occurs in response to microbial challenge, involves proliferation and somatic hypermutation of B-cells but does not expand repertoire or generate classical, isotype-switched memory B-cells. We suggest that microbial colonisation of neonatal piglets drives immunological competence in two stages: first, an antigen non-specific, follicular reaction which expands immunological compartments; and second, microbe driven changes in V-segment usage which expand immunological repertoire.