Stepwise Evolution of a Klebsiella pneumoniae Clone within a Host Leading to Increased Multidrug Resistance

ABSTRACT Five blaCTX-M-14-positive Klebsiella pneumoniae isolates (KpWEA1, KpWEA2, KpWEA3, KpWEA4-1, and KpWEA4-2) were consecutively obtained from a patient with relapsed acute myeloid leukemia who was continuously administered antimicrobials. Compared with KpWEA1 and KpWEA2, KpWEA3 showed decrease...

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Main Authors: Mai Yoshino, Masamune Aihara, Yasuhiro Gotoh, Masaru Akimoto, Wakana Tatsuhara, Makiko Kiyosuke, Yuichi Matsushima, Takeshi Uchiumi, Tetsuya Hayashi, Dongchon Kang
Format: Article
Language:English
Published: American Society for Microbiology 2021-12-01
Series:mSphere
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Online Access:https://journals.asm.org/doi/10.1128/mSphere.00734-21
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author Mai Yoshino
Masamune Aihara
Yasuhiro Gotoh
Masaru Akimoto
Wakana Tatsuhara
Makiko Kiyosuke
Yuichi Matsushima
Takeshi Uchiumi
Tetsuya Hayashi
Dongchon Kang
author_facet Mai Yoshino
Masamune Aihara
Yasuhiro Gotoh
Masaru Akimoto
Wakana Tatsuhara
Makiko Kiyosuke
Yuichi Matsushima
Takeshi Uchiumi
Tetsuya Hayashi
Dongchon Kang
author_sort Mai Yoshino
collection DOAJ
description ABSTRACT Five blaCTX-M-14-positive Klebsiella pneumoniae isolates (KpWEA1, KpWEA2, KpWEA3, KpWEA4-1, and KpWEA4-2) were consecutively obtained from a patient with relapsed acute myeloid leukemia who was continuously administered antimicrobials. Compared with KpWEA1 and KpWEA2, KpWEA3 showed decreased susceptibility to antimicrobials, and KpWEA4-1 and KpWEA4-2 (isolated from a single specimen) showed further-elevated multidrug-resistance (MDR) phenotypes. This study aims to clarify the clonality of the five isolates and their evolutionary processes leading to MDR by comparison of these complete genomes. The genome comparison revealed KpWEA1 was the antecedent of the other four isolates, and KpWEA4-1 and KpWEA4-2 independently emerged from KpWEA3. Increasing levels of MDR were acquired by gradual accumulation of genetic alterations related to outer membrane protein expression: the loss of OmpK35 and upregulation of AcrAB-TolC occurred in KpWEA3 due to ramA overexpression caused by a mutation in ramR; then OmpK36 was lost in KpWEA4-1 and KpWEA4-2 by different mechanisms. KpWEA4-2 further acquired colistin resistance by the deletion of mgrB. In addition, we found that exuR and kdgR, which encode repressors of hexuronate metabolism-related genes, were disrupted in different ways in KpWEA4-1 and KpWEA4-2. The two isolates also possessed different amino acid substitutions in AtpG, which occurred at very close positions. These genetic alterations related to metabolisms may compensate for the deleterious effects of major porin loss. Thus, our present study reveals the evolutionary process of a K. pneumoniae clone leading to MDR and also suggests specific survival strategies in the bacteria that acquired MDR by the genome evolution. IMPORTANCE Within-host evolution is a survival strategy that can occur in many pathogens and is often associated with the emergence of novel antimicrobial-resistant (AMR) bacteria. To analyze this process, suitable sets of clinical isolates are required. Here, we analyzed five Klebsiella pneumoniae isolates which were consecutively isolated from a patient and showed a gradual increase in the AMR level. By genome sequencing and other analyses, we show that the first isolate was the antecedent of the later isolates and that they gained increased levels of antimicrobial resistance leading to multidrug resistance (MDR) by stepwise changes in the expression of outer membrane proteins. The isolates showing higher levels of MDR lost major porins but still colonized the patient’s gut, suggesting that the deleterious effects of porin loss were compensated for by the mutations in hexuronate metabolism-related genes and atpG, which were commonly detected in the MDR isolates.
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spelling doaj.art-f6c9b938967d4d3381371e57ede960212022-12-21T19:38:11ZengAmerican Society for MicrobiologymSphere2379-50422021-12-016610.1128/mSphere.00734-21Stepwise Evolution of a Klebsiella pneumoniae Clone within a Host Leading to Increased Multidrug ResistanceMai Yoshino0Masamune Aihara1Yasuhiro Gotoh2Masaru Akimoto3Wakana Tatsuhara4Makiko Kiyosuke5Yuichi Matsushima6Takeshi Uchiumi7Tetsuya Hayashi8Dongchon Kang9Department of Clinical Chemistry and Laboratory Medicine, Kyushu University Hospital, Fukuoka, JapanDepartment of Clinical Chemistry and Laboratory Medicine, Kyushu University Hospital, Fukuoka, JapanDepartment of Bacteriology, Graduate School of Medical Sciences, Kyushu University, Fukuoka, JapanDepartment of Clinical Chemistry and Laboratory Medicine, Kyushu University Hospital, Fukuoka, JapanDepartment of Health Science, Graduate School of Medical Sciences, Kyushu University, Fukuoka, JapanDepartment of Clinical Chemistry and Laboratory Medicine, Kyushu University Hospital, Fukuoka, JapanDepartment of Clinical Chemistry and Laboratory Medicine, Graduate School of Medical Sciences, Kyushu University, Fukuoka, JapanDepartment of Health Science, Graduate School of Medical Sciences, Kyushu University, Fukuoka, JapanDepartment of Bacteriology, Graduate School of Medical Sciences, Kyushu University, Fukuoka, JapanDepartment of Clinical Chemistry and Laboratory Medicine, Kyushu University Hospital, Fukuoka, JapanABSTRACT Five blaCTX-M-14-positive Klebsiella pneumoniae isolates (KpWEA1, KpWEA2, KpWEA3, KpWEA4-1, and KpWEA4-2) were consecutively obtained from a patient with relapsed acute myeloid leukemia who was continuously administered antimicrobials. Compared with KpWEA1 and KpWEA2, KpWEA3 showed decreased susceptibility to antimicrobials, and KpWEA4-1 and KpWEA4-2 (isolated from a single specimen) showed further-elevated multidrug-resistance (MDR) phenotypes. This study aims to clarify the clonality of the five isolates and their evolutionary processes leading to MDR by comparison of these complete genomes. The genome comparison revealed KpWEA1 was the antecedent of the other four isolates, and KpWEA4-1 and KpWEA4-2 independently emerged from KpWEA3. Increasing levels of MDR were acquired by gradual accumulation of genetic alterations related to outer membrane protein expression: the loss of OmpK35 and upregulation of AcrAB-TolC occurred in KpWEA3 due to ramA overexpression caused by a mutation in ramR; then OmpK36 was lost in KpWEA4-1 and KpWEA4-2 by different mechanisms. KpWEA4-2 further acquired colistin resistance by the deletion of mgrB. In addition, we found that exuR and kdgR, which encode repressors of hexuronate metabolism-related genes, were disrupted in different ways in KpWEA4-1 and KpWEA4-2. The two isolates also possessed different amino acid substitutions in AtpG, which occurred at very close positions. These genetic alterations related to metabolisms may compensate for the deleterious effects of major porin loss. Thus, our present study reveals the evolutionary process of a K. pneumoniae clone leading to MDR and also suggests specific survival strategies in the bacteria that acquired MDR by the genome evolution. IMPORTANCE Within-host evolution is a survival strategy that can occur in many pathogens and is often associated with the emergence of novel antimicrobial-resistant (AMR) bacteria. To analyze this process, suitable sets of clinical isolates are required. Here, we analyzed five Klebsiella pneumoniae isolates which were consecutively isolated from a patient and showed a gradual increase in the AMR level. By genome sequencing and other analyses, we show that the first isolate was the antecedent of the later isolates and that they gained increased levels of antimicrobial resistance leading to multidrug resistance (MDR) by stepwise changes in the expression of outer membrane proteins. The isolates showing higher levels of MDR lost major porins but still colonized the patient’s gut, suggesting that the deleterious effects of porin loss were compensated for by the mutations in hexuronate metabolism-related genes and atpG, which were commonly detected in the MDR isolates.https://journals.asm.org/doi/10.1128/mSphere.00734-21Klebsiellacarbapenemsdrug resistance evolutionmultidrug resistance
spellingShingle Mai Yoshino
Masamune Aihara
Yasuhiro Gotoh
Masaru Akimoto
Wakana Tatsuhara
Makiko Kiyosuke
Yuichi Matsushima
Takeshi Uchiumi
Tetsuya Hayashi
Dongchon Kang
Stepwise Evolution of a Klebsiella pneumoniae Clone within a Host Leading to Increased Multidrug Resistance
mSphere
Klebsiella
carbapenems
drug resistance evolution
multidrug resistance
title Stepwise Evolution of a Klebsiella pneumoniae Clone within a Host Leading to Increased Multidrug Resistance
title_full Stepwise Evolution of a Klebsiella pneumoniae Clone within a Host Leading to Increased Multidrug Resistance
title_fullStr Stepwise Evolution of a Klebsiella pneumoniae Clone within a Host Leading to Increased Multidrug Resistance
title_full_unstemmed Stepwise Evolution of a Klebsiella pneumoniae Clone within a Host Leading to Increased Multidrug Resistance
title_short Stepwise Evolution of a Klebsiella pneumoniae Clone within a Host Leading to Increased Multidrug Resistance
title_sort stepwise evolution of a klebsiella pneumoniae clone within a host leading to increased multidrug resistance
topic Klebsiella
carbapenems
drug resistance evolution
multidrug resistance
url https://journals.asm.org/doi/10.1128/mSphere.00734-21
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