Validation of Bartonella henselae Western Immunoblotting for Serodiagnosis of Bartonelloses in Dogs - PubMed (original) (raw)

Validation of Bartonella henselae Western Immunoblotting for Serodiagnosis of Bartonelloses in Dogs

Pradeep Neupane et al. J Clin Microbiol. 2020.

Abstract

Bartonella spp. are etiological agents of life-threatening zoonotic diseases in dogs worldwide. Due to the poor sensitivity of immunofluorescent-antibody assays (IFAs), a reliable serodiagnostic test for canine bartonelloses is of clinical importance. The utility of Western blotting (WB) for the serodiagnosis of canine bartonelloses has not been critically investigated. The objective of this study was to characterize WB immunodominant proteins that could be used to confirm a serodiagnosis of bartonelloses. Using agar-grown Bartonella henselae San Antonio type 2 (SA2) whole-cell proteins, sera derived from four dog groups were tested by WB to assess immunodominant protein recognition patterns: group I consisted of 92 serum samples (10 preexposure and 82 postexposure serum samples) from 10 adult beagles experimentally inoculated with Bartonella spp., group II consisted of 36 serum samples from Bartonella PCR-positive naturally infected dogs, group III consisted of 26 serum samples from Bartonella PCR-negative and IFA-negative dogs, and group IV consisted of serum samples from 8 Brucella canis IFA-positive and 10 Rickettsia rickettsii IFA-positive dogs. Following experimental inoculation, 9 (90%) group I dogs were variably seroreactive to one or more of six specific immunodominant proteins (13, 17, 29, 50, 56, and 150 kDa). There was a strong but variable recognition of these proteins among 81% of group II dogs. In contrast, 24/26 group III dogs were not reactive to any immunodominant protein. In this study, the sensitivity and diagnostic accuracy of B. henselae SA2 WB were higher than those of B. henselae SA2 IFA testing. Some B. henselae SA2 immunodominant proteins were recognized by dogs experimentally and naturally infected with Bartonella spp. other than B. henselae Additional research is necessary to more fully define the utility of WB for the serodiagnosis of canine bartonelloses.

Keywords: Bartonella henselae; Western blotting; antigen; serology; vector borne; zoonosis.

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Figures

FIG 1

Representative images of WB results for group I dogs (dogs experimentally inoculated with Bartonella henselae), group II dogs (naturally infected Bartonella PCR-positive dogs), and group III dogs (Bartonella PCR-negative and IFA-negative dogs). Whole-cell proteins extracted from agar-grown Bartonella henselae San Antonio type 2 were used as WB antigens. WB was performed using alkaline phosphatase-conjugated goat anti-dog IgG (H+L). (A) Image of WB results for a group I dog. Lanes 1 to 5, image of blotting results for a dog (6GCB11) experimentally inoculated with B. henselae for preexposure serum (lane 1) and serum collected at postinoculation days 6, 10, 13, and 24 (lanes 2 to 5, respectively). (B) Image of WB results for selected group II dogs. Lanes 1 to 5, each lane represents the blot image for one of five group II dogs. (C) Image of WB results for group III dogs. Lanes 1 and 2, WB images for two group III dogs. The gel images in panels A, B and C are from three separate gels. The white space within each image differentiates different areas of the same gel to show the variation in WB seroreactivity over the experimental study period (for the group I dog) or for group II dogs infected with different Bartonella spp. Images were selected and are depicted by group designations I to III. IgG IFA titers of ≥1:64 were considered positive for Bartonella exposure. Bh, B. henselae; Bvb I, B. vinsonii subsp. berkhoffii genotype I; Bcl, B. clarridgeiae; NEG, negative; lanes L, Bio-Rad Precision Plus Kaleidoscope protein standard.

FIG 2

Frequency of WB bands among group I dogs (dogs experimentally inoculated with Bartonella spp.) and group II dogs (Bartonella PCR-positive naturally infected dogs). The percentage of dogs recognizing each B. henselae protein on WB is represented as WB seroreactivity (in percent) in the histogram.

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Validation of Bartonella henselae Western Immunoblotting for Serodiagnosis of Bartonelloses in Dogs - PubMed (original) (raw)