Background: A growing number of Campylobacter species other than C. jejuni and C. coli have been considered as emerging human and animal pathogens. However, the contribution of these species to human gastroenteritis is poorly documented. This study aimed at detecting Campylobacter species from human and cattle faecal samples in Kilosa district, Tanzania using Polymerase Chain Reaction (PCR) amplification of the 16S rRNA gene, and Sanger sequencing . Methods: A total number of 100 faecal samples (70 from human and 30 from cattle) were collected from diarrheic and non-diarrheic patients and healthy cattle in Kilosa district, Tanzania from July to October 2019. Species identification was conducted by PCR and 16S rRNA sequencing. The phylogenetic analysis was carried out by comparison of the 16S rRNA gene sequences to reference strains by the Neighbor-Joining method in MEGA X. Results: Campylobacter species detection rate by PCR was 65.7% (46/70) and 20% (6/30) in humans and cattle, respectively. There were five human diarrheic cases, four showed Campylobacter presence and two were from children ≤15 years of age. In humans, the 16S rRNA sequencing revealed that C. concisus was the most predominant species occurring at a frequency of 37.8% (14/37), followed by uncultured Campylobacter spp. 24.3% (9/37) and C. hominis 21.6% (8/37). The least represented species were C. jejuni and C. lanienae all occurring at 2.7% (1/37). In cattle, five (100%) sequenced PCR products matched with C. lanienae . Phylogenetic analysis revealed that Campylobacter 16S rRNA sequences were closely related to C. concisus , uncultured Campylobacter sp., C. hominis , and C. gracilis . Conclusion: The non- C. jejuni / C. coli species are present in human and cattle faecal samples and their true occurrence is probably under-reported due to shortcomings of conventional techniques used in most diagnostic microbiology laboratories. Based on our findings, we recommend that molecular techniques be adopted for direct detection of Campylobacter species during routine laboratory screening and surveillance studies. Keywords: Campylobacter , molecular diagnostics, polymerase chain reaction, sequencing, gastroenteritis, Tanzania