Recipient Organization
UNIV OF PENNSYLVANIA
(N/A)
PHILADELPHIA,PA 19104
Performing Department
CLINICAL STUDIES
Non Technical Summary
Calf diarrhea is a major cause of illness and death in dairy calves, and it represents a major economic loss for the dairy farmer. Many factors can contribute to the development of calf diarrhea, including management practices on the farm and animal nutrition, but ultimately this disease is caused by the flourishing of pathogenic bacteria in the animal's gut that overwhelm the "good bacteria" that keep the intestinal tract healthy. Studies have shown that the bacterial make-up of the calf gut (the "microbiome") is critical to calf health.One particular microbe that can be found in young calves is called Clostridioides difficile, or "C. diff". In people and in certain species of animals, C. diff can cause debilitating and sometimes life-threatening diarrhea. One of the ways it does this is by outcompeting the native flora of the gut and producing toxins that damage the lining of the intestine. It is unclear whether C. diff can cause diarrhea in calves. However, the presence of C diff alone, even at levels insufficient to cause diarrhea, has been shown to be associated with an altered gut microbiome in people and in several species of animals. We recently demonstrated such a finding in puppies, and we now intend to explore whether similar associations between C. diff and the gut microbiome occur in dairy calves. We propose to compare the gut microbiome of dairy calves that carry C. diff and calves that do not. Having a better understanding of the factors that contribute to a healthy gut can eventually lead to better control of calf diarrhea and the prevention of illness and death in calves and economic loss for the farmer.
Animal Health Component
0%
Research Effort Categories
Basic
100%
Applied
0%
Developmental
0%
Goals / Objectives
Infectious diarrheal disease is one of the main causes of mortality in dairy calves (1, 2), and calves less than 30 days of age are at highest risk of developing diarrhea (3, 4). Studies have shown that gut microbial composition is associated with gut health and the likelihood of diarrhea: reductions in microbial diversity are associated with an increased incidence of diarrhea (5), and the colonization of the calf gut with beneficial bacteria along with the decreased colonization of potential pathogens decreases the likelihood of calf diarrhea (6). 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 there is some evidence that infection with C. difficile can result in diarrhea in calves (12), the effect of the asymptomatic colonization of calves on the gut microbiome is unknown. Given the crucial role of the gut microbiome in providing colonization resistance against pathogens that cause diarrhea (13, 14), a better understanding of the effect of pathogens such as C. difficile on the calf gut microbiome is needed. The goal of this study is to define the gut microbiota features associated with C. difficile colonization in dairy calves and to define the effects of calf age, diet, and farm on the risk of colonization.
Project Methods
Methods:We are currently in possession of a collection of 109 fecal samples from calves less than 14 days of age from 20 dairy farms in Pennsylvania. Pooled samples from each farm have undergone anaerobic culture for C. difficile as previously described (9). We propose to individually culture samples from the pooled samples that previously tested positive (n=36) and to submit all fecal samples (n=109) for 16S sequencing. Briefly, DNA will be extracted from the fecal samples, and the V4 region of the 16S rRNA gene will be amplified using barcoded primers for use on the Illumina platform (15). Sequencing will be performed using 250-base paired-end chemistry on an Illumina MiSeq instrument at the Center for Host Microbial Interactions at the University of Pennsylvania School of Veterinary Medicine.The effects of calf age, farm, and diet on culture status will be analyzed by logistic regression. Metrics of alpha and beta diversity of the fecal microbiota will be calculated using the qiime diversity core-metrics-phylogenetic function in qiime2 and visualized using QIIME2 and Emperor (16). The relative contributions of different microbial taxa that characterize the differences between C. difficile culture positive and negative calves will be assessed through linear discriminant analysis effect size (LEfSe).