Live attenuated bacterial vaccine to reduce or inhibit carriage and shedding of enterohemorrhagic Escherichia coli in cattle
Inventors
Zhu, Chengru • Boedeker, Edgar
Assignees
US Department of Veterans Affairs • UNM Rainforest Innovations
Publication Number
US-8858930-B2
Publication Date
2014-10-14
Expiration Date
2025-10-26
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Abstract
The invention provides live, attenuated enterohemorrhagic Escherichia coli (EHEC) bacteria in which the Shiga toxin coding sequences are deleted to abolish Shiga toxin production and one or more of the nucleotide sequences for the bacterial adhesin protein intimin, the locus of enterocyte effacement encoded regulator, and the translocated intimin receptor are mutated to inactivate the activity of the encoded protein(s). This live, attenuated E. coli bacteria is used in a vaccine for reducing or inhibiting carriage and shedding of EHEC in cattle.
Core Innovation
The invention provides live, attenuated enterohemorrhagic Escherichia coli (EHEC) bacteria in which the Shiga toxin coding sequences are deleted to abolish Shiga toxin production and one or more of the nucleotide sequences for the bacterial adhesin protein intimin, the locus of enterocyte effacement encoded regulator (ler), and the translocated intimin receptor (tir) are mutated to inactivate the activity of the encoded protein(s). This live, attenuated E. coli bacteria is used in a vaccine for reducing or inhibiting carriage and shedding of EHEC in cattle.
The problem being solved is that Shiga toxin-producing strains of Escherichia coli (STEC), also known as enterohemorrhagic Escherichia coli (EHEC), are important foodborne pathogens associated with epidemics of bloody diarrhea, hemorrhagic colitis, and serious complications including fatal hemolytic uremic syndrome (HUS) in humans. Cattle are frequently identified as the primary source of EHEC infection due to colonization and shedding that causes contamination of livestock-derived food products. There are no proven vaccines or therapeutic agents for STEC (EHEC) infections, and antibiotic therapy may worsen outcomes. Controlling EHEC carriage and shedding in cattle is thus critical to reduce the incidence of human infections.
The invention overcomes the limitations of no effective human or animal vaccines by providing a live attenuated EHEC vaccine targeting cattle asymptomatically colonized by EHEC strains. The vaccine induces mucosal immunity to prevent or inhibit EHEC colonization and fecal shedding. Live attenuated EHEC mutants are constructed by deleting the Shiga toxin coding sequences to abolish toxin production and mutating eae (intimin), ler, or tir genes to inactivate bacterial adherence and virulence functions, thereby attenuating bacterial virulence while maintaining immunogenicity. The mutants retain colonization persistence facilitating the development of local immunity. Vaccination of cattle with the live attenuated strains is expected to reduce or inhibit EHEC colonization, thus providing safer meat and a cleaner environment.
Claims Coverage
There is one independent claim defining an isolated, live attenuated, double mutant of EHEC strains with specific genetic modifications.
Live attenuated double mutant EHEC with deleted Shiga toxin gene and truncated intimin
The invention covers an isolated, live attenuated, double mutant of enterohemorrhagic Escherichia coli (EHEC) O157:H7, O26, or O111 in which the Shiga toxin coding sequence is deleted to abolish its production and the nucleotide sequence coding for intimin (Eae) is mutated by deletion of the C-terminal Tir-binding domain. This mutation eliminates intimin's Tir-binding activity while retaining the rest of the intimin protein that comprises the immunodominant regions.
Live attenuated mutant containing no antibiotic resistance marker
The double mutant as above is free of any antibiotic resistance marker.
Live attenuated mutant including pharmaceutically acceptable carriers and adjuvants
The inventions include the live attenuated double mutant formulated with a pharmaceutically acceptable carrier, with or without added adjuvants.
The claim covers live attenuated EHEC mutants with deleted Shiga toxin genes and targeted mutations in intimin that abolish adherence, making safe, immunogenic vaccine strains for cattle. These mutants lack antibiotic resistance markers and can be formulated with suitable carriers and adjuvants.
Stated Advantages
Live attenuated EHEC vaccines induce effective mucosal immunity preventing or inhibiting EHEC colonization and shedding in the bovine gut.
Truncation mutation of intimin abolishes bacterial virulence while retaining immunogenicity to generate protective antibody responses.
Deletion of Shiga toxin genes abolishes toxin production, making vaccine strains safe for use in cattle.
Persistence of mutant bacteria in the intestinal tract promotes local immunity development.
Vaccination of cattle reduces EHEC shedding, leading to safer meat and decreased human infection risk.
Documented Applications
Use as a vaccine for reducing or inhibiting carriage and shedding of enterohemorrhagic Escherichia coli in cattle.
Immunization of cattle with live attenuated EHEC containing deletion mutations in Shiga toxin and mutations in intimin, ler or tir genes.
Oral or intrarectal administration of live attenuated EHEC vaccines to cattle to induce mucosal immune responses.
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