Genomic epidemiology analysis of Enterobacter hormaechei subsp. hoffmannii / 中华检验医学杂志

Objectives:To investigate the genetic characteristics of the blaNDM-1-carrying plasmid of the multidrug resistant Enterobacter hormaechei subsp. hoffmannii clinical isolate C37, and constructing a phylogenetic tree of the 66 publicly available genomes of the Enterobacter hormaechei subsp. hoffmannii to explore its global epidemic situation. Methods:Carbapenem-resistant Enterobacter cloacae complex (CRECC) strains isolated from the First Affiliated Hospital of Sun Yat-sen University from August 2014 to August 2021 were collected. Matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS) and 16S rDNA Sanger sequencing were used for species identification. Micro broth dilution method was used for antibacterial susceptibility test. The polymerase chain reaction (PCR) was used to detect the β-lactamase resistance gene and plasmid-mediated quinolone resistance (PMQR) gene carried by the C37 strain. The conjugation experiment was used to confirm the conjugative metastasis of the resistance genes in C37 strain. Whole genome of the C37 strain was sequenced. Core genome was extracted and the phylogenetic analysis of 66 publicly available Enterobacter hormaechei subsp. hoffmannii was performed. Results:Enterobacter hormaechei subsp. hoffmannii C37 strain is resistant to third-generation cephalosporins, carbapenems, aminoglycosides and quinolones, and carries blaACT-5, blaNDM-1, qnrA1, aac(6′)-Ib-cr, oqxAB, fosA, dfrA15 and other resistance genes, as well as IncX3, IncX4, IncFIB and IncFII plasmids. Multilocus sequence typing (MLST) analysis showed that C37 strain belongs to the ST78 type of Enterobacter cloacae complex (ECC). Conclusions:ST78 type Enterobacter hormaechei subsp. hoffmannii is closely related to the spread of carbapenem resistance genes. It is a potential high-risk clone to spread carbapenem resistance genes. The prevalence and trends of ST78 type Enterobacter hormaechei subsp. Hoffmannii should be monitored.

Molecular, biochemical, and phenotypic characterization of a newly isolated Enterobacter hormaechei subsp. xiangfangensis strain associated with diarrhea cases in Iraq

Aims@#The current study aimed to isolate and characterize bacterial strains associated diarrhea with the focus on Enterobacter species strains and test for susceptibility to antibiotics.@*Methodology and results@#A total of 400 stool samples from inpatients suffering from diarrhea in Al-Qasim Hospital at Al-Hilla City of Iraq were screened form January 2018 to January 2019. Phenotypic, molecular, and biochemical methods were used to identify the isolated bacteria. A new strain of Enterobacter hormaechei was obtained from two stool samples of inpatients suffering from diarrhea for more than two weeks. This strain is Gram negative, rod shaped, and facultative anaerobic. Multiple sequence alignment analysis and phylogenetic tree construction of the sequenced 16S rRNA gene of the isolated strain suggested that this strain can be identified as E. hormaechei subsp. xiangfangensis, named as E. hormaechei subsp. xiangfangensis strain AA1. This strain was resistant to augmentin, ampicillin, cephalothin, cefoxitin, ceftazidime, cefixime, ticracillin/clavulanic acid, cefotaxime, streptomycin, erythromycin, amikacin, ciprofloxacin, and chloramphenicol, while it was susceptible to meropenem along with imipenem. @*Conclusion, significance and impact of study@# In the present study, E. hormaechei subsp xiangfangensis was isolated for the first time in Iraq and was resistant to most of the tested antibiotics, making it an etiologic agent that is not easy to be treated.

Estudo genotípico e fenotípico de bacilos Gram-negativos produtores de carbapenemase do tipo New Delhi metalo-ß-lactamase / Genotypic and fenotypic study of Gram-negative bacilli producers carbapenemase type New Delhi metallo--ß-lactamase

Os carbapenêmicos são os antimicrobianos mais amplamente utilizados no tratamento empírico de infecções graves por bacilos Gram-negativos. A pressão seletiva gerada pelo uso desses antimicrobianos ao longo das últimas três décadas contribuiu para a disseminação de enterobactérias e Gram-negativos não fermentadores produtores de carbapenemases, particularmente as do tipo KPC e NDM. Os genes que codificam essas enzimas usualmente estão localizados em plasmídeos e/ou transpósons. A hipótese atualmente mais aceita é que o gene blaNDM-1 seja uma quimera criada em Acinetobacter baumannii. A NDM-1 foi descrita em paciente proveniente da Índia e subsequentemente evidenciou-se sua ampla disseminação nesse país. A epidemiologia que tem sido observada nos casos detectados na Europa e Estados Unidos tem sido viagem à Índia, ou seja, sem casos autóctones. No Brasil, os primeiros casos foram identificados no Rio Grande do Sul, e a seguir no Rio de Janeiro e em São Paulo. Diferentemente dos casos da Europa e América do Norte, os casos do Brasil não tem relação epidemiológica com a Índia. O sequenciamento integral dos plasmídeos e cromossomos albergando o gene blaNDM permitirá entender como ocorre a disseminação desse mecanismo de resistência no Brasil. Para isso, foi avaliado o perfil de susceptibilidade dos isolados, bem como a capacidade conjugativa e clonalidade. Das vinte e oito amostras utilizadas neste trabalho, treze delas pertencem à espécie Enterobacter hormaechei, uma à espécie Citrobacter freundii, sete à espécie Escherichia coli, quatro à Klebsiella pneumoniae e três ao gênero Acinetobacter spp. Os primeiros isolados incluídos neste estudo (Escherichia coli e Enterobacter hormaechei produzindo NDM-1) foram isolados em agosto de 2013, de uma mesma amostra de swab retal de um paciente do Rio de Janeiro que nunca viajou para o exterior. O sequenciamento completo do DNA plasmidial utilizando a plataforma Illumina e a anotação de ambos os plasmídeos albergando o gene blaNDM-1 revelou que estes pertencem a grupos de incompatibilidade diferentes, IncFIIK (E. hormaechei) e IncX3 (E. coli), e abrigam um novo transpóson composto designado Tn3000. A comparação da sequência nucleotídica do Tn3000 com aquelas disponíveis no GenBank evidencia que a mesma estrutura está presente em plasmídeos de isolados da cidade de Porto Alegre e também em diferentes continentes. As espécies de Acinetobacter (A. radioresistens, A. ursingii e A. guillouiae) isoladas em São Paulo e Porto Alegre, possuem o gene blaNDM-1 albergados em um mesmo plasmídeo não tipável de 41.087 pb. A avaliação da clonalidade dos isolados de Enterobacter hormaechei "subsp. oharae" mostrou dois perfis diferentes através da técnica de PFGE, sendo que todos os microrganismos foram isolados de um surto no mesmo hospital no Rio de Janeiro. Isolados de Klebsiella pneumoniae de uma mesma paciente internada em hospital em Salvador, de sítios distintos - swab retal, hemocultura e urina, em ordem cronológica - obtiveram o mesmo perfil clonal pela técnica de PFGE. O mesmo ocorreu com três isolados de Escherichia coli, de um mesmo paciente do Rio de Janeiro, em amostras de swab retal. Os achados deste estudo evidenciam que no Brasil, Nepal, Marrocos e Índia há uma disseminação do gene blaNDM-1 mediada por um novo elemento móvel designado Tn3000 em enterobactérias. A detecção de um mesmo plasmídeo em diferentes espécies de Acinetobacter evidencia que neste gênero bacteriano, no Brasil, a disseminação do gene blaNDM-1 ocorre por conjugação.

Carbapenems are the antimicrobials most widely used in the empirical treatment of severe infections caused by Gram-negative bacilli. The selective pressure generated by the use of these antibiotics over the last three decades has contributed to the spread of enterobacteria and Gram-negative non-fermenting producing carbapenemases, mainly KPC and NDM. Genes encoding these enzymes are usually located in plasmids and/or transposons. Currently the most accepted hypothesis is that the blaNDM-1 gene is a chimera created in Acinetobacter baumannii. The NDM-1 was described in a patient from India and subsequently was reported to be broadly disseminate in this country. The epidemiology that has been observed in cases detected in Europe and United States is traveling to India, but no autochthonous cases. In Brazil, the first cases were identified in Rio Grande do Sul, and then in Rio de Janeiro and São Paulo. Differently from the cases described in Europe and North America, the cases from Brazil have no epidemiological link with India. The complete sequencing of plasmids and chromosomes harboring blaNDM gene will understanding how the dissemination of this resistance mechanism in Brazil occurs. In this work we will be evaluate the susceptibility profile of the isolates, and their conjugal capacity and clonality. Of the twenty-eight samples used in this study, thirteen of them belong to the species Enterobacter hormaechei, one to Citrobacter freundii, seven to Escherichia coli, four to Klebsiella pneumoniae and three to the genus Acinetobacter sp. The first two isolates included in this study (Escherichia coli and Enterobacter hormaechei) were isolated in August 2013, from the same rectal swab sample from a patient from Rio de Janeiro that never traveled abroad. Complete sequencing of plasmid DNA using Illumina platform and annotation of both plasmids harboring the blaNDM-1 gene revealed that they belong to different incompatibility groups, IncFIIK (E. hormaechei) and IncX3 (E. coli), and are harbor to a new transposon designated Tn3000. The comparison of the Tn3000 nucleotide sequence with those available at GenBank shows that the same structure is present in plasmids from other Porto Alegre and also in different continents. The Acinetobacter species (A. radioresistens, A. ursingii and A. guillouiae) isolated in São Paulo and Porto Alegre, have the blaNDM-1 gene harbored in a single non-typing plasmid of 41,087 bp. The evaluation of clonal relationship of Enterobacter hormaechei "subsp. oharae" showed two different profiles by PFGE technique; of note all microorganisms were isolated from an outbreak in the same hospital in Rio de Janeiro. Isolates of Klebsiella pneumoniae from a single patient hospitalized in Salvador, from different anatomical sites - rectal swab, blood culture and urine, in chronological order - obtained the same clonal profile by the PFGE technique. The same occurred with three Escherichia coli isolates, from the same patient from Rio de Janeiro, in swab rectal strains. Our findings suggest that in Brazil, Nepal, Morocco and India there is a spread of blaNDM-1 gene mediated by Tn3000 in enterobacteria. The detection of a same plasmid in different species of Acinetobacter shows that in this bacterial genus, in Brazil, the dissemination of the blaNDM-1 gene occurs by conjugation.

Molecular identification of multidrug resistant Enterobacter hormaechei in Venezuela / Identificación molecular de Enterobacter hormaechei multidrogoresistente en Venezuela

Besides the importance of Enterobacter cloacae species complex as a nosocomial pathogen, little is known about the frequency of each species/genotype. Here, we describe a strain of E. hormaechei subsp. hormaechei isolated from a bronchial secretion of a patient, in the Intensive Care Unit at the General Hospital of Cumaná, Venezuela, who died due to complications of his infection. The molecular identification was done by sequencing the 16S rRNA gene and comparing it to sequences from the GenBank. This strain showed resistance to multiple families of antibiotics (MDR), and the genes blaKPC and blaVIM were detected by PCR. This is the first time E. hormaechei has been identified in Venezuela.

A pesar de la importancia de las especies del complejo Enterobacter cloacae como patógeno nosocomial, poco se conoce sobre la frecuencia de cada especie/genotipo. Aquí se describe una cepa de E. hormaechei subsp. hormaechei aislada de una secreción bronquial de un paciente internado en la Unidad de Cuidados Intensivos del Hospital General de Cumaná, Venezuela, quien murió producto de complicaciones de su infección. La identificación molecular fue hecha por secuenciación del gen ARNr 16S y porcomparación con las secuencias del GenBank. Esta cepa mostró resistencia a múltiples familias de antibióticos (MDR) y se detectaron los genes blaKPCyblaVIMpor PCR. Este es el primer reporte de E. hormaechei en Venezuela.

Two Cases of Enterobacter hormaechei Infection Misidentified as Cedecea davisae Infection

Enterobacteriaceae is a family of gram-negative, rod-shaped bacteria that consists of various species. Among these, members of the genus Cedecea has been reported as relatively rare causative pathogens of human infections. Commercially available automated identification systems that use biochemical reactions are known to accurately identify Enterobacteriaceae species. However, the accurate identification of some organisms with diverse biochemical profiles by these automated identification systems may be problematic. In this study, we report two cases of isolate misidentification, from patients with acute cholecystitis and deep vein thrombosis, as Cedecea davisae with VITEK II system. Both the isolates were correctly identified as Enterobacter hormaechei using gyrB gene sequence analysis. We also performed 16S rRNA sequence analyses and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry analyses; however, indeterminate results were obtained from both the assays. Therefore, the sequence analysis of alternative genes, like gyrB, might be useful for accurate identification of species that belong to the family of Enterobacteriaceae.