Immunizing Compositions and Methods of Use
The present invention provides compositions including siderophore receptor polypeptides and porins from gram negative microbes, and preferably, lipopolysaccarhide at a concentration of no greater than about 10.0 endotoxin units per milliliter. The present invention also provides methods of making and methods of using such compositions.
1 . A composition comprising:
at least two siderophore receptor polypeptides (SRPs) isolated from a gram negative microbe;
at least two porins isolated from the gram negative microbe; and
lipopolysaccharide (LPS) at a concentration of no greater than about 10.0 endotoxin units per milliliter (EU/ml).
2 - 71 . (canceled)
72 . An antibody composition that specifically binds at least one siderophore receptor polypeptide (SRP), the composition prepared by a method comprising:
collecting a body fluid from an animal that comprises antibody that specifically binds to at least one SRP, wherein the animal has been administered an effective amount of a composition comprising at least two siderophore receptor polypeptides (SRPs) isolated from a gram negative microbe.
73 . The antibody composition of claim 72 wherein the composition administered to the animal further comprises lipopolysaccharide (LPS) at a concentration of no greater than about 10.0 EU/ml.
74 . The antibody composition of claim 72 further comprising a pharmaceutically acceptable carrier.
75 . The antibody composition of claim 72 wherein the gram negative microbe is an enteropathogen.
76 . The antibody composition of claim 72 wherein the gram negative microbe is a member of the family Enterobacteriaceae, family Vibrionaceae, family Pasteurellaceae, or family Pseudomonadaceae.
77 . The antibody composition of claim 76 wherein the member of the family Enterobacteriaceae is Shigella spp., Proteus spp., Klebsiella spp., Serratia spp., or Yersinia spp.
78 . The antibody composition of claim 76 wherein the member of the family Vibrionaceae is Vibrio cholerae.
79 . The antibody composition of claim 76 wherein the member of the family Pasteurellaceae is Pasturella spp.
80 . The antibody composition of claim 79 wherein the Pasturella spp. is Pasturella multocida or Pasteurella haemolytica.
81 . The antibody composition of claim 76 wherein the member of the family Pseudomonadaceae is Pseudomonas spp.
82 . The antibody composition of claim 81 wherein the Pseudomonas spp. is Pseudomonas aeruginosa.
83 . The antibody composition of claim 72 wherein the gram negative microbe is a member of the tribe Escherichieae or Salmonelleae.
84 . The antibody composition of claim 83 wherein the gram negative microbe comprises a Salmonella spp., and wherein the composition administered to the animal comprises at least two SRPs having a molecular weight of between about 87 kDa and about 91 kDa, between about 82 kDa and about 86 kDa, or between about 69 kDa and about 75 kDa, wherein the molecular weight is determined by separation by sodium dodecyl sulfate-polyacrylamide gel electrophoresis.
85 . The antibody composition of claim 72 wherein the gram negative microbe is Salmonella spp., Escherichia coli, Actinobacillus spp., Haemophilus spp., or Campylobacter spp.
86 . The antibody composition of claim 72 wherein the composition administered to the animal comprises at least two SRPs having molecular weights of between about 60 kDa and about 100 kDa as determined by separation by sodium dodecyl sulfate-polyacrylamide gel electrophoresis.
87 . The antibody composition of claim 72 wherein the composition administered to the animal further comprises at least one porin.
88 . The antibody composition of claim 87 wherein the at least one porin comprises a porin having molecular weight of about 30 kDa or about 43 kDa as determined by separation by sodium dodecyl sulfate-polyacrylamide gel electrophoresis.
89 . The antibody composition of claim 87 wherein the at least one porin comprises two or more porins.
90 . The antibody composition of claim 72 wherein the composition administered to the animal comprises lipopolysaccharide (LPS) at a concentration no greater than the concentration of LPS in a reference composition comprising at least two SRPs isolated from a gram negative microbe, wherein the reference composition is produced by a process comprising:
disrupting the gram negative microbe in a buffer;
solubilizing the disrupted gram negative microbe for greater than about 24 hours in a solution comprising sarcosine to result in solubilized and insoluble cellular material, wherein a ratio of the sarcosine to gram weight of disrupted gram negative microbe is between about 0.8 gram sarcosine per about 4.5 grams of disrupted gram negative microbe and about 1.2 grams sarcosine per about 4.5 grams of disrupted gram negative microbe; and
isolating molecules of the gram negative microbe, wherein the isolated molecules comprise the at least two SRPs and LPS.
91 . The antibody composition of claim 72 wherein the animal is an avian, a bovine, a caprine, a porcine, an ovine, a bisontine, a cervine, or a companion animal.
92 . The antibody composition of claim 72 comprising antibody that specifically binds to an SRP present on the surface of a second gram negative microbe.
93 . The antibody composition of claim 92 wherein the second gram negative microbe is a member of the family Enterobacteriaceae, family Vibrionaceae, family Pasteurellaceae, or family Pseudomonadaceae.
94 . The antibody composition of claim 72 wherein the body fluid comprises serum, plasma, blood, milk, or colostrum.
95 . The antibody composition of claim 72 wherein the antibody that specifically binds to at least one SRP comprises an antibody fragment.
96 . The antibody composition of claim 72 wherein the antibody fragment comprises an scFv, a Fab, a F(ab′) 2 or a Fv.
97 . An antibody composition that specifically binds at least one siderophore receptor polypeptide (SRP), the composition prepared by a method comprising:
collecting a body fluid from an animal that comprises antibody that specifically binds to at least one SRP, wherein the animal has been administered a genetically modified gram negative whole cell that expresses at least two siderophore receptor polypeptides (SRPs).
98 . The antibody composition of claim 97 wherein the composition administered to the animal further comprises lipopolysaccharide (LPS) at a concentration of no greater than about 10.0 EU/ml.
99 . The antibody composition of claim 97 further comprising a pharmaceutically acceptable carrier.
100 . The antibody composition of claim 97 wherein the genetically modified gram negative whole cell is an enteropathogen.
101 . The antibody composition of claim 97 wherein the genetically modified gram negative whole cell is a member of the family Enterobacteriaceae, family Vibrionaceae, family Pasteurellaceae, or family Pseudomonadaceae.
102 . The antibody composition of claim 101 wherein the member of the family Enterobacteriaceae is Shigella spp., Proteus spp., Klebsiella spp., Serratia spp., or Yersinia spp.
103 . The antibody composition of claim 101 wherein the member of the family Vibrionaceae is Vibrio cholerae.
104 . The antibody composition of claim 101 wherein the member of the family Pasteurellaceae is Pasturella spp.
105 . The antibody composition of claim 104 wherein the Pasturella spp. is Pasturella multocida or Pasteurella haemolytica.
106 . The antibody composition of claim 101 wherein the member of the family Pseudomonadaceae is Pseudomonas spp.
107 . The antibody composition of claim 106 wherein the Pseudomonas spp. is Pseudomonas aeruginosa.
108 . The antibody composition of claim 97 wherein the genetically modified gram negative whole cell is a member of the tribe Escherichieae or Salmonelleae.
109 . The antibody composition of claim 108 wherein the genetically modified gram negative whole cell comprises a Salmonella spp., and wherein the composition administered to the animal comprises at least two SRPs having a molecular weight of between about 87 kDa and about 91 kDa, between about 82 kDa and about 86 kDa, or between about 69 kDa and about 75 kDa, wherein the molecular weight is determined by separation by sodium dodecyl sulfate-polyacrylamide gel electrophoresis.
110 . The antibody composition of claim 97 wherein the genetically modified gram negative whole cell is Salmonella spp., Escherichia coli, Actinobacillus spp., Haemophilus spp., or Campylobacter spp.
111 . The antibody composition of claim 110 wherein genetically modified gram negative whole cell comprises a cell modified to possess lipopolysaccharide (LPS) having decreased outer oligosaccharide side chains compared to a comparable wild-type microbe.
112 . The antibody composition of claim 111 wherein the genetically modified gram negative whole cell comprises E. coli J5.
113 . The antibody composition of claim 111 wherein the genetically modified gram negative whole cell comprises Salmonella minnesota.
114 . The antibody composition of claim 97 wherein the genetically modified gram negative whole cell administered to the animal expresses at least two SRPs having molecular weights of between about 60 kDa and about 100 kDa as determined by separation by sodium dodecyl sulfate-polyacrylamide gel electrophoresis.
115 . The antibody composition of claim 97 wherein the genetically modified gram negative whole cell administered to the animal further expresses at least one porin.
116 . The antibody composition of claim 115 wherein the at least one porin comprises a porin having molecular weight of about 30 kDa or about 43 kDa as determined by separation by sodium dodecyl sulfate-polyacrylamide gel electrophoresis.
117 . The antibody composition of claim 115 wherein the at least one porin comprises two or more porins.
118 . The antibody composition of claim 97 wherein the animal is an avian, a bovine, a caprine, a porcine, an ovine, a bisontine, a cervine, or a companion animal.
119 . The antibody composition of claim 97 comprising antibody that specifically binds to an SRP present on the surface of a second gram negative microbe.
120 . The antibody composition of claim 119 wherein the second gram negative microbe is a member of the family Enterobacteriaceae, family Vibrionaceae, family Pasteurellaceae, or family Pseudomonadaceae.
121 . The antibody composition of claim 97 wherein the body fluid comprises serum, plasma, blood, milk, or colostrum.
122 . The antibody composition of claim 97 wherein the antibody that specifically binds to at least one SRP comprises an antibody fragment.
123 . The antibody composition of claim 97 wherein the antibody fragment comprises an scFv, a Fab, a F(ab′) 2 or a Fv.
124 . An isolated composition comprising:
antibody that specifically binds to at least two siderophore receptor polypeptides (SRPs) from a gram negative microbe.
125 . The antibody composition of claim 124 further comprising a pharmaceutically acceptable carrier.
126 . The antibody composition of claim 124 wherein the gram negative microbe is an enteropathogen.
127 . The antibody composition of claim 124 wherein the gram negative microbe is a member of the family Enterobacteriaceae, family Vibrionaceae, family Pasteurellaceae, or family Pseudomonadaceae.
128 . The antibody composition of claim 127 wherein the member of the family Enterobacteriaceae is Shigella spp., Proteus spp., Klebsiella spp., Serratia spp., or Yersinia spp.
129 . The antibody composition of claim 127 wherein the member of the family Vibrionaceae is Vibrio cholerae.
130 . The antibody composition of claim 127 wherein the member of the family Pasteurellaceae is Pasturella spp.
131 . The antibody composition of claim 130 wherein the Pasturella spp. is Pasturella multocida or Pasteurella haemolytica.
132 . The antibody composition of claim 127 wherein the member of the family Pseudomonadaceae is Pseudomonas spp.
133 . The antibody composition of claim 132 wherein the Pseudomonas spp. is Pseudomonas aeruginosa.
134 . The antibody composition of claim 124 wherein the gram negative microbe is a member of the tribe Escherichieae or Salmonelleae.
135 . The antibody composition of claim 134 wherein the gram negative microbe comprises a Salmonella spp., and wherein the at least two SRPs have molecular weights of between about 87 kDa and about 91 kDa, between about 82 kDa and about 86 kDa, or between about 69 kDa and about 75 kDa, wherein the molecular weight is determined by separation by sodium dodecyl sulfate-polyacrylamide gel electrophoresis.
136 . The antibody composition of claim 124 wherein the gram negative microbe is Salmonella spp., Escherichia coli, Actinobacillus spp., Haemophilus spp., or Campylobacter spp.
137 . The antibody composition of claim 124 wherein the at least two SRPs have molecular weights of between about 60 kDa and about 100 kDa as determined by separation by sodium dodecyl sulfate-polyacrylamide gel electrophoresis.
138 . The antibody composition of claim 124 further comprising antibody that specifically binds to at least one porin.
139 . The antibody composition of claim 138 wherein the at least one porin comprises a porin having molecular weight of about 30 kDa or about 43 kDa as determined by separation by sodium dodecyl sulfate-polyacrylamide gel electrophoresis.
140 . The antibody composition of claim 138 wherein the at least one porin comprises two or more porins.
141 . The antibody composition of claim 124 comprising antibody that specifically binds to an SRP present on the surface of a second gram negative microbe.
142 . The antibody composition of claim 141 wherein the second gram negative microbe is a member of the family Enterobacteriaceae, family Vibrionaceae, family Pasteurellaceae, or family Pseudomonadaceae.
143 . The antibody composition of claim 124 comprising a body fluid from an animal.
144 . The antibody composition of claim 143 wherein the animal is an avian, a bovine, a caprine, a porcine, an ovine, a bisontine, a cervine, or a companion animal.
145 . The antibody composition of claim 143 wherein the body fluid comprises serum, plasma, blood, milk, or colostrum.
146 . A composition comprising:
a body fluid from an animal, or a fraction thereof, comprising antibody that specifically binds to at least two siderophore receptor polypeptides (SRPs) from a gram negative microbe; and
a pharmaceutically acceptable excipient.
147 . The composition of claim 146 wherein the body fluid comprises serum, plasma, blood, milk, or colostrum.
148 . The composition of claim 146 wherein the animal is an avian, a bovine, a caprine, a porcine, an ovine, a bisontine, a cervine, or a companion animal.
149 . The antibody composition of claim 146 wherein the gram negative microbe is an enteropathogen.
150 . The antibody composition of claim 146 wherein the gram negative microbe is a member of the family Enterobacteriaceae, family Vibrionaceae, family Pasteurellaceae, or family Pseudomonadaceae.
151 . The antibody composition of claim 150 wherein the member of the family Enterobacteriaceae is Shigella spp., Proteus spp., Klebsiella spp., Serratia spp., or Yersinia spp.
152 . The antibody composition of claim 150 wherein the member of the family Vibrionaceae is Vibrio cholerae.
153 . The antibody composition of claim 150 wherein the member of the family Pasteurellaceae is Pasturella spp.
154 . The antibody composition of claim 153 wherein the Pasturella spp. is Pasturella multocida or Pasteurella haemolytica.
155 . The antibody composition of claim 150 wherein the member of the family Pseudomonadaceae is Pseudomonas spp.
156 . The antibody composition of claim 155 wherein the Pseudomonas spp. is Pseudomonas aeruginosa.
157 . The antibody composition of claim 146 wherein the gram negative microbe is a member of the tribe Escherichieae or Salmonelleae.
158 . The antibody composition of claim 157 wherein the gram negative microbe comprises a Salmonella spp., and wherein the at least two SRPs have molecular weights of between about 87 kDa and about 91 kDa, between about 82 kDa and about 86 kDa, or between about 69 kDa and about 75 kDa, wherein the molecular weight is determined by separation by sodium dodecyl sulfate-polyacrylamide gel electrophoresis.
159 . The antibody composition of claim 146 wherein the gram negative microbe is Salmonella spp., Escherichia coli, Actinobacillus spp., Haemophilus spp., or Campylobacter spp.
160 . The antibody composition of claim 146 wherein the at least two SRPs have molecular weights of between about 60 kDa and about 100 kDa as determined by separation by sodium dodecyl sulfate-polyacrylamide gel electrophoresis.
161 . The antibody composition of claim 146 further comprising antibody that specifically binds to at least one porin.
162 . The antibody composition of claim 161 wherein the at least one porin comprises a porin having molecular weight of about 30 kDa or about 43 kDa as determined by separation by sodium dodecyl sulfate-polyacrylamide gel electrophoresis.
163 . The antibody composition of claim 161 wherein the at least one porin comprises two or more porins.
164 . The antibody composition of claim 146 comprising antibody that specifically binds to an SRP present on the surface of a second gram negative microbe.
165 . The antibody composition of claim 164 wherein the second gram negative microbe is a member of the family Enterobacteriaceae, family Vibrionaceae, family Pasteurellaceae, or family Pseudomonadaceae.