Genome misclassification of Klebsiella variicola and Klebsiella quasipneumoniae isolated from plants, animals and humans

Artículo original Martínez-Romero E y col. Genome misclassification of Klebsiella variicola and Klebsiella quasipneumoniae isolated from plants, animals and humans Esperanza Martínez-Romero, PhD,(1) Nadia Rodríguez-Medina, M in C,(2) Marilu Beltrán-Rojel, Biol,(2) Jesús Silva-Sánchez, PhD,(2) Humberto Barrios-Camacho, PhD,(2) Ernesto Pérez-Rueda, PhD,(3,4) Ulises Garza-Ramos, PhD.(2) Martínez-Romero E, Rodríguez-Medina N, Beltrán-Rojel M, Silva-Sánchez J, Barrios-Camacho H, Pérez-Rueda E, Garza-Ramos U. Genome misclassification of Klebsiella variicola and Klebsiella quasipneumoniae isolated from plants, animals and humans. Salud Publica Mex 2018;60:56-62. http://doi.org/10.21149/8149 Martínez-Romero E, Rodríguez-Medina N, Beltrán-Rojel M, Silva-Sánchez J, Barrios-Camacho H, Pérez-Rueda E, Garza-Ramos U. Clasificación errónea de aislados de Klebsiella variicola y Klebsiella quasipneumoniae obtenidos de plantas, animales y humanos. Salud Publica Mex 2018;60:56-62. http://doi.org/10.21149/8149 Abstract Resumen Keywords: bacterial infections; Gram-negative bacterial infections; Enterobacteriaceae infections; nitrogen fixation Palabras clave: infecciones bacterianas; infecciones por bacterias gramnegativas; fijación de nitrógeno Objective. Due to the fact that K. variicola, K. quasipneumoniae and K. pneumoniae are closely related bacterial species, misclassification can occur due to mistakes either in normal biochemical tests or during submission to public databases. The objective of this work was to identify K. variicola and K. quasipneumoniae genomes misclassified in GenBank database. Materials and methods. Both rpoB phylogenies and average nucleotide identity (ANI) were used to identify a significant number of misclassified Klebsiella spp. genomes. Results. Here we report an update of K. variicola and K. Quasipneumoniae genomes correctly classified and a list of isolated genomes obtained from humans, plants, animals and insects, described originally as K. pneumoniae or K. variicola, but known now to be misclassified. Conclusions. This work contributes to recognize the extensive presence of K. variicola and K. quasipneumoniae isolates in diverse sites and samples. (1) (2) (3) (4) Objetivo. Identificar genomas mal clasificados de K. variicola, y K. quasipneumoniae en la base de datos del GenBank. Material y métodos. En el presente estudio se usaron tanto análisis filogenéticos usando rpoB como la identidad media de nucleótidos (ANI, por sus siglas en ingles) para identificar un número significativo de genomas del género Klebsiella. Resultados. Se reportó una actualización de genomas de K. variicola y K. quasipneumoniae correctamente clasificados y una lista de aquellos aislamientos obtenidos de seres humanos, plantas, animales e insectos, descritos originalmente como K. pneumoniae o K. variicola pero ahora se conoce que están mal clasificados. Conclusiones. Este trabajo contribuye a la presencia extensiva de aislamientos de K. variicola y K. quasipneumoniae en diversos sitios y muestras. Centro de Ciencias Genómicas, Universidad Nacional Autónoma de México. Cuernavaca, Morelos, México. Grupo de Resistencia Bacteriana, Centro de Investigación sobre Enfermedades Infecciosas, Instituto Nacional de Salud Pública. Cuernavaca, Morelos, México. Departamento de Ingeniería Celular y Biocatálisis, Instituto de Biotecnología, Universidad Nacional Autónoma de México. México. Instituto de Investigaciones en Matemáticas Aplicadas y en Sistemas, Universidad Nacional Autónoma de México. Mérida,Yucatán, México. Received on: August 22, 2016 • Accepted on: June 13, 2017 Corresponding author: Dr. Ulises Garza Ramos. Departamento de Diagnóstico Epidemiológico, Instituto Nacional de Salud Pública. Av. Universidad 655 col. Santa María Ahuacatitlán. 62100, Cuernavaca, Morelos, México. E-mail: 56 salud pública de méxico / vol. 60, no. 1, enero-febrero de 2018 Genome misclassification of Klebsiella A recent taxonomic description of the genus Klebsiella spp. includes three taxonomic subspecies; K. pneumoniae subsp. pneumoniae (KpI), K. pneumoniae subsp. rhinoscleromatis and K. pneumoniae subsp. ozaenae and our novel species; Klebsiella variicola (KpIII), K. singaporensis, K. michiganensis and K. quasipneumoniae (KpII), which has two subspecies: Klebsiella quasipneumoniae subsp. quasipneumoniae (KpII-A) and K. quasipneumoniae subsp. similipneumoniae (KpII-B).1 For these bacterial species novel genome sequences have been described from the strains 07A044, 18A0691 and FIHV2014.2 The misclassification of K. variicola and Klebsiella spp. genomes was first identified by whole-genome and rpoB phylogenies.3,4 Subsequently by K. quasipneumoniae and other K. variicola genomes the misclassification was evidenced by Average Nucleotide Identity (ANI).5 The aim of this work was detect misclassified K. variicola and K. quasipneumoniae genomes. We used an rpoB gene phylogenetic analysis and the results were validated by ANI. We identified the isolates misclassified previously described5 and identified new isolates that corresponded to K. variicola and K. quasipneumoniae that were deposited as K. pneumoniae. Likewise, we identified one K. variicola genome misclassified that corresponds to K. quasipneumoniae subsp. similipneumoniae. Materials and methods Identification of the K. variicola and K. quasipneumoniae genomes The complete nucleotide sequence of rpoB (KVR801 v1_470024) gene from K. variicola 801 (CDMV00000000) was downloaded from the European Nucleotide Archive database.6 The rpoB sequence was analyzed by the BLASTn option (with default values) against 419 complete and draft Klebsiella spp. (tax id: 570) genomes deposited in GenBank database (last analysis 13/07/2015).7 The rpoB genes from Escherichia coli K-12-MG1655, Salmonella enterica Ty21a, Klebsiella oxytoca KCTC1686, K. quasipneumoniae subsp. quasipneumoniae 18A069 (KpII-A), K. quasipneumoniae subsp. quasipneumoniae 01A030 (KpII-A) and K. quasipneumoniae subsp. similipneumoniae 07A044 (KpII-B) genomes1 were included in the phylogenetic analysis. Subsequently, the rpoB nucleotide sequences of K. pneumoniae genomes with 100% nucleotide identity were filtered out to exclude a possible bias. All strain genomes identified as K. variicola and K. quasipneumoniae were included. Finally, a phylogenetic reconstruction was performed using the maximum-likelihood method with a Tamura-Nei-parameter model and 1 000 bootstrap replications (Mega v6.06).8 salud pública de méxico / vol. 60, no. 1, enero-febrero de 2018 Artículo original The ANI was determined using the Average Nucleotide Identity Calculator9 the default parameters. In total, thirty-one K. variicola, nine K. quasipneumoniae and two K. pneumonia genomes were downloaded from GenBank and analyzed (table I). K. variicola At-22 (CP001891), K. quasipneumoniae subsp. quasipneumoniae 18A069 (CBZM000000000), K. quasipneumoniae subsp. similipneumoniae 07A044 (CBZR000000000) and K. pneumon (...truncated)