Comparison of microbiome and culture techniques for determination of gastrointestinal microbial communities in ceca of chickens

Abstract

The use of 16S next generation sequencing (NGS) technology to identify the relative abundance of microbial communities have become the standard when studying the intestinal microbiome. The increased use is due to the ability to identify a proportion of bacteria that cannot be observed with culture-based methods. However, culture-based techniques are acceptable to identify key bacterial groups, yet may grossly underestimate the microbial community in question. Since there is limited research comparing NGS results to colony forming units (CFU), the objective of this study was to compare total Enterobacteriaceae and lactic acid bacteria (LAB) recovery with culture techniques (CFU/g ceca) to total number of reads from operational taxonomic units (OTU) categorized as Enterobacteriaceae or LAB from Illumina MiSeq platform from matched chick cecal samples at three and 10 days of age. Both CFU recovery (1.09x109 {+/-} 2.42x108; 1.37x108 {+/-} 5.57x107) and reads (5460 {+/-} 1164; 282 {+/-} 163) belonging to Enterobacteriaceae decreased by 10 days of age (p < 0.001). Similarly, LAB reads decreased over time (21,128 {+/-} 2262; 6220 {+/-} 817, respectively p < 0.0001). However, LAB CFU recovery increased by 10 days (1.18x108 {+/-} 1.91x107; 1.62x109 {+/-} 5.00x108, respectively p < 0.01). At three days the Pearsons correlation was -0.082 between CFU of culturable Enterobacteriaceae to reads and culturable LAB CFU to reads at 0.097, showing no correlation (p = 0.606, 0.551; respectively). By 10 days, no correlation of reads and CFU occurred with Enterobacteriaceae (r=-0.049; p-value = 0.769) while with LAB the correlation was 0.290 (p = 0.066) at 10 days. The CFU may be appropriate to identify a few families that change due to treatment or product. Without identifying viable cells to DNA recovered from NGS, there will always be the question whether the reads within the binned OTU in the intestinal tract is accurate.