Culturomics, a high scale culture approach based on the multiplication of culture conditions, coupled with extended incubation times proved effective for discovering unknown microorganisms. While significant efforts were made to reduce the number of conditions, preliminary work suggested that strict anaerobic species do not necessarily require prolonged pre-incubation time. The aim of this study is to apply culturomics to faecal samples with one pre-incubation step, following very early and late subcultures. From the same stool specimen from a healthy donor, we first developed our rapid culturomics method by handling the sample Microbiological Safety Cabinet (MSC) and then validate our approach on the same sample but manipulated in anaerobic chamber. We used a single culture condition consisting of pre-incubation in an anaerobic blood culture bottle containing modified YCFA (Yeast Casitone Fatty Acids Agar), supplemented with blood and rumen fluid, followed by early (H0 -H24) and late (D2 -D30) subcultures. A total of 88 bacterial species were isolated during the initial experiment without anaerobic chamber, including Christensenella and Faecalibacterium species. Subculturing at H0 was the most diverse subculture time, and species considered as fastidious, including Coprococcus comes, Faecalibacterium massiliense and Ruminococcus torques, that were subcultured immediately following pre-incubation. We then reduced the less fertile subculture times by reducing their number from 15 to 8 and validated our protocol on a second aliquot from the same specimen, from which we cultured 103 bacterial species, mostly strict anaerobes. This study demonstrates that strict anaerobes do not require extended pre-incubation times. Even if our data are limited by the analysis of one single stool sample, we propose herein a rapid and effective culturomics protocol targeting anaerobes by combining early and late subculture times from one single culture condition, paving the way for high-scale culturomics studies.