Gut microbiota features associated with Clostridioides difficile colonization in puppies

Abstract

In people, colonization with Clostridioides difficile , the leading cause of antibiotic-associated diarrhea, has been shown to be associated with distinct gut microbial features, including reduced bacterial community diversity and depletion of key taxa. In dogs, the gut microbiota features that define C . difficile colonization are less well understood. We sought to define the gut microbiota features associated with C . difficile colonization in puppies, a population where the prevalence of C . difficile has been shown to be elevated, and to define the effect of puppy age and litter upon these features and C . difficile risk. We collected fecal samples from weaned (n = 27) and unweaned (n = 74) puppies from 13 litters and analyzed the effects of colonization status, age and litter on microbial diversity using linear mixed effects models. Colonization with C . difficile was significantly associated with younger age, and colonized puppies had significantly decreased bacterial community diversity and differentially abundant taxa compared to non-colonized puppies, even when adjusting for age. C . difficile colonization remained associated with decreased bacterial community diversity, but the association did not reach statistical significance in a mixed effects model incorporating litter as a random effect. Even though litter explained a greater proportion (67%) of the variability in microbial diversity than colonization status, we nevertheless observed heterogeneity in gut microbial community diversity and colonization status within more than half of the litters, suggesting that the gut microbiota contributes to colonization resistance against C . difficile . The colonization of puppies with C . difficile has important implications for the potential zoonotic transfer of this organism to people. The identified associations point to mechanisms by which C . difficile colonization may be reduced.

Gut microbiota features associated with Clostridioides difficile colonization in dairy calves

“…Clostridioides difficile is a spore-forming anaerobic, gram-positive bacillus that is a significant enteric pathogen in many species of animals. Colonization with C. difficile has been shown to be associated with reduced gut microbial diversity and increased colonization of pathogenic bacteria in people (7, 8), and we recently demonstrated a similar association in puppies (9). Dairy calves, like the neonates of other species, are colonized with C. difficile at high rates, with reported prevalences ranging from 28-56% (10, 11).…”

“…While the role of C. difficile in calf diarrhea remains equivocal (12), exploring the association between this pathogen and the gut microbiome is important for understanding factors that affect gut health and enteric diseases. While a number of studies have examined the epidemiology of C. difficile in animals of veterinary importance, the association between the microbiome and C. difficile is only beginning to be explored in dogs (9), horses (29), and pigs (30). Notably, in pigs, the presence of C. difficile is associated with significantly reduced microbial diversity and increased levels of enteropathogens associated with neonatal diarrhea (30).…”

“…The differential levels in calves compared to people with enteric disease may be due to species-specific patterns of development of the neonatal gut. Species-specific differences may also explain why C. difficile colonized calves had higher levels of Clostridial genuses but colonized puppies had lower levels (9). While rumen development is minimal in pre-weaned calves, they are nevertheless ruminants and thus have fundamentally different enteric physiologies and microbial ecologies compared to true monogastric species.…”

“…DNA was extracted from fecal samples using Qiagen PowerSoil DNA extraction kit. 16S rRNA sequencing was performed as described previously (9, 15). Briefly, the V4 region of the 16S rRNA gene was amplified using PCR, which was performed using Accuprime Pfx Supermix and custom primers for 30 cycles (15).…”

Gut microbiota features associated with Clostridioides difficile colonization in dairy calves

“…Colonization with C . difficile has been shown to be associated with reduced gut microbial diversity and increased colonization of pathogenic bacteria in people [ 7 , 8 ], and we recently demonstrated a similar association in puppies [ 9 ]. Dairy calves, like the neonates of other species, are colonized with C .…”