b6 mice carrying myd88 and tlr mutations were backcrossed 10 - 12 times to nod / ltj males and intercrossed to produce ko and heterozygous animals . gf animals were re - derived from nod / ltj females impregnated by nod.myd88ko males and kept gf at taconic farms , germantown , ny , the university of chicago and washington university ( st . asf was introduced to gf nod.myd88ko mice by adding cecal contents from donor mice to sterile drinking water . wild - type gf mice were colonized with microbiota from spf nod.myd88ko animals by co - housing gf nod females and newborn progeny with spf myd88.ko females . damage to the islets was scored in a blinded fashion , and graded as follows : 0 no visible infiltration ; i  periinsulitis ; ii - insulitis with < 50% and iii  with > 50% islet infiltration ( supplemental fig . at least 100 islets in each group of 5 to 12 animals were scored . in microbiota transfer experiments 20 sections per pancreas cut at 40m intervals ( 10 islets / section ) were examined and scored ( combining grades ii and iii ) . 610 splenocytes alone , or 210 lymph node cells mixed with 410 irradiated splenocytes from b6.nod-(d17mit21-d17mit10)/ltj ( b6.g7 ) mice , per well of 96-well plates pretreated with anti - ifn- antibodies were incubated overnight with listed peptides . lymphocytes were stained with antibodies against cd4 and cd8 to calculate the frequency of peptide - specific cells per 10 cd4 or cd8 t cells . 16s rrna gene sequences were edited and assembled into consensus sequences using phred and phrap , aided by xplorseq23 , and bases with a phrap quality score of < 20 were trimmed . sequences were aligned using the nast online alignment tool ( http://greengenes.lbl.gov/cgi-bin/nph-index.cgi ) , and checked for chimeras using the online greengenes server ( http://greengenes.lbl.gov/cgi-bin/nph-bel3_interface.cgi ) with a window size of 300 and using the nast - aligned sequences31 . b6 mice carrying myd88 and tlr mutations were backcrossed 10 - 12 times to nod / ltj males and intercrossed to produce ko and heterozygous animals . gf animals were re - derived from nod / ltj females impregnated by nod.myd88ko males and kept gf at taconic farms , germantown , ny , the university of chicago and washington university ( st . asf was introduced to gf nod.myd88ko mice by adding cecal contents from donor mice to sterile drinking water . wild - type gf mice were colonized with microbiota from spf nod.myd88ko animals by co - housing gf nod females and newborn progeny with spf myd88.ko females . damage to the islets was scored in a blinded fashion , and graded as follows : 0 no visible infiltration ; i  periinsulitis ; ii - insulitis with < 50% and iii at least 100 islets in each group of 5 to 12 animals were scored . in microbiota transfer experiments 20 sections per pancreas cut at 40m intervals ( 10 islets / section ) were examined and scored ( combining grades ii and iii ) . 610 splenocytes alone , or 210 lymph node cells mixed with 410 irradiated splenocytes from b6.nod-(d17mit21-d17mit10)/ltj ( b6.g7 ) mice , per well of 96-well plates pretreated with anti - ifn- antibodies were incubated overnight with listed peptides . lymphocytes were stained with antibodies against cd4 and cd8 to calculate the frequency of peptide - specific cells per 10 cd4 or cd8 t cells . 16s rrna gene sequences were edited and assembled into consensus sequences using phred and phrap , aided by xplorseq23 , and bases with a phrap quality score of < 20 were trimmed . sequences were aligned using the nast online alignment tool ( http://greengenes.lbl.gov/cgi-bin/nph-index.cgi ) , and checked for chimeras using the online greengenes server ( http://greengenes.lbl.gov/cgi-bin/nph-bel3_interface.cgi ) with a window size of 300 and using the nast - aligned sequences31 .