Purification and characterization of Borrelia burgdorferi from feeding nymphal ticks (Ixodes scapularis).
Here we describe a protocol for purifying Borrelia burgdorferi from feeding ticks by velocity centrifugation and Percoll density gradient centrifugation. The purified spirochetes were motile and 10- to 20-fold purer than the bacteria in crude tick homogenates. The purified bacteria were present in sufficient quantity for protein and gene expression studies. In comparison to culture-grown bacteria, tick-borne spirochetes had several proteins that were upregulated and a few that were downregulated. When the levels of B. burgdorferi outer surface proteins OspA and OspC were measured, OspC protein and mRNA levels were lower in cultured bacteria than in bacteria purified from ticks. Although differences in OspA mRNA levels were observed between cultured and tick-borne bacteria, no differences were observed at the protein level. These experiments demonstrate that tick-transmitted borreliae display a gene expression and antigen profile different from that of spirochetes cultured in vitro.
Rathinavelu
S
S
Department of Microbiology and Immunology, University of North Carolina, Chapel Hill 27599, USA.
de Silva
A M
AM
D000818:Animals; D000942:Antigens, Bacterial / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D000954:Antigens, Surface / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D001425:Bacterial Outer Membrane Proteins / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D001428:Bacterial Vaccines; D001431:Bacteriological Techniques; D015748:Borrelia burgdorferi Group / Q000254:growth & development / Q000302:isolation & purification*; D002499:Centrifugation, Density Gradient / Q000379:methods; D003470:Culture Media; D005247:Feeding Behavior; D005260:Female; D018884:Ixodes / Q000382:microbiology* / Q000502:physiology; D008074:Lipoproteins; D008193:Lyme Disease / Q000382:microbiology / Q000635:transmission*; D022123:Lyme Disease Vaccines / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D051379:Mice; D008809:Mice, Inbred C3H
D000942:Antigens, Bacterial; D000954:Antigens, Surface; D001425:Bacterial Outer Membrane Proteins; D001428:Bacterial Vaccines; D003470:Culture Media; D008074:Lipoproteins; D022123:Lyme Disease Vaccines; C072045:OspA protein; C079616:OspC protein