title: Tissue-Associated Bacterial Alterations in Rectal Carcinoma Patients Revealed by 16S rRNA Community Profiling.
abstract: Sporadic and inflammatory forms of colorectal cancer (CRC) account for more than 80% of cases. Recent publications have shown mechanistic evidence for the involvement of gut bacteria in the development of both CRC-forms. Whereas, colon and rectal cancer have been routinely studied together as CRC, increasing evidence show these to be distinct diseases. Also, the common use of fecal samples to study microbial communities may reflect disease state but possibly not the tumor microenvironment. We performed this study to evaluate differences in bacterial communities found in tissue samples of 18 rectal-cancer subjects when compared to 18 non-cancer controls. Samples were collected during exploratory colonoscopy (non-cancer group) or during surgery for tumor excision (rectal-cancer group). High throughput 16S rRNA amplicon sequencing of the V4-V5 region was conducted on the Ion PGM platform, reads were filtered usingQiimeand clustered usingUPARSE. We observed significant increases in species richness and diversity in rectal cancer samples, evidenced by the total number of OTUs and the Shannon and Simpson indexes. Enterotyping analysis divided our cohort into two groups, with the majority of rectal cancer samples clustering into one enterotype, characterized by a greater abundance ofBacteroidesandDorea. At the phylum level, rectal-cancer samples had increased abundance of candidate phylumOD1(also known asParcubacteria) whilst non-cancer samples had increased abundance ofPlanctomycetes. At the genera level, rectal-cancer samples had higher abundances ofBacteroides, Phascolarctobacterium, Parabacteroides, Desulfovibrio, andOdoribacterwhereas non-cancer samples had higher abundances ofPseudomonas, Escherichia, Acinetobacter, Lactobacillus, andBacillus. TwoBacteroides fragilisOTUs were more abundant among rectal-cancer patients seen through 16S rRNA amplicon sequencing, whose presence was confirmed by immunohistochemistry and enrichment verified by digital droplet PCR. Our findings point to increased bacterial richness and diversity in rectal cancer, along with several differences in microbial community composition. Our work is the first to present evidence for a possible role of bacteria such asB. fragilisand the phylumParcubacteriain rectal cancer, emphasizing the need to study tissue-associated bacteria and specific regions of the gastrointestinal tract in order to better understand the possible links between the microbiota and rectal cancer.
authors.affiliation: Medical Genomics Laboratory, CIPE/A.C. Camargo Cancer CenterSão Paulo, Brazil; Departamento de Bioquímica, Instituto de Química, Universidade de São PauloSão Paulo, Brazil; Curso de Pós-Graduação em Bioinformática, Universidade de São PauloSão Paulo, Brazil.
authors.forename: Andrew M
authors.initials: AM
authors.lastname: Thomas
authors.affiliation: Medical Genomics Laboratory, CIPE/A.C. Camargo Cancer CenterSão Paulo, Brazil; Department of Pelvic Surgery, A.C. Camargo Cancer CenterSão Paulo, Brazil.
authors.forename: Eliane C
authors.initials: EC
authors.lastname: Jesus
authors.affiliation: Department of Pelvic Surgery, A.C. Camargo Cancer Center São Paulo, Brazil.
authors.forename: Ademar
authors.initials: A
authors.lastname: Lopes
authors.affiliation: Department of Pelvic Surgery, A.C. Camargo Cancer Center São Paulo, Brazil.
authors.forename: Samuel
authors.initials: S
authors.lastname: Aguiar
authors.affiliation: Department of Pathology, A.C. Camargo Cancer Center São Paulo, Brazil.
authors.forename: Maria D
authors.initials: MD
authors.lastname: Begnami
authors.affiliation: Laboratory of Molecular Gynecology, Department of Gynecology, Medicine College, Federal University of São Paulo São Paulo, Brazil.
authors.forename: Rafael M
authors.initials: RM
authors.lastname: Rocha
authors.affiliation: Centro de Oncologia Molecular, Hospital Sirio-Libanês São Paulo, Brazil.
authors.forename: Paola Avelar
authors.initials: PA
authors.lastname: Carpinetti
authors.affiliation: Centro de Oncologia Molecular, Hospital Sirio-Libanês São Paulo, Brazil.
authors.forename: Anamaria A
authors.initials: AA
authors.lastname: Camargo
authors.affiliation: Departamento de Alimentos e Nutrição Experimental, Faculdade de Ciências Farmacêuticas, Food Research Center (FoRC), Universidade de São Paulo São Paulo, Brazil.
authors.forename: Christian
authors.initials: C
authors.lastname: Hoffmann
authors.affiliation: Medical Genomics Laboratory, CIPE/A.C. Camargo Cancer CenterSão Paulo, Brazil; Department of Clinical Oncology, A.C. Camargo Cancer CenterSão Paulo, Brazil.
authors.forename: Helano C
authors.initials: HC
authors.lastname: Freitas
authors.affiliation: Laboratory of Computational Biology and Bioinformatics, A.C. Camargo Cancer Center São Paulo, Brazil.
authors.forename: Israel T
authors.initials: IT
authors.lastname: Silva
authors.affiliation: Medical Genomics Laboratory, CIPE/A.C. Camargo Cancer Center São Paulo, Brazil.
authors.forename: Diana N
authors.initials: DN
authors.lastname: Nunes
authors.affiliation: Departamento de Bioquímica, Instituto de Química, Universidade de São PauloSão Paulo, Brazil; Biocomplexity Institute, Virginia TechBlacksburg, VA, USA.
authors.forename: João C
authors.initials: JC
authors.lastname: Setubal
authors.affiliation: Medical Genomics Laboratory, CIPE/A.C. Camargo Cancer CenterSão Paulo, Brazil; Laboratory of Neurosciences (LIM-27) Alzira Denise Hertzog Silva, Institute of Psychiatry, Faculdade de Medicina, Universidade de São PauloSão Paulo, Brazil.
authors.forename: Emmanuel
authors.initials: E
authors.lastname: Dias-Neto
mesh_terms: D000328:Adult; D000368:Aged; D001419:Bacteria / Q000145:classification* / Q000235:genetics*; D044822:Biodiversity; D001706:Biopsy; D001938:Brazil; D016000:Cluster Analysis; D003106:Colon / Q000382:microbiology / Q000473:pathology; D003113:Colonoscopy / Q000379:methods; D004269:DNA, Bacterial / Q000235:genetics; D004275:DNA, Ribosomal; D005243:Feces / Q000382:microbiology; D005260:Female; D041981:Gastrointestinal Tract / Q000382:microbiology; D005798:Genes, Bacterial; D006801:Humans; D007150:Immunohistochemistry; D008297:Male; D059013:Microbial Consortia / Q000235:genetics*; D008875:Middle Aged; D058889:Molecular Typing; D010802:Phylogeny; D012336:RNA, Ribosomal, 16S / Q000235:genetics*; D012004:Rectal Neoplasms / Q000382:microbiology*; D017422:Sequence Analysis, DNA
chemical_list: D004269:DNA, Bacterial; D004275:DNA, Ribosomal; D012336:RNA, Ribosomal, 16S
keywords: 16S rRNA gene sequencing; Bacterial diversity and community composition; Bacteroides fragilis; mucosa-associated microbiota; rectal cancer