Therapeutic potential of egg yolk antibodies for treating Clostridium difficile infection

Abstract

Herein we present evidence for the therapeutic potential of colonization factor (CF)-specific egg yolk antibodies (IgY) for potentially treating acute and recurring Clostridium difficile infection (CDI) in humans. The study involved cloning, expressing as 6¾His-tagged proteins in Escherichia coli, and Ni-affinity purifying three previously identified CFs (FliC, FliD and Cwp84) from C. difficile. The recombinant CF antigens were then used to immunize Leghorn chickens and CF-specific IgY antibodies were prepared from their eggs. The specificity and titre of the resulting C. difficile CF-specific IgY antibodies were assessed by ELISA and Western immunoblotting techniques. The antibodies were also screened for their ability to inhibit C. difficile adherence to human colon-derived T84 cells, and, based on these findings, one of them (FliD-specific IgY) was evaluated for its potential to prevent C. difficile-mediated morbidity and mortality in Syrian hamsters. The results revealed that purified FliD-specific IgY significantly protected hamsters from C. difficile strain 630 infection relative to control animals treated with carbonate buffer alone or IgY produced from unimmunized chicken eggs. The results suggest that egg yolk preparations obtained from chickens immunized with recombinant C. difficile CFs may represent another safe and cost-effective treatment option in humans suffering from acute or recurring CDI. INTRODUCTIONConventional antibiotics alter both the numbers and complexity of the normal gastrointestinal (GI) microflora, thereby reducing the ability of these protective organisms to competitively exclude opportunists from colonizing the GI tract. This disruption may render hospitalized subjects susceptible to colonization by C. difficile (Job & Jacobs, 1997). Once established in the colon, C. difficile expresses two very large exotoxins, toxin A (TcdA) and toxin B (TcdB), which are primarily responsible for the clinical symptoms of C. difficile infection (CDI) (Voth & Ballard, 2005). These range from diarrhoea to pseudomembranous colitis, toxic megacolon and multi-system organ failure, which may be fatal.The accepted approach to treating CDI is to replace the antibiotic responsible for initiating the condition with one, typically metronidazole, to which C. difficile is sensitive. In about 80 % of cases, this approach leads to complete recovery without further complications. In the remaining cases, however, recurring CDI (rCDI) can occur, due, presumably, to a failure of the normal GI microflora to become re-established after the metronidazole is discontinued. This leaves the colon of these individuals susceptible to C. difficile relapse or reinfection. Multiple recurrences are not unusual, resulting in a significant reduction in the quality of life in these individuals.In response to the increasing risk of CDI to human health, much research has focused on new and improved treatment regimens, particularly in subjects suffering from rCDI. These studies have concentrated on novel toxin neutralizing agents, an...

Neutralization of Clostridium difficile Toxin B Mediated by Engineered Lactobacilli That Produce Single-Domain Antibodies

“…These results confirm the potential of using VHH antibodies as an affordable antibody-based approach for the treatment of CDI. Although systemic administration of monoclonal antibodies was previously found to be protective against a C. difficile challenge (42,47), very few studies have reported protection for oral delivery of antibodies, showing the difficulty of this approach (44,48,49). Using a similar experimental setup, Kink and Williams (44) observed that hamsters fed daily for 4 consecutive days with a high dose of antitoxin chicken immunoglobulin were protected over a period of 20 days following spore challenge.…”

The potency of chicken egg yolk immunoglobulin (IgY) specific as immunotherapy to Mycobacterium tuberculosis infection

“…The deposition of IgY into the egg yolks of the immunized bird then provides an elegant source of polyclonal immunoglobulins. [10] Since polyclonal IgY can be recovered from the eggs of laying hens for prolonged periods, this approach provides a long-term supply of substantial amounts of antibodies.…”

“…[9] Chicken antibodies also recognize other epitopes than mammalian antibodies. [10] Thus, the eggs of immunized chickens are an economical and abundant source of polyclonal antibody. Immunoglobulin therapy may diminish the development of antibiotic resistant microorganisms.…”

The Prospect for Vaccines to Prevent Clostridium difficile Infection

“…54 Avian immunoglobulins in the form of egg IgY antibodies against recombinant peptides spanning the TcdA and TcdB when given orogastrically to hamsters provided complete protection from CDI and death. 55 Ovine hyperimmune serum against different regions of the RBDs of the toxins has helped narrow down the essential regions needed for a toxin-neutralizing immune response to protect against disease and death. 56 Fully human monoclonal antibodies against the RBD of TcdA and TcdB have been developed by many groups and a combination of these parenterally given monoclonal antibodies were shown to protect antibiotic-treated mice or hamsters from C difficile challenge.…”