Clostridium tetani and Tetanus
Introduction
By Tyler Stearns
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The Tetanus Toxin: Genetics and Mechanism of Action
Genetics
The genome of C. tetani consists of a single 2,799,250-bp chromosome with 2,372 open reading frames (ORFs); the tetanus toxin (TeTx) is encoded on a 74,082-bp plasmid containing 61 ORFs (Brüggemann 2003). The genus Clostridium is a member of the Firmicutes phylum, which are known for a low G-C content (S & F 2011). The C. tetani chromosome has a G-C content of 28.6%, while the TeTx-encoding plasmid pE88 only has a G-C content of 24.5%. The G-C ratio is relatively stable in the main chromosome, indicating a lack of recent horizontal gene transfer (Brüggemann 2003).
The plasmid pE88 holds the genes for TeTx (tetX) and its direct transcriptional regulator TetR. The tetR gene is located just upstream from tetX (Brüggemann 2005). TetR was thought to possibly be a positive regulatory of toxin expression, but current research has shown that TetR is in fact a sigma factor of a subgroup unique to clostridial species. This research found that TetR only associated with target DNA in the presence of the RNA polymerase core enzyme, bound to the core enzyme, and triggered transcription of the target DNA at the promoter in vitro (Raffestin 2005). Other regulatory genes are present on the plasmid. CTP05, CTP10, and CTP11 are sigma factor-like proteins, and CTP21 and CTP22 form a two-component system of unknown regulatory function. The three sigma-like proteins do not appear to be involved in the regulation of toxin production like TetR (Brüggemann 2005; Raffestin 2005).
Other virulence factors may be present on pE88, in addition to the primary tetanus toxin. The 114-kDa collagenase ColT is also encoded on the plasmid (Brüggemann 2003). Collagenase is an enzyme that degrades collagen, which makes up almost 25 to 33% of the total protein in mammalian organisms (Harrington 1996). Thus, ColT may help destroy tissue in an infected host.
The main chromosome of C. tetani also possesses potential virulence factors. ORFs for tetanolysin O, hemolysin, and fibronectin-binding proteins, as well as genes for surface-layer (S-layer) proteins have been identified (Brüggemann 2003). Tetanolysin, discussed below, is one of the two exotoxins produced by C. tetani, in addition to TeTx. The S-layer has been characterized as a molecular sieve or as a possible defense against parasites. However, it also has been implicated in host cell adhesion or as a possible mechanism of evading host immune systems in another pathogenic clostridial species, C. difficile (Spigaglia 2011).
The origin of the plasmid pE88 is still unclear. Over 50% of the ORFs on the plasmid are unique to C. tetani (Brüggemann 2003). Toxin genes are currently—or were in their evolutionary history—part of a flexible clostridial gene pool. Many of the toxin genes in pathogenic Clostridium species are on plasmids or capable of transduction by phages (Brüggemann 2005). C. botulinum, which produces various forms of the botulinum neurotoxin (BoNT), has been shown to transfer its toxin-encoding plasmid by conjugation. In one study, a tagged BoNT-encoding plasmid was transferred from one strain of C. botulinum to another. Bacteriophages for transduction were not observed, and gene transfer was not inhibited by DNase, ruling out transformation of free-floating DNA (Marshall 2010). The epsilon-toxin plasmids in C. perfringens Type D have also been shown to be transferable to other cells by conjugation (Hughes 2007). Given this evidence, and the fact TeTx is quite similar to BoNT, it seems highly probably that the TeTx plasmid was acquired via some form of horizontal gene transfer, though it has clearly undergone its own divergent evolution since that point.
Infection
Tetanus infections most commonly occur after deep-tissue puncture wounds that are exposed to C. tetani. Contamination of the wound often involves contact with soil, fecal matter, or rusty metal that contains C. tetani (Cook 2001; Campbell 2009). C. tetani is an obligate anaerobe. The conditions in a wound are particularly well suited to the bacteria's anaerobic needs (Campbell 2009). Tetanolysin is one of the two exotoxins excreted by C. tetani. Tetanolysin damages tissue surrounding an infection, optimizing conditions within the wound (Cook 2001; Campbell 2009). Bacteria grow and ferment in the wound, releasing in small quantities the actual causative agent of tetanus disease, the tetanospasmin protein (also referred to as tetanus toxin, TeTx, or TeNT). The toxin is mainly released during the stationary phase of growth and a significant majority of the toxin is not freed until a cell lyses, releasing its contents into the body of the host (Mellanby 1981). TeTx is distributed to motor neurons and in the bloodstream to other pre-synaptic nerve terminals in the peripheral nervous system (Kerr 1979).
Cell Entry
Mechanism of Action within the Cell
Clinical Tetanus
Symptoms
Prevention and Treatment
Vaccination
Treating the Infection
Treating the Symptoms
Conclusion
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References
Edited by student of Joan Slonczewski for BIOL 238 Microbiology, 2011, Kenyon College.
