Kenaf (Hibiscus cannabinus), a native warm-seasonal crop in Africa, is being considered for genetic improvement for local bast fiber production. To expedite its genetic improvement through breeding, kenaf genotypes from Ghana were assessed for genomic diversity regarding their chromosomal composition and ploidy levels. To gain insight into the repetitive DNA fractions in kenaf, the organization of 5S and 35S rRNA genes, as well as telomeric signal patterns were studied by a molecular cytogenetic approach. Using multi-color fluorescent in situ hybridization, distinct rDNA loci and Arabidopsis-type telomeres were revealed. The 5S rRNA genes were conserved in kenaf and localized in interstitial regions of two chromosomes across all accessions. The 35S rRNA genes were variable across the kenaf accessions and localized at sub-terminal ends and rarely interstitially in eight or six chromosome arms. Telomeric signals were observed at terminal ends of all chromosomes, with smaller signals also interstitially. The chromosome configuration of Ghana kenaf accessions was confirmed to be 2n=2x=36, each. We discuss the chromosomal variability and the likely genomic instability in the kenaf breeding lines from Ghana. To our knowledge, this is the first report on molecular cytogenetics on kenaf and thus, provides valuable insights into the genome of kenaf that will be useful for breeding. Additionally, this study provides a basis for further studies to analyze the repetitive DNA sequences and develop reference karyotypes to reveal genetic and evolutionary relationships between cultivated and wild Hibiscus species.