As participating molecules in nucleic acids, nucleotides also have signaling functions and act as second messengers in both prokaryotic and eukaryotic cells, the most common example being cyclic AMP (cAMP). In bacteria, a great interest has arisen in studying nucleotide signaling. It has been determined that cyclic di-AMP (c-di-AMP), cAMP, and cyclic di-GMP (c-di-GMP) participate in biological events such as bacterial growth, biofilm formation, sporulation, cell differentiation, motility, virulence, and other. Moreover, these cyclic-di-nucleotides (c-di-nucleotides) produced in pathogenic intracellular bacteria can affect eukaryotic host cells for infection. On the other hand, the nucleotide molecules pppGpp, and ppGpp are alarmones involved in regulating bacterial nutritional stress; they are also considered second messengers. These second messengers can be used as therapeutic agents because of their immunological functions on eukaryotic cells. In this review, different topics on the role of c-di-nucleotides as second messengers in different bacterial processes are addressed.