ERP128624 PRJEB44558 b3a96b61-5eab-4058-a9a8-63cdbe5fe031 Molecular systematics, higher-rank classification and Gondwanan origins of Cryptocephalinae leaf beetles With 5,300 species, the Cryptocephalinae is the fourth largest subfamily of the megadiverse Chrysomelidae beetles. This subfamily currently merges groups that were traditionally considered subfamilies of their own, including the fulcidacines, clytrines and cryptocephalines, and it is believed to be sister to the Lamprosomatinae, forming a natural group characterized by immature stages developing within the protection of a solid case of organic matter known as scatoshell. Nonetheless, the hypotheses on higher-level systematics of this diverse group and the recognition of its closest relatives have been nearly as varied as the number of authors who struggled to elaborate its system. But none of the hypotheses have been tested with phylogenetic approaches, save for the unpublished morphology-based conclusions of a 1990 PhD thesis. In this work, we provide the first molecular phylogenetic hypothesis for a large dataset of Cryptocephalinae (167 species, 66 genera) based on the analysis of five gene fragments, including up to 3488 aligned nucleotide positions of the mitochondrial DNA cox1 and rrnS, and the nuclearly encoded LSU, elongation factor 1-alpha and polyadenylate binding protein 1. The phylogeny confirms the sister group relationship between Cryptocephalinae and Lamprosomatinae and identifies the Eumolpinae as the sister to this assemblage. Moreover, it partly refutes the current system for Cryptocephalinae giving support to five tribes in the subfamily: Fulcidacini and Clytrini, matching the current system, Pachybrachini separated from Cryptocephalini, and a new subantarctic tribe, Mylassini trib. n., divergent from the other lineages. Among Clytrini, data support inclusion of Ischyopachina within New World Babiina, which would be sister to New World Megalostomina, and together sister to Old World Clytrina. In Cryptocephalini, after exclusion of pachybrachines, four subtribes are distinguished, of which only Stylosomina matches the current system. Otherwise, the current Monachulina are dismantled into three clades deserving subtribal status, one of which, former New World Monachulina genera, are unambiguously transferred to Cryptocephalina. A molecular clock calibrated using paleontological and biogeographic information places the origin of the subfamily in the Early Cretaceous and the split of the main evolutionary lineages temporally and geographically congruent with the fragmentation of major Gondwanan territories. undefined With 5,300 species, the Cryptocephalinae is the fourth largest subfamily of the megadiverse Chrysomelidae beetles. This subfamily currently merges groups that were traditionally considered subfamilies of their own, including the fulcidacines, clytrines and cryptocephalines, and it is believed to be sister to the Lamprosomatinae, forming a natural group characterized by immature stages developing within the protection of a solid case of organic matter known as scatoshell. Nonetheless, the hypotheses on higher-level systematics of this diverse group and the recognition of its closest relatives have been nearly as varied as the number of authors who struggled to elaborate its system. But none of the hypotheses have been tested with phylogenetic approaches, save for the unpublished morphology-based conclusions of a 1990 PhD thesis. In this work, we provide the first molecular phylogenetic hypothesis for a large dataset of Cryptocephalinae (167 species, 66 genera) based on the analysis of five gene fragments, including up to 3488 aligned nucleotide positions of the mitochondrial DNA cox1 and rrnS, and the nuclearly encoded LSU, elongation factor 1-alpha and polyadenylate binding protein 1. The phylogeny confirms the sister group relationship between Cryptocephalinae and Lamprosomatinae and identifies the Eumolpinae as the sister to this assemblage. Moreover, it partly refutes the current system for Cryptocephalinae giving support to five tribes in the subfamily: Fulcidacini and Clytrini, matching the current system, Pachybrachini separated from Cryptocephalini, and a new subantarctic tribe, Mylassini trib. n., divergent from the other lineages. Among Clytrini, data support inclusion of Ischyopachina within New World Babiina, which would be sister to New World Megalostomina, and together sister to Old World Clytrina. In Cryptocephalini, after exclusion of pachybrachines, four subtribes are distinguished, of which only Stylosomina matches the current system. Otherwise, the current Monachulina are dismantled into three clades deserving subtribal status, one of which, former New World Monachulina genera, are unambiguously transferred to Cryptocephalina. A molecular clock calibrated using paleontological and biogeographic information places the origin of the subfamily in the Early Cretaceous and the split of the main evolutionary lineages temporally and geographically congruent with the fragmentation of major Gondwanan territories. ENA-FIRST-PUBLIC 2021-10-19 ENA-LAST-UPDATE 2021-10-19