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Improved culture from lymph nodes of patients with cat scratch disease and genotypic characterization of Bartonella henselae isolates in Australia.

ABSTRACT: Over a 4-year period we detected Bartonella henselae isolates in 104 of 297 specimens (35.1%) from Australian patients clinically suspected of having cat scratch disease by amplification of a fragment of the htrA gene. We isolated 17 B. henselae strains (20.5%) from the 83 PCR-positive human specimens available for culture. Our culture method was based on prolonged incubation in a moist atmosphere of blood agar to which hemin was added. We obtained more B. henselae isolates than the number of all other isolates from lymph nodes reported in the literature. In order to identify and study the genetic variation of Australian B. henselae isolates, we determined the sequence of a fragment of the pap31 gene from our 17 human isolates and also from 8 Australian cat isolates. Thirteen of the human B. henselae isolates belonged to the Houston genotype, variant Houston-1 (76.5%), and four belonged to the Marseille genotype, variant CAL-1 (23.5%). In contrast, seven cat isolates were classified as B. henselae Marseille, variant CAL-1 (87.5%), and one was classified as B. henselae Houston, variant Houston-1 (12.5%). Our study describes an efficient culture method for the diagnosis of cat scratch disease and contributes to the description of the genotypic distribution of B. henselae in Australia.

SUBMITTER: Fournier PE

PROVIDER: S-EPMC130866 | biostudies-literature | 2002 Oct

REPOSITORIES: biostudies-literature

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Improved culture from lymph nodes of patients with cat scratch disease and genotypic characterization of Bartonella henselae isolates in Australia.

Fournier Pierre-Edouard PE Robson Jenny J Zeaiter Zaher Z McDougall Rodney R Byrne Shane S Raoult Didier D

Journal of clinical microbiology 20021001 10

Over a 4-year period we detected Bartonella henselae isolates in 104 of 297 specimens (35.1%) from Australian patients clinically suspected of having cat scratch disease by amplification of a fragment of the htrA gene. We isolated 17 B. henselae strains (20.5%) from the 83 PCR-positive human specimens available for culture. Our culture method was based on prolonged incubation in a moist atmosphere of blood agar to which hemin was added. We obtained more B. henselae isolates than the number of al ...[more]

PMID: 12354855

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Project description:Cat scratch disease (CSD) is an infectious disease caused by Bartonella henselae, usually characterized by self-limiting regional lymphadenopathy and fever. Given the low clinical diagnostic sensitivity and specificity of conventional anti-B. henselae indirect immunofluorescence assays (IFAs), real-time polymerase chain reaction (PCR)-based detection of B. henselae is now being proposed as a more sensitive tool to diagnose CSD. Thus, here we have assessed the efficacy of real-time PCR in detecting B. henselae in different specimens from patients with suspected CSD and compared it to that of IFA. From March 2011 to May 2016, at the Microbiology and Virology Unit, Azienda Ospedaliera Universitaria Città della Salute e della Scienza di Torino, Turin, Italy, 115 clinical specimens (56 aspirated pus, 39 fresh lymph node biopsies, and 20 whole blood samples) and 99 sera from 115 patients with suspected CSD (62 females and 53 males between the ages of 3 months and 68 years) were analyzed by both real-time PCR, used in a qualitative way, and IFA (IgM and IgG) for the presence of B. henselae. For 16 patients, serological results were not available due to a clinical decision not to request the test. B. henselae DNA positivity was detected by real-time PCR in 37.39% of patients, while 62.61% of them were negative. Thus, patients were divided into two groups: real-time PCR+ (n = 43) and real-time PCR- (n = 72). Real-time PCR screening of whole blood, biopsies, and aspirated pus revealed B. henselae positivity in 40%, 38.46%, and 35.71% of patients, respectively. When we analyzed samples by IFA, we found the presence of B. henselae in 28 out of 99 (28.28%) patients, of which 11 (11.11%) belonged to the real-time PCR+ group and 17 (17.17%) to the real-time PCR- group. Among the 71 seronegative subjects, 16 (16.16%) were found positive for B. henselae by real-time PCR. Thus, by combining the results of both assays, we were able to increase the percentage of B. henselae positive specimens from 27.27% (real-time PCR) or 28.28% (IFA) to 44.44% (real-time PCR+IFA). Altogether, these findings indicate that the early detection of B. henselae in patients with suspicious CSD through combined real-time PCR and serological analyses can lead to a more accurate diagnosis of CSD, thereby allowing prompt and appropriate disease management.

Project description:Bartonella henselae is a Gram-negative bacterium that causes cat scratch disease (CSD), as well as bacteremia, endocarditis, and other clinical presentations. CSD remains one of the most common infections caused by bacteria in the genus Bartonella, and it is transmitted to humans through a scratch or cat bite. Vaccination and more efficient diagnostic methods would represent a promising and sustainable alternative measure for CSD control in humans and animals. Here, we described the in silico analyses and design of three recombinant chimeric proteins (rC1, rC2, and rC3), for use in the control of CSD. The chimeras were constructed with epitopes identified from the sequences of the GroEL, 17 kDa, P26, BadA, Pap31, OMP 89, and OMP 43, previously described as the most important B. henselae antigens. The rC1, rC2, and rC3 were expressed and purified using a heterologous system based on Escherichia coli and reacted with antibodies present in the sera of humans naturally infected. The chimeric proteins were used to immunize mice using Freund adjuvant, and the humoral immune response was evaluated. Animals immunized with rC1 and rC3 showed a significant IgG antibodies response from the 28th day (P < 0.05), and the animals immunized with the rC2 from the 35th day (P < 0.05) remained until the 56th day of experimentation, with a titer of 1:3200 (P < 0.05), 1:1600 (P < 0.05) and 1:1600 (P < 0.05) from rC1, rC2, and rC3, respectively. Significant production of IgA and IgG1 isotype was detected in animals immunized with rC1 and rC2 proteins. Additionally, analysis using 13 serum samples from naturally infected patients showed that the proteins are recognized by antibodies present in sera, reinforcing the possibility of using these chimeras for CSD control. KEY POINTS: • The recombinant chimeras were expressed in Escherichia coli with 37 kDa (rC1), 35 kDa (rC2), and 38 kDa (rC3). • Animals immunized with rC1, rC2, and rC3 showed significant antibody response. • The chimeras were recognized by the sera of naturally infected patients.

Dataset Information

Improved culture from lymph nodes of patients with cat scratch disease and genotypic characterization of Bartonella henselae isolates in Australia.

Publications

Improved culture from lymph nodes of patients with cat scratch disease and genotypic characterization of Bartonella henselae isolates in Australia.

Similar Datasets

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