The present invention relates to modified Mannheimia haemolytica ( M. haemolytica ) lktCA gene cluster cassettes, compositions comprising such cassettes, methods of using such cassettes and compositions, and kits comprising such cassettes and compositions. Also described herein are Mycoplasma bovis ( M. bovis ) protective antigens, compositions comprising such antigens, methods of using such antigens and compositions, and kits comprising such antigens and compositions. Also described herein are modified M. haemolytica lktCA gene cluster cassettes engineered to express M. bovis protective antigens, compositions comprising such cassettes, methods of using such cassettes and compositions, and kits comprising such cassettes and compositions.
1. A cassette comprising a modified Mannheimia haemolytica lktCA gene cluster, wherein the modified M. haemolytica lktCA gene cluster comprises two polynucleotides encoding leukotoxin neutralizing epitopes, a first polynucleotide encoding the native leukotoxin neutralizing epitope, and a second polynucleotide encoding an additional leukotoxin neutralizing epitope inserted immediately downstream of the native leukotoxin A start codon.
2. A composition comprising the cassette of claim 1 , wherein the composition is a polynucleotide, a plasmid, an expression vector, a host cell, a vaccine, or an immunogenic composition.
3. The composition of claim 2 , wherein the composition is a replacement plasmid, an attenuated M. haemolytica strain A1 bacterium, or an attenuated M. haemolytica strain A6 bacterium.
4. The cassette of claim 1 , further comprising at least one polynucleotide encoding a heterologous antigen.
5. The cassette of claim 4 , wherein the at least one polynucleotide encoding a heterologous antigen is inserted upstream of the polynucleotide encoding the added leukotoxin neutralizing epitope.
6. A composition comprising the cassette of claim 4 , wherein the composition is a polynucleotide, a plasmid, an expression vector, a host cell, a vaccine, or an immunogenic composition.
7. The cassette of claim 4 , wherein the at least one polynucleotide encoding a heterologous antigen is at least one M. bovis antigen.
8. A kit comprising the cassette of claim 1 .
9. A kit comprising the cassette of claim 4 .
10. A kit comprising the composition of claim 6 .