Phage lysin that specifically eliminates Clostridium botulinum Group I cells

Abstract

Clostridium botulinum poses a serious threat to food safety and public health by producing potent neurotoxin during its vegetative growth and causing life-threatening neuroparalysis, botulism. While high temperature can be utilized to eliminate C. botulinum spores and the neurotoxin, non-thermal elimination of newly germinated C. botulinum cells before onset of toxin production could provide an alternative or additional factor controlling the risk of botulism in some applications. Here we introduce a putative phage lysin that specifically lyses vegetative C. botulinum Group I cells. This lysin, called CBO1751, efficiently kills cells of C. botulinum Group I strains at the concentration of 5 µM, but shows little or no lytic activity against C. botulinum Group II or III or other Firmicutes strains. CBO1751 is active at pH from 6.5 to 10.5. The lytic activity of CBO1751 is tolerant to NaCl (200 mM), but highly susceptible to divalent cations Ca2+ and Mg2+ (50 mM). CBO1751 readily and effectively eliminates C. botulinum during spore germination, an early stage preceding vegetative growth and neurotoxin production. This is the first report of an antimicrobial lysin against C. botulinum, presenting high potential for developing a novel antibotulinal agent for non-thermal applications in food and agricultural industries.

Novel Lytic Enzyme of Prophage Origin from Clostridium botulinum E3 Strain Alaska E43 with Bactericidal Activity against Clostridial Cells

“…Two of them, CD27L and PlyCD, are active against several C. difficile isolates and were characterized in detail [21,22]. Single endolysins targeting Clostridium tyrobutyricum (CTP1L), Clostridium sporogenes (CS74L), and C. botulinum Group I cells (CBO1751) were also reported [23][24][25]. Despite extensive efforts to understand the structure-function relationship of endolysins from Clostridium bacteriophages [26,27], there is still little demonstration of their practical applications.…”

Phage and phage lysins: New era of bio‐preservatives and food safety agents

“…Phage lysins have also been shown great potential against Clostridium botulinum (with more than 10 lysins being reported) and represent a potential nonthermal alternative method to control outbreaks of botulism. More recently, Zhang et al (2020) reported development of a putative recombinantly expressed CBO1751 phage lysin, that exhibited specific lytic activity against cells of C. botulinum Group I over a pH range from 6.5 to 10.5 and tolerant to high NaCl concentration (200 mM). Also, CBO1751 was effective to eliminate C. botulinum during spore germination stage itself , well in advance before active vegetative growth and neurotoxin production.…”

Advances in the use of biocontrol applications in preharvest and postharvest environments: A food safety milestone

“…Several endolysin studies have focused on, among others, identification, efficacy and application in various food systems such as milk (Van Tassell, Ibarra‐Sánchez, Hoepker, & Miller, 2017) and other products. Genetically modified endolysins, PlyP100 (Van Tassell et al, 2017) and CBO1751 (Z. Zhang, Lahti, Douillard, Korkeala, & Lindström, 2020) inhibited L . monocytogenes and Clostridium botulinum , respectively.…”