Dynamic changes of the respiratory microbiota and its relationship to fecal and blood microbiota in healthy young cats

Abstract

Advances in the field of metagenomics using culture-independent methods of microbial identification have allowed characterization of rich and diverse communities of bacteria in the lungs of healthy humans, mice, dogs, sheep and pigs. These data challenge the long held belief that the lungs are sterile and microbial colonization is synonymous with pathology. Studies in humans and animals demonstrate differences in the composition of airway microbiota in health versus disease suggesting respiratory dysbiosis occurs. Using 16S rRNA amplicon sequencing of DNA extracted from rectal and oropharyngeal (OP) swabs, bronchoalveolar lavage fluid (BALF), and blood, our objective was to characterize the fecal, OP, blood, and lower airway microbiota over time in healthy cats. This work in healthy cats, a species in which a respiratory microbiota has not yet been characterized, sets the stage for future studies in feline asthma in which cats serve as a comparative and translational model for humans. Fecal, OP and BALF samples were collected from six healthy research cats at day 0, week 2, and week 10; blood was collected at week 10. DNA was extracted, amplified via PCR, and sequenced using the Illumina MiSeq platform. Representative operational taxonomic units (OTUs) were identified and microbial richness and diversity were assessed. Principal component analysis (PCA) was used to visualize relatedness of samples and PERMANOVA was used to test for significant differences in microbial community composition. Fecal and OP swabs provided abundant DNA yielding a mean±SEM of 65,653±6,145 and 20,6323±4,360 sequences per sample, respectively while BALF and blood samples had lower coverage (1,489±430 and 269±18 sequences per sample, respectively). Oropharyngeal and fecal swabs were significantly richer than BALF (mean number OTUs 93, 88 and 36, respectively; p < 0.001) with no significant difference (p = 0.180) in richness between time points. PCA revealed site-specific microbial communities in the feces, and upper and lower airways. In comparison, blood had an apparent compositional similarity with BALF with regard to a few dominant taxa, but shared more OTUs with feces. Samples clustered more by time than by individual, with OP swabs having subjectively greater variation than other samples. In summary, healthy cats have a rich and distinct lower airway microbiome with dynamic bacterial populations. The microbiome is likely to be altered by factors such as age, environmental influences, and disease states. Further data are necessary to determine how the distinct feline microbiomes from the upper and lower airways, feces and blood are established and evolve. These data are relevant for comparisons between healthy cats and cats with respiratory disease.

Cultural Isolation and Characteristics of the Blood Microbiome of Healthy Individuals

“…We note, that some phyla are enriched by the applied resuscitation strategy according to the OTUs combined abundancy (Table 1). Other studies also identified Proteobacteria as predominant in the blood of healthy individuals [14], healthy broilers [16] and cats [17].…”

Oral Probiotics Alter Healthy Feline Respiratory Microbiota

“…Samples were collected as previously described (Vientos-Plotts et al, 2017). Briefly, cats were anesthetized and a sterile cotton-tip swab was inserted rectally, while avoiding contact with the perianal area.…”

“…However, recent studies using sequencing of microbial 16S rRNA amplicons (Sidiq et al, 2016) have demonstrated that healthy humans (Charlson et al, 2012), dogs (Ericsson et al, 2016), and cats (Vientos-Plotts et al, 2017) harbor rich, complex commensal microbial populations in the respiratory tract. There is increasing evidence that commensal microbes play a critical role in the maturation, education, and function of the immune system and, in particular, regulation of inflammatory responses.…”

Consensus on standard biobanking of gut microbiota

“…Specimens should be directly frozen or added to RNA later stabilizer solution after collection and treatment and then transferred to −80°C for storage. To avoid factors that may influence the results, such as reagents and batches, entire specimens should be collected and then tested together . The order of specimen processing should be randomized.…”