Codon usage bias refers to the phenomenon where synonymous codons are used with unequal frequencies. To understand the patterns of codon usage in mitochondrial cytochrome B (MT-CYB) gene of phylum platyhelminthes we used bioinformatic approaches to analyze the protein coding sequences of five different classes - cestoda, monogenea, rabditophora, trematoda and turbellaria. It was found from nucleotide composition analysis that in all the classes, A/T-ended codons were preferred to G/C -ended codons. From box plot analysis GC1 was found to have highest response to codon usage bias. Correspondence analysis indicated that besides mutation other factors such as natural selection might also affect the codon usage pattern. Neutrality plot reveals that both mutation and natural selection played role in codon usage pattern in five classes of MT-CYB gene. Various factors namely nucleotide composition, natural selection and mutation pressure affected the codon usage pattern.