Formula Feeding Predisposes Neonatal Piglets to Clostridium difficile Gut Infection

Abstract

Spontaneous outbreaks of Clostridium difficile infection (CDI) occur in neonatal piglets, but the predisposing factors are largely not known. To study the conditions for C. difficile colonization and CDI development, 48 neonatal piglets were moved into isolators, fed bovine milk-based formula, and infected with C. difficile 078. Analyses included clinical scoring; measurement of the fecal C. difficile burden, toxin B level, and calprotectin level; and postmortem histopathological analysis of colon specimens. Controls were noninfected suckling piglets. Fecal specimens from suckling piglets, formula-fed piglets, and formula-fed, C. difficile-infected piglets were used for metagenomics analysis. High background levels of C. difficile and toxin were detected in formula-fed piglets prior to infection, while suckling piglets carried about 3-fold less C. difficile, and toxin was not detected. Toxin level in C. difficile-challenged animals correlated positively with C. difficile and calprotectin levels. Postmortem signs of CDI were absent in suckling piglets, whereas mesocolonic edema and gas-filled distal small intestines and ceca, cellular damage, and reduced expression of claudins were associated with animals from the challenge trials. Microbiota in formula-fed piglets was enriched with Escherichia, Shigella, Streptococcus, Enterococcus, and Ruminococcus species. Formula-fed piglets were predisposed to C. difficile colonization earlier as compared to suckling piglets. Infection with a hypervirulent C. difficile ribotype did not aggravate the symptoms of infection. Sow-offspring association and consumption of porcine milk during early life may be crucial for the control of C. difficile expansion in piglets.

Porcine Colostrum Protects the IPEC-J2 Cells and Piglet Colon Epithelium against Clostridioides (syn. Clostridium) difficile Toxin-Induced Effects

“…We have recently demonstrated that neonatal piglets fed a formula milk develop gut symptoms similar to that observed in CDI piglets and they also show lower expression of TJ proteins, as compared to suckling piglets [8]. Moreover, neonatal piglets that are switched to formula feeding are known to develop gut microbial dysbiosis, which may predispose them to CDI [8,29]. Our findings as well as previous data clearly support the importance of porcine milk including colostrum in offspring nutrition as a gut barrier for protection against CDI.…”

“…Furthermore, C. difficile toxins have been reported to mediate acute inflammation by induction of microtubule depolymerisation in human colonocytes and induce a pro-inflammatory response in an experimental piglet model [7,28]. We have recently demonstrated that neonatal piglets fed a formula milk develop gut symptoms similar to that observed in CDI piglets and they also show lower expression of TJ proteins, as compared to suckling piglets [8]. Moreover, neonatal piglets that are switched to formula feeding are known to develop gut microbial dysbiosis, which may predispose them to CDI [8,29].…”

“…To obtain the spent culture supernatant with toxins, spores of infectious C. difficile ribotype 078, known to produce toxins A and B and carrying genes encoding a binary toxin, were inoculated into BHI media supplemented with yeast extract and taurocholate and without L-cysteine [8,[15][16][17]. They were then incubated at 37 • C for 1-2 weeks to produce a sufficient concentration of toxins.…”

“…This leads to disruption of tight junction (TJ) proteins and finally to a loss of epithelial integrity, as demonstrated in porcine and human cell lines [4][5][6][7]. Such a scenario of events may lead to the typical signs of C. difficile infection (CDI) [8,9] with severe consequences, such as animal death or growth retardation in piglets that survived the infection [10]. However, why some piglets are asymptomatic carriers of high concentrations of C. difficile spores and toxins and others are not, is not exactly clear [11].…”

“…Porcine milk is rich in bioactive components and has been shown to be beneficial for the offspring [12,13]. Porcine milk has also been demonstrated to strengthen the epithelial barrier function in the piglet gut [14] and previous evidence suggests that it could have the potential to neutralise clostridial toxins [8] and possibly prevent CDI.…”