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Open Access Research article

Decrease of virulence for BALB/c mice produced by continuous subculturing of Nocardia brasiliensis

Janeth A Almaguer-Chávez1, Oliverio Welsh1, Hector G Lozano-Garza2, Salvador Said-Fernández2, Víktor J Romero-Díaz3, Jorge Ocampo-Candiani1 and Lucio Vera-Cabrera1*

Author Affiliations

1 Servicio de Dermatología, Hospital Universitario "José E. González", Monterrey, N.L., 64460 México

2 Centro de Investigación Biomédica del Noreste, IMSS, Monterrey, N.L., 64720, México

3 Departamento de Histología, Facultad de Medicina, UANL., Monterrey, N.L., 64460 México

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BMC Infectious Diseases 2011, 11:290  doi:10.1186/1471-2334-11-290

Published: 26 October 2011

Abstract

Background

Subculturing has been extensively used to attenuate human pathogens. In this work we studied the effect of continuous subculturing of Nocardia brasiliensis HUJEG-1 on virulence in a murine model.

Methods

Nocardia brasiliensis HUJEG-1 was subcultured up to 130 times on brain heart infusion over four years. BALB/c mice were inoculated in the right foot pad with the bacteria subcultured 0, 40, 80, 100 and 130 times (T0, T40, T80 T100 and T130). The induction of resistance was tested by using T130 to inoculate a group of mice followed by challenge with T0 12 weeks later. Biopsies were taken from the newly infected foot-pad and immunostained with antibodies against CD4, CD8 and CD14 in order to analyze the in situ immunological changes.

Results

When using T40, T80 T100 and T130 as inoculums we observed lesions in 10, 5, 0 and 0 percent of the animals, respectively, at the end of 12 weeks. In contrast, their controls produced mycetoma in 80, 80, 70 and 60% of the inoculated animals. When studying the protection of T130, we observed a partial resistance to the infection. Immunostaining revealed an intense CD4+ lymphocytic and macrophage infiltrate in healing lesions.

Conclusions

After 130 in vitro passages of N. brasiliensis HUJEG-1 a severe decrease in its virulence was observed. Immunization of BALB/c mice, with these attenuated cells, produced a state of partial resistance to infection with the non-subcultured isolate.

Keywords:
Mycetoma; Nocardia; attenuation