Lipoteichoic acids (LTAs) are membrane-anchored molecules in the cell envelopes of Gram-positive bacteria. Until recently, they were considered to be restricted to the Firmicutes, which include important pathogens such as Staphylococcus aureus and Streptococcus pneumoniae. Polyanionic LTAs have fundamentally important roles in divalent cation retention within the Gram-positive cell envelope and thereby influence bacterial cell division. Thus, LTA biosynthesis provides an attractive target for the development of novel antimicrobial interventions. Recent studies, notably two investigations of S. aureus and another of Bacillus subtilis, have greatly improved our understanding of the genetic basis of LTA biosynthesis. In addition, reports have revealed that at least some members of the Actinobacteria (another phylum of Gram-positive bacteria) produce LTAs, rather than the lipoglycans previously assumed to be typical of this taxon. The availability of whole bacterial genome sequences has enabled us to perform comparative analyses to shed light on the distribution of putative LTA biosynthetic genes among bacteria. Here, we discuss the results of these genomic analyses, together with the current literature, and propose that LTA biosynthesis in Actinobacteria might be fundamentally different to that in most Firmicutes.