Clostridium perfringens group infections

General introduction

Clostridium perfringens, previously referred to as Clostridium welchii, was first described in 1891 by Achalme as the Bacillus of Acute Articular Rheumatism.¹⁰³ In the historical literature it was also known by names such as Bacillus phlegmonis emphysematosae, Bacillus emphysematis vaginae, Bacillus cadaveris butyricus, Bacillus aerogenes capsulatus, Granulobacillus saccharobutyricus liquefaciens immobilis, and as Bacillus perfringens.^{103, 106}

Clostridium perfringens is a typical representative of the genus Clostridium, which requires anaerobic growth conditions. It is widely distributed in nature and occurs in soil, sewage and water, as well as in the intestinal tract of humans and warm- and some cold-blooded animals. Clostridium perfringens is usually absent from the stomach, but present in variable numbers (vegetative bacteria and spores) in the small and large intestines of most animals. Vegetative organisms may multiply in soil¹⁰⁵ and counts as high as 5 × 10⁴ C. perfringens organisms per gram of soil have been recorded.^{105, 111} Normal human intestinal contents can contain as many as 10⁹ organisms per gram,¹ the number being dependent on the diet of the host.² In pigs fed on a high protein diet, the number of C. perfringens is significantly higher than in pigs on a standard protein diet.⁵⁰ Higher faecal counts of C. perfringens are found in calves aged between one and ten days than in older animals.⁴

The different Clostridium perfringens types produce six major toxins: alpha (CPA), beta (CPB), epsilon (ETX),  iota (ITX), enterotoxin (CPE), and necrotic enteritis B-like (NetB), which are used for typing them into toxin types A, B, C, D, E, F and G (Table 1).⁷⁷ Each one of these types also produces a number of so-called minor, less lethal toxins. Under certain in vivo and in vitro conditions the major and minor toxins are produced alone or in combination, depending on the specific type, and are responsible for the pathogenicity of the bacterium.^{77, 116, 117} These toxins, alone or in combination, are responsible for the various syndromes associated with this group of organisms. They cause disease as a consequence of their local effects on the intestinal tract, which may vary from insignificant to outspoken, and/or the systemic effect of absorbed toxins — a situation referred to as enterotoxaemia.

The effects of these toxins, also referred to as virulence factors, are divided into three groups, namely:¹⁰⁴

• the alpha and kappa toxins, which are phospholipase C (lecithinase) and collagenase enzymes, respectively, hydrolyze substances essential to the integrity of cellular membranes or other body structures;
• a group that includes the beta, epsilon, iota, enterotoxin and NetB toxins, that are pore forming toxins
• a group that includes the haemolytic toxins such as the theta and delta toxins.

Clostridium perfringens is associated with a wide variety of diseases (Table 2) that affect most domestic animal species and humans and include enterotoxaemia, haemorrhagic and necrotic enteritis, and gas gangrene.^{53, 62, 64, 106, 110, 122} It is one of the most widespread and potential pathogenic organisms in nature. C. perfringens type F, producing enterotoxin is an important cause of food poisoning in humans.^{53, 102, 103}

Clostridium perfringens grows well on rabbit, human, ovine, bovine and equine blood agar at 35 to 37 °C in an anaerobic atmosphere. After 18 to 24 hours’ incubation, the colonies are 2 to 5mm in diameter, white or grey and translucent, with a glossy appearance, low convex, and round with edges that may vary from irregular/serrated in some isolates to entire in most isolates. Pure cultures of C. perfringens frequently contain two or more colony forms, with aberrant forms sometimes occurring alongside the typical colony forms. On ovine, bovine, rabbit, and human blood agar, colonies are surrounded by a narrow zone of complete haemolysis caused by the theta toxin, surrounded by a wider zone of partial haemolysis caused by the alpha toxin (Figure 1). On horse blood agar, only the theta toxin is responsible for haemolysis. The alpha toxin has relatively little effect on horse and goat red blood cells.^{53, 80, 103} Clostridium perfringens tolerates a wide temperature range when incubated and, although the optimal range is 35 to 37 °C, temperatures of 43 to 45 °C, are tolerated. When grown under optimal conditions the generation time (i.e. the time it takes an organism to double itself) of C. perfringens may be as short as eight minutes, which makes it probably the most rapidly growing organism known.¹⁰⁶

Different media have been composed for the selective isolation of C. perfringens in the presence of other organisms. The addition of antibiotics, such as neomycin, to the tryptonesulphite- neomycin medium of Marshall, Steenbergen and McClung,⁵¹ allows...