Different storage conditions affect microbial profiles in samples of chicken caecal chyme and lung

This study has evaluated the effect of storage temperature on microbial composition to provide a suitable preparation method for clinical samples. In this study, fresh chicken pulmonary and caecal samples were harvested and set as the control, after which they were stored at different temperatures: −20℃, −80℃, or −196℃ for 7 days. The total ribosomal deoxyribonucleic acid content of the samples was extracted and sequenced to compare differences in microbial composition. At the phylum level, the abundances of Campilobacteria stored at −80°C and −196℃ were lower than in the control and at −20℃. The richness of Proteobacteria in the samples at −20℃ and −80℃ was increased than those in the control. At the genus level, Bacteroides, Rikenellaceas_RC9, Clostrida_vadinBB60, norank_f_norank _o_rhodospirllales, and norank_ f_ Barnesiellaceae accounted for the largest proportion. The abundances of Bacteroides, Helicobacter and Clostridia_vadinBB60 were reduced at −20℃ and −80℃ compared with the control. For the caecal samples, the number of OTUs in control was higher than those stored at −196℃, whereas no difference was observed between the fresh samples and those stored at −20℃ and −80℃. At the phylum level, the abundance of Proteobacteria and unclassified_k_norank_d_bacteria increased in the treated samples compared to the control, whereas the diversity of Firmicute and Bacteroidota was reduced compared with that in the control. Especially, the Cyanobacteria, Campilobacterota, and Synergistota phyla at −80℃ and −196℃ were further reduced than those of the control and at −20℃. At the genus level, Bacteroides, Lactobacillus, Megasphaera, norank_f_ruminococcaceae, Helicobacter, and norank_f_ norank_o_Gastranaerophilales were more abundant in the control samples than in the treated samples. However, the richness of unclassified_k_norank_d_bacteria, Sphingomonas and norank_f_ norank_o_SJA－15 were higher in all of the treated samples than in the control. The pulmonary and caecal samples stored at −80℃ and −196℃ had reduced the bacterial diversities compared with those in the control and at −20℃ as suggested by the linear discriminant analysis. In conclusion, samples of the lung and caecal chyme stored at −20℃ for 7 days can promote the colonisation of microbiota, leading to inaccurate sequencing data. Samples stored at −80℃ and −196℃can decrease the abundance of the microbial composition.