## 4 Clostridal Binary Toxins  
### C. perfringens  
_C. perfringens_ is an anaerobic, Gram-positive, nonmotile, subterminal, spore-forming rod expressing an extensive arsenal of PFTs across different strains [35]. _C. perfringens_ is a major cause of human foodborne disease and is one of the most common instigators of food poisoning in the United States of America (USA) [35]. Contaminated poultry, meat, and meat-based products serve as the main source of infection for humans, resulting in nearly a million cases of food-related illness in the USA annually [35]. In humans, illness is typically short-term (12-24 h) and includes symptomatology such as watery diarrhea and abdominal pain [35]. Analogous to _C. difficile_, interventions utilizing broad-spectrum antibiotics can lead to _C. perfringens_ infection as a consequence of gut microbiome disruption [35]. Furthermore, depending on the strain of _C. perfringens_, severe symptoms can manifest, including intestinal wound infection, septicemia, enterotoxaemia, and enteritis [35].  
There are a large number of varying PFT toxins produced by _C. perfringens_. In the case of _C. perfringens_ type B and D, the PFT known as epsilon toxin (Ext) causes enterotox anemia in domestic ruminants by entering the blood stream through the intestinal wall and preferentially accumulating in brain and kidney tissue [36]. _C. perfringens_ type C produces _C. perfringens_ beta toxin (CPB), a potent hemolysin-like \(\beta\)-PFT that causes intestinal damage, and lethal necrotic enteritis in both animals and humans through the targeting of endothelial cells [36; 37]. Other \(\beta\)-PFTs produced by _C. perfringens_ strains are specific to livestock, including _C. perfringens_ type F Enterotoxin (CPE) [38]. CPE oligomerizes after binding to cell surface receptors termed claudins, leading to the formation of cytotoxic \(\beta\)-pores that disrupt gastrointestinal homeostasis in mammals [38]. Another hemolysin-like \(\beta\)-PFT produced by _C. perfringens_ type G is necrotic enteritis B-like toxin (NetB), which causes necrotic enteritis in chickens [39; 40]. In addition to these toxins, _C. perfringens_ expresses two binary toxins, the iota toxin, which is expressed by _C. perfringens_ type E, and the binary enterotoxin of _C. perfringens_ (BEC) expressed in _C. perfringens_ strains OS1 and TS1 [41; 42]. For our purposes, this review focuses only on the iota toxin expressed by _C. perfringens_ type E.  
Overfeeding, stress, and cold conditions can trigger _C. perfringens_ type E overgrowth in bovines, promoting the expression of iota toxin [41]. Iota toxin consists of two non-linked subunits: Ia, the catalytic subunit, and Ib, the translocase/pore-forming subunit that forms a heptameric assembly of Ib monomers [14; 41]. Once Ia enters the cytosol of intestinal mucosal cells, it ADP-ribosylates G-actin and prevents the polymerization of F-actin, leading to cytoskeletal destruction, intestinal epithelial cell permeability, and cell death [15; 43; 41]. Fascinatingly, Ia (as well as CDTa and C2I) was shown to have ART activity against Arp2 of the Arp2/3 complex at R179, suggesting these toxins use a multifaceted approach to inhibit actin polymerization [44]. Iota toxin expression was also found to induce antibiotic-associated enterotoxaemia in rabbits and hemorrhagic enterocolitis and sudden death in calves and lambs [43]. This sudden induction of iota toxin expression and the havoc it can wreak on livestock make it an important virulence factor to study with regard to _C. perfringens_ pathogenesis.