This paper documents a difference in isotopic compositions between Upper Cambrian microbial and non-microbial micritic deposits and proposes implications for diagenesis of calcimicrobial deposits. The δ18O values (−10.98 to −8.71; average −9.88 ‰ VPDB) of calcimicrite comprising shallow subtidal microbialites from the southern Appalachians are more negative than: (1) the calcimicrite from associated subtidal non-microbial deposits (−8.98 to −7.16; average −7.82), suggesting a different diagenetic history; and (2) estimates of Late Cambrian marine calcite values (−5 to −3 ‰), indicating postdepositional modifications. Early diagenetic calcification of microbial deposits promoted the formation of growth cavities and borings rimmed with marine fibrous and prismatic calcite cement. Some of the voids remained open and provided pathways for fluids during later diagenesis. The microbial deposits, therefore, experienced more pronounced diagenetic alteration than the less porous non-microbial micritic deposits. The δ18O compositions provided invaluable insights into the influence of early lithification on the later diagenesis of microbialites and into the processes that can result in poor preservation of syndepositional marine isotope signatures in these deposits.