Prevalence and diversity of non-tuberculous mycobacteria isolated from slaughtered tuberculin-positive bovine samples: Epidemiological and Diagnostic implications

Natalia Jiménez-Pizarro; David Risco; Felipe Molina; Remigio Martínez; Alfredo García; José Manuel Benítez-Medina; Javier Hermoso-de-Mendoza

doi:10.1016/j.vetmic.2026.110909

Prevalence and diversity of non-tuberculous mycobacteria isolated from slaughtered tuberculin-positive bovine samples: Epidemiological and Diagnostic implications

Vet Microbiol. 2026 Jan 29:315:110909. doi: 10.1016/j.vetmic.2026.110909. Online ahead of print.

Authors

Natalia Jiménez-Pizarro¹, David Risco², Felipe Molina³, Remigio Martínez⁴, Alfredo García⁵, José Manuel Benítez-Medina⁶, Javier Hermoso-de-Mendoza⁷

Affiliations

¹ Animal Health Department, Faculty of Veterinary, University of Extremadura, Cáceres 10003, Spain. Electronic address: nataliajp@unex.es.
² Animal Medicine Department. Faculty of Veterinary, University of Extremadura, Cáceres 10003, Spain. Electronic address: riscope@unex.es.
³ Biochemistry and Molecular Biology and Genetics Department, Faculty of Science, University of Extremadura, Badajoz 06071, Spain. Electronic address: fmolina@unex.es.
⁴ Departamento de Sanidad Animal, Grupo de Investigación en Sanidad Animal y Zoonosis (GISAZ), UIC Zoonosis y Enfermedades Emergentes ENZOEM, Universidad de Córdoba, Córdoba 14014, Spain. Electronic address: sa2maper@uco.es.
⁵ Animal Production Area, CICYTEX-La Orden, Badajoz 06187, Spain. Electronic address: alfredo.garcia@juntaex.es.
⁶ Animal Health Department, Faculty of Veterinary, University of Extremadura, Cáceres 10003, Spain. Electronic address: jmbenimed@unex.es.
⁷ Animal Health Department, Faculty of Veterinary, University of Extremadura, Cáceres 10003, Spain. Electronic address: jhermoso@unex.es.

PMID: 41637965
DOI: 10.1016/j.vetmic.2026.110909

Abstract

The isolation of non-tuberculous mycobacteria (NTM) is common in cattle positive for tuberculosis (TB) in official diagnostic tests, whereas data on specific NTM species in Spanish cattle remain limited. This study identifies the most frequently isolated NTM species from Single Intradermal Tuberculin Test (SITT)-positive cattle in Extremadura, western Spain. Among 1669 Mycobacterium Growth Indicator Tube (MGIT) positive cultures collected in 2018, 493 (29.54 %) were identified as NTM, and 194 were randomly selected for further analysis. Polymerase Chain Reaction (PCR)-restriction analysis of the hsp65 gene and partial sequencing of 16S ribosomal DNA (rDNA) confirmed a diverse range of species. The most prevalent complex was Mycobacterium avium (40.12 %), including M. senriense, M. intracellulare, and M. avium subsp. paratuberculosis. Other notable NTM species (23.35 %) included M. bourgelatii, M. kansasii, M. gordonae, and M. shinjukuense. Less frequent complexes included M. simiae (11.38 %), M. ulcerans (3.59 %), M. parafortuitum (2.99 %), and M. terrae (1.20 %), along with M. holsaticum (1.20 %), a species related to the M. tuberculosis complex. Phylogenetic analysis and geographic mapping revealed weak correlation between genetic and geographic distances (Mantel test: R_xy = 0.015, P = 0.253), suggesting limited spatial structuration of genetic diversity. Alpha diversity metrics indicated moderate diversity (Shannon's H = 2.641, Simpson's D = 0.106), with some zones exhibiting greater species evenness. Diversity analyses showed moderate dissimilarity among clusters. These findings enhance understanding of Mycobacterium diversity and distribution while emphasizing the diagnostic challenges posed by NTM in TB detection and the importance of molecular tools in species identification and epidemiological surveillance.

Keywords: 16S rDNA sequencing; Alpha-Beta diversity; Cattle; Genetic diversity; Non-tuberculous mycobacteria; PCR-restriction analysis; Shannon-Simpson indices.