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22508862[PMID] - PMC - NCBI

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1.
Fig 3

Fig 3. From: Borrelia burgdorferi Needs Chemotaxis To Establish Infection in Mammals and To Accomplish Its Enzootic Cycle.

Detection of plasmid contents in the cheA2mut (A) and cheA2com (B) strains by PCR. The primers for PCR were described previously ().

Ching Wooen Sze, et al. Infect Immun. 2012 Jul;80(7):2485-2492.
2.
Fig 2

Fig 2. From: Borrelia burgdorferi Needs Chemotaxis To Establish Infection in Mammals and To Accomplish Its Enzootic Cycle.

Detection of CheA2 in the cheA2mut strain and the complemented cheA2com strain. (A) Construction of CheA2/pBBE22G for complementation of cheA2mut. The fused PflgB-cheA2 fragment was released from pFlgBA2com, a previously constructed vector for complementing an avirulent cheA2 mutant (), and cloned into pBBE22G (), a shuttle vector of B. burgdorferi, yielding CheA2/pBBE22G. (B) Western blot analysis of cheA2mut and cheA2com strains. The same amounts of wild-type, cheA2mut, and cheA2com whole-cell lysates were analyzed by SDS-PAGE and then probed with CheA2 and DnaK antibodies as previously documented (, ).

Ching Wooen Sze, et al. Infect Immun. 2012 Jul;80(7):2485-2492.
3.
Fig 7

Fig 7. From: Borrelia burgdorferi Needs Chemotaxis To Establish Infection in Mammals and To Accomplish Its Enzootic Cycle.

Detection of spirochete burdens in C3H mice infected via tick bite, using qRT-PCR. At day 14 after tick feeding, mice were sacrificed and tissues (skin, heart, joint, and bladder) collected for RNA isolations and then subjected to qRT-PCR analysis as described in the legend to . The data shown here are the results for skin and heart tissues. No trace of flaB message was detected in the mouse tissues infected by cheA2mut; since its PCR cycle thresholds (CT) are higher than that of the negative controls (no transcriptase added), it is considered negative.

Ching Wooen Sze, et al. Infect Immun. 2012 Jul;80(7):2485-2492.
4.
Fig 1

Fig 1. From: Borrelia burgdorferi Needs Chemotaxis To Establish Infection in Mammals and To Accomplish Its Enzootic Cycle.

Detection of CheA2 under various culture conditions. The wild-type A3-68 Δbbe02 or B31A3 (, ) strain was cultivated under various conditions: UF conditions (23°C and pH 7.6), 34°C and pH 7.6, 37°C and pH 6.8, or FT conditions (switch from 23°C and pH 7.6 to 37°C and pH 6.8) (A) or in the presence of DMC (B) (). Similar amounts of whole-cell lysates were analyzed by SDS-PAGE and then probed with CheA2, DnaK (a loading control), and OspC antibodies as previously described (). Increased OspC expression was used as a marker for cells cultured in DMC.

Ching Wooen Sze, et al. Infect Immun. 2012 Jul;80(7):2485-2492.
5.
Fig 6

Fig 6. From: Borrelia burgdorferi Needs Chemotaxis To Establish Infection in Mammals and To Accomplish Its Enzootic Cycle.

Detection of spirochete burdens in microinjected nymphal ticks before and after feeding, using qRT-PCR. RNA samples were extracted from whole unfed ticks (10 days after injection) and fed ticks (after repletion; 5 to 7 days) and subjected to PCR analysis. The bacterial burdens in ticks were measured by the number of copies of flaB mRNA compared to the number of copies of tick β-actin transcript as previously described (, ). The data are presented as the means of relative levels of flaB transcript ± SEM for three groups (each group containing 3 unfed ticks or 4 replete ticks) for each strain (wild type, cheA2mut, and cheA2com).

Ching Wooen Sze, et al. Infect Immun. 2012 Jul;80(7):2485-2492.
6.
Fig 5

Fig 5. From: Borrelia burgdorferi Needs Chemotaxis To Establish Infection in Mammals and To Accomplish Its Enzootic Cycle.

Detection of spirochete burdens in needle-infected SCID mice by qRT-PCR. Groups of three mice were needle inoculated with 105 cells of the wild-type, cheA2mut, and cheA2com strains. Infected animals were sacrificed at 24, 48, and 72 h. Skin samples from areas around the inoculation sites were collected and subjected to qRT-PCR. The bacterial burdens in these samples were measured by determining the number of copies of flaB mRNA compared to the number of copies of mouse β-actin transcript as previously documented (, ). The data are expressed as the means of relative levels of flaB transcript ± SEM for three independent experiments. At 72 h, the flaB mRNA was undetectable in the skin tissues infected by cheA2mut. *, significant difference (P < 0.01). Bb, B. burgdorferi.

Ching Wooen Sze, et al. Infect Immun. 2012 Jul;80(7):2485-2492.
7.
Fig 4

Fig 4. From: Borrelia burgdorferi Needs Chemotaxis To Establish Infection in Mammals and To Accomplish Its Enzootic Cycle.

The B. burgdorferi cheA2 mutant is nonchemotactic. (A) Swarm plate assay. The assay was carried out on 0.35% agarose containing 1:10-diluted BSK-II medium as described previously (). The flaB mutant, a previously constructed nonmotile mutant (), was included to determine the sizes of the inocula. The data are presented as mean diameters (in millimeters) of rings ± SEM for four plates. (B) Capillary tube chemotaxis assay. Assays were carried out using 100 mM N-acetyl-d-glucosamine as an attractant according to a previous report (). The results are expressed as fold increases in cell number entering the capillary tubes containing the attractant relative to the number entering control tubes without attractant (buffer alone). Results are expressed as means ± SEM for five tubes. A 2-fold increase is considered the threshold for a positive response. *, significant difference (P < 0.01).

Ching Wooen Sze, et al. Infect Immun. 2012 Jul;80(7):2485-2492.

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