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B chromosomes (B) are supernumerary, additional and non-homologous to standard (A) chromosomes. Significant biological questions concerning the origin and structural organization of Bs are under discussion. Previous reports found that B chromosomes are enriched with repetitive elements and can carry functional genes. The present study is aimed to extend our knowledge of the structure and composition of B chromosomes. We have combined cytogenetics and genomics approaches to undertsand the B chromosome problem. Cytogenetics analysis, applied to cichlid fish Astatotilapia latifasciata , revealed the presence of 1 or 2 B chromosomes. Next generation sequencing data analysis, based on differential coverage approach, among B- (individual without B) and B+ (individual with B) samples, detected a higher number of copies for the Indian Hedgehog b (Ihhb) gene and 45S RNA transcriptional gene-cluster. Fluorescence in situ hybridization (FISH) confirmed that these genes are located on the B chromosome. In addition, the excessively abundant and strong signals of IHHB on B shows that this gene has accumulated and amplified its copies due to frequent duplication events. The findings also demonstrate that B chromosome have incorporated the entire 45S RNA cluster (18S ribosomal RNA, internal transcribed spacer 1, 5.8S ribosomal RNA, internal transcribed spacer 2, and 28S ribosomal RNA) from the A complement set. Our study contributes to explain that these genic sequences are vital components in constitution of B chromosomes.

FISH mapping of 45S rRNA and IHHB genes on autosomes and B chromosome of cichlid fish Astatotilapia latifasciata