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Brazilian journal of microbiology : [publication of the Brazilian Society for Microbiology]2025; 56(2); 1321-1331; doi: 10.1007/s42770-025-01640-x

Phenotypical and molecular characterization of Rhodococcus equi isolated from foals in the Agreste region of Pernambuco – Brazil.

Abstract: Equine rhodococcosis is caused by Rhodococcus equi, an intracellular coccobacillus whose main virulence factor is a plasmid that harbors genes encoding proteins from the Vap family, with the vapA gene being the most important in equine isolates. Furthermore, other factors observed in R. equi strains, such as antimicrobial resistance and biofilm production, may represent significant challenges in the treatment of affected animals. The objective of this study was to characterize four isolates of R. equi from foals in the state of Pernambuco, Brazil. All isolates were identified as R. equi through biochemical tests, amplification of the choE gene, and sequencing of 16 S rRNA. PCR analysis revealed that three isolates were positive for the plasmid virulence genes (vapA, -C, -D, -E, -F, -H and traA), although vapD was absent in one of the three isolates. One isolate did not present any virulence genes, possibly due to the loss of the plasmid after repeated passages at 37ºC. In the antimicrobial susceptibility test, all isolates were susceptible to erythromycin, clarithromycin, azithromycin, rifampicin, gentamicin, and doxycycline. However, all isolates were capable of forming biofilms, with moderate biofilm formation in isolates Rhodo1 and Rhodo2, and weak biofilm formation in isolates Rhodo3 and Rhodo4, which may be associated with increased antimicrobial tolerance. This molecular characterization demonstrated, for the first time, the presence of the virulence plasmid in R. equi isolates from foals in Northeast Brazil, as well as their capacity for biofilm formation.
© 2025. The Author(s) under exclusive licence to Sociedade Brasileira de Microbiologia.
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Publication Date: 2025-03-06 PubMed ID: 40048142PubMed Central: PMC12095708DOI: 10.1007/s42770-025-01640-xGoogle Scholar: Lookup
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  • Journal Article

Summary

This research summary has been generated with artificial intelligence and may contain errors and omissions. Refer to the original study to confirm details provided. Submit correction.

Overview

  • This study analyzes four Rhodococcus equi bacteria isolates from foals in Pernambuco, Brazil, focusing on their genetic traits, virulence factors, antimicrobial susceptibility, and ability to form biofilms.
  • It is the first report demonstrating that these isolates from Northeast Brazil carry key virulence plasmids and can form biofilms, which have implications for disease treatment and management.

Research Background

  • Rhodococcus equi is a bacterium causing equine rhodococcosis, a serious lung infection in foals.
  • It is an intracellular coccobacillus, meaning it can live and multiply within host cells, complicating treatment.
  • The primary virulence factor is a plasmid — a circular DNA segment separate from chromosomal DNA — carrying genes encoding virulence-associated proteins (Vap family).
  • The vapA gene in the plasmid is especially important because it is strongly linked to the bacterium’s ability to cause disease in horses.
  • Other bacterial traits like antimicrobial resistance and biofilm formation can further challenge effective treatment.

Study Objectives

  • To characterize phenotypically (biochemical identification, antibiotic susceptibility) and molecularly (gene amplification and sequencing) four R. equi isolates from foals located in the Agreste region of Pernambuco, Brazil.
  • To determine the presence of virulence genes and evaluate the biofilm-forming ability of these isolates.

Methodology

  • The four bacterial isolates were confirmed as R. equi by:
    • Performing biochemical tests
    • Amplifying the choE gene (a species-specific gene)
    • Sequencing the 16S rRNA gene, a common method for bacterial identification
  • Polymerase Chain Reaction (PCR) was used to detect virulence plasmid genes, including vapA, vapC, vapD, vapE, vapF, vapH, and traA.
  • Antimicrobial susceptibility tests were conducted for several antibiotics commonly used in treatment (erythromycin, clarithromycin, azithromycin, rifampicin, gentamicin, and doxycycline).
  • Biofilm formation was assessed, categorizing the isolates’ ability to form biofilms as weak or moderate.

Key Findings

  • Identification and Virulence Factors:
    • All isolates were confirmed as R. equi through phenotypic and molecular methods.
    • Three out of four isolates carried the virulence plasmid genes, suggesting pathogenic potential in these strains.
    • One isolate lacked the vapD gene within this set, indicating some variability in virulence gene content.
    • One isolate did not contain any tested virulence genes, potentially due to plasmid loss after repeated culture at 37ºC, which can occur during laboratory passages.
  • Antibiotic Susceptibility:
    • All isolates were susceptible to the antibiotics tested, including macrolides (erythromycin, clarithromycin, azithromycin), rifampicin, gentamicin, and doxycycline.
    • This susceptibility profile is favorable for treatment but still requires caution due to possible biofilm-associated tolerance.
  • Biofilm Formation:
    • All isolates demonstrated the capacity to form biofilms, a protective mode that bacteria adopt making them less vulnerable to antibiotics and host defenses.
    • Two isolates (Rhodo1 and Rhodo2) formed moderate biofilms, while Rhodo3 and Rhodo4 formed weaker biofilms.
    • Biofilm formation might contribute to persistent infection and challenges in antimicrobial therapy despite antibiotic susceptibility.

Significance and Implications

  • This is the first molecular characterization report confirming the presence of the virulence plasmid in R. equi isolates from foals in Northeastern Brazil, particularly Pernambuco.
  • Understanding virulence gene presence helps in assessing the pathogenic potential of bacterial strains circulating in local horse populations.
  • Recognition of biofilm formation ability highlights a potential mechanism of increased tolerance to antibiotics, which could complicate treatment protocols despite in vitro susceptibility.
  • The knowledge gained supports better diagnostic, therapeutic, and preventive measures for equine rhodococcosis in the region.
  • Further studies may be needed to investigate plasmid stability, long-term antibiotic resistance trends, and biofilm-related pathogenicity in these regional isolates.

Cite This Article

APA
Godoi APDS, Sobral GG, da Silva Vieira JC, Carneiro GF, Conceição FR, da Silva ER, Mendonça M. (2025). Phenotypical and molecular characterization of Rhodococcus equi isolated from foals in the Agreste region of Pernambuco – Brazil. Braz J Microbiol, 56(2), 1321-1331. https://doi.org/10.1007/s42770-025-01640-x

Publication

Brazilian journal of microbiology : [publication of the Brazilian Society for Microbiology]
ISSN: 1678-4405
NlmUniqueID: 101095924
Country: Brazil
Language: English
Volume: 56
Issue: 2
Pages: 1321-1331

Researcher Affiliations

Godoi, Alysson Paulo Dos Santos
  • Departamento de Medicina Veterinária, Universidade Federal Rural de Pernambuco, Rua Dom Manuel de Medeiros, Recife, Pernambuco, 52171-900, Brazil.
  • Programa de Pós-Graduação em Sanidade e Reprodução de Animais de Produção, Universidade Federal do Agreste de Pernambuco, Avenida Bom Pastor, Boa Vista, Garanhuns, 55292-270, Pernambuco, Brazil.
Sobral, Gilvannya Gonçalves
  • Departamento de Medicina Veterinária, Universidade Federal Rural de Pernambuco, Rua Dom Manuel de Medeiros, Recife, Pernambuco, 52171-900, Brazil.
da Silva Vieira, Júlio César
  • Programa de Pós-Graduação em Sanidade e Reprodução de Animais de Produção, Universidade Federal do Agreste de Pernambuco, Avenida Bom Pastor, Boa Vista, Garanhuns, 55292-270, Pernambuco, Brazil.
Carneiro, Gustavo Ferrer
  • Departamento de Medicina Veterinária, Universidade Federal Rural de Pernambuco, Rua Dom Manuel de Medeiros, Recife, Pernambuco, 52171-900, Brazil.
Conceição, Fabricio Rochedo
  • Centro de Desenvolvimento Tecnológico, Núcleo de Biotecnologia, Universidade Federal de Pelotas, Campus Capão do Leão, Pelotas, 96010-610, Rio Grande do Sul, Brazil.
da Silva, Elizabete Rodrigues
  • Programa de Pós-Graduação em Sanidade e Reprodução de Animais de Produção, Universidade Federal do Agreste de Pernambuco, Avenida Bom Pastor, Boa Vista, Garanhuns, 55292-270, Pernambuco, Brazil.
Mendonça, Marcelo
  • Programa de Pós-Graduação em Sanidade e Reprodução de Animais de Produção, Universidade Federal do Agreste de Pernambuco, Avenida Bom Pastor, Boa Vista, Garanhuns, 55292-270, Pernambuco, Brazil. marcelo.mendonca@ufape.edu.br.

MeSH Terms

  • Animals
  • Rhodococcus equi / genetics
  • Rhodococcus equi / isolation & purification
  • Rhodococcus equi / drug effects
  • Rhodococcus equi / classification
  • Rhodococcus equi / physiology
  • Horses / microbiology
  • Brazil
  • Horse Diseases / microbiology
  • Actinomycetales Infections / veterinary
  • Actinomycetales Infections / microbiology
  • Virulence Factors / genetics
  • Anti-Bacterial Agents / pharmacology
  • Microbial Sensitivity Tests
  • Plasmids / genetics
  • RNA, Ribosomal, 16S / genetics
  • Bacterial Proteins / genetics
  • Phylogeny
  • Phenotype
  • Virulence
  • Biofilms / growth & development

Grant Funding

  • 001 - Scholarship / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior

Conflict of Interest Statement

Declarations. Conflict of interest: None of the authors has any financial or personal relationships that could inappropriately influence or bias the content of the paper.

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