Downloaded from www.microbiologyresearch.org by IP: 54.70.40.11 On: Sun, 04 Aug 2019 09:20:17 Caecibacterium sporoformans gen. nov., sp. nov., an anaerobic, butyrate-producing, spore-forming bacterium isolated from chicken caecum Lonneke Onrust, 1 Diana Petzoldt, 2,3 Venessa Eeckhaut, 1 Celine De Maesschalck, 1 Freddy Haesebrouck, 1 Silke Rautenschlein, 3 Richard Ducatelle, 1 Filip Van Immerseel 1, * and David Taras 2 Abstract Strains of a Gram-stain-negative, rod-shaped and immotile bacterium were isolated from broiler chicken caecal content. The isolates required strict anaerobic conditions for growth, formed spores, were catalase-positive and oxidase-negative. They produced butyrate as the major metabolic end product in reinforced clostridial medium broth. The genomic DNA G+C content of the isolated strains was 32.5–34.6 mol%. The major cellular fatty acids were C 16 : 0 FAME ,C 14 : 0 FAME ,C 19 : 0CYC 9,10DMA and C 16 : 0DMA . The fatty acid composition of the cell wall showed no similarity to any strain in the MIDI database. 16S rRNA gene sequence analysis showed that the nearest phylogenetic neighbours were Anaerostipes hadrus and Clostridium populeti (92 % sequence similarity) within Clostridium cluster XIVa of the phylum Firmicutes. Therefore, a novel genus is proposed, with the name Caecibacterium sporoformans gen. nov., sp. nov. The type strain of Caecibacterium sporoformans is LMG 27730 T =DSM 26959 T . A chicken’s caecum harbours approximately 10 10 to 10 11 bacteria per gram content (wet weight) [1, 2], consisting of up to 640 species [3], of which up to 85 % are still unknown [4, 5]. The majority of the caecal microbiota are strict anae- robes [6, 7]. The abundance of spore-forming anaerobes in chicken faeces is estimated to be 10 4 c.f.u. g 1 [8]. In the past, the genus Clostridium was thought to consist of spore- forming, Gram-stain-positive, anaerobic rod-shaped bacter- ia within the class Clostridia in the phylum Firmicutes. Now the genus includes Gram-stain-negative bacteria, non-spore formers, cocci and non-anaerobes [9]. In 1994, Collins et al. demonstrated the marked phylogenetic incoherence of the genus Clostridium with distinct clusters[10]. Almost one- half of the clostridial species belonged to cluster I and the remaining clostridial species exhibited very considerable degrees of phylogenetic diversity, belonging to numerous clusters [10]. The need for extensive taxonomic revision has been recognized and only Clostridia sensu stricto has been retained in cluster I with Clostridium butyricum as type spe- cies. However, even after extensive reassignment in the past years, many organisms are still misplaced in the genus Clos- tridium, e.g. the Gram-stain-negative spore-formers. Recently, the assignment of all cluster XIVa organisms that are still listed as Clostridium species to the new genus Lach- noclostridium was proposed [9]. However, this proposed genus would still consist of a very heterogeneous group of organisms. The Clostridium cluster XVIa, or the family Lachnospira- ceae, contains numerous butyrate-producing bacteria that colonize the distal intestinal tract of humans and animals including chickens [11]. Butyric acid plays an important role in the reduction of inflammation and pathogen coloni- zation, and increased production of mucins and host anti- microbial peptides [12, 13]. Due to these properties, butyrate producers are interesting probiotic candidates. Sev- eral butyrate-producing strains have been isolated from chicken caecal content [14], however only a minority have been characterized and are spore-forming bacteria [15–17]. Utilization of spores as probiotics is advantageous because of their higher resistance to acidic conditions in the stom- ach, and the requirement of smaller numbers compared to probiotic vegetative cells [18]. In this study spore-forming butyrate-producing strains with 99 % 16S rRNA gene sequence similarity to each other, obtained from chicken Author affiliations: 1 Department of Pathology, Bacteriology and Avian Diseases, Research Group Veterinary Public Health and Zoonoses, Faculty of Veterinary Medicine, Ghent University, Salisburylaan 133, B-9820 Merelbeke, Belgium; 2 Lohmann Animal Health GmbH, Heinz-Lohmann-Straße 4, D- 27472 Cuxhaven, Germany; 3 Clinic for Poultry, University of Veterinary Medicine Hannover, Buenteweg 17, D-30559 Hannover, Germany. *Correspondence: Filip Van Immerseel, filip.vanimmerseel@ugent.be Keywords: chicken; caecum; spore-former; anaerobe; Clostridium cluster XIVa. The GenBank accession numbers for the 16S rRNA sequences of strain LMG 27730 T and strain DSM 100432 (=CECT 8891) are HQ452857 and KX017589 respectively One supplementary table is available with the online Supplementary Material. TAXONOMIC DESCRIPTION Onrust et al., Int J Syst Evol Microbiol 2017;67:4589–4594 DOI 10.1099/ijsem.0.002338 002338 ã 2017 IUMS 4589