SD Ospedaliera di Microbiologia, Azienda Ospedaliero-Universitaria Pisana, Pisa, Italy.
University of Groningen, University Medical Center Groningen, Department of Medical Microbiology, Groningen, Netherlands.
mSphere. 2019 Nov 6;4(6):e00551-19. doi: 10.1128/mSphere.00551-19.
In the present study, we provide the results of a detailed genomic analysis and the growth characteristics of a colistin-resistant KPC-3-producing sequence type 512 (ST512) isolate (the colR-KPC3-KP isolate) with a mutated and isogenic isolates of colR-KPC3-KP with isolated from an immunocompromised patient. From 2014 to 2017, four colR-KPC3-KP isolates were detected in rectal swab samples collected from a pediatric hematology patient at the Azienda Ospedaliero-Universitaria Pisana in Pisa, Italy. Whole-genome sequencing was performed by MiSeq sequencing (Illumina). Growth experiments were performed using different concentrations of colistin. The growth lag phases both of an isolate harboring a deletion in and of clonal variants with were assessed by the use of real-time light-scattering measurements. In the first isolate (isolate 1000-Δ, recovered in September 2014), a 17-nucleotide deletion in was detected. In subsequent isolates, the gene associated with the plasmid pIncX4-AOUP was found, while was intact. Additionally, plasmid pIncQ-AOUP, harboring aminoglycoside resistance genes, was detected. The growth curves of the first three isolates were identical without colistin exposure; however, at higher concentrations of colistin, the growth curves of the isolate with a deletion in showed longer lag phases. We observed the replacement of mutated colR-KPC3-KP by isogenic isolates with multiple resistance plasmids, including carrying pIncX4, probably due to coselection under gentamicin treatment in a patient with prolonged colR-KPC3-KP carriage. The carriage of these isolates persisted in follow-up cultures. Coselection and the advantages in growth characteristics suggest that the plasmid-mediated resistance conferred by has fewer fitness costs in colR-KPC3-KP than mutations in chromosomal , contributing to the success of this highly resistant hospital-adapted epidemiological lineage. Our study shows a successful prolonged human colonization by a colistin-resistant isolate harboring An intense antibiotic therapy contributed to the maintenance of this microorganism through the acquisition of new resistance genes. The isolates carrying showed fewer fitness costs than isogenic isolates with a mutation in the chromosome. Coselection and reduced fitness costs may explain the replacement of isolates with the mutation by other isolates and the ability of the microorganism to persist despite antibiotic treatment.
在本研究中，我们提供了详细的基因组分析结果和对耐粘菌素 KPC-3 产生的序列型 512（ST512）分离株（colR-KPC3-KP 分离株）的生长特征的研究，该分离株携带突变的 和具有相同遗传特性的分离株，这些分离株是从一名免疫功能低下的患者中分离出来的。从 2014 年到 2017 年，在意大利比萨皮萨大学附属医院，从一名儿科血液病患者的直肠拭子样本中检测到了 4 株耐 colR-KPC3-KP 分离株。使用 MiSeq 测序（Illumina）进行全基因组测序。使用不同浓度的粘菌素进行生长实验。通过实时光散射测量评估携带缺失 和具有相同遗传特性的克隆变体的分离株的生长迟滞期。在第一个分离株（2014 年 9 月分离的 1000-Δ 分离株）中，检测到 中的 17 个核苷酸缺失。在随后的分离株中，发现与质粒 pIncX4-AOUP 相关的 基因，而 是完整的。此外，还检测到携带氨基糖苷类耐药基因的质粒 pIncQ-AOUP。在没有粘菌素暴露的情况下，前三个分离株的生长曲线是相同的；然而，在更高浓度的粘菌素下，缺失 基因的分离株的生长曲线表现出更长的迟滞期。我们观察到突变的 colR-KPC3-KP 通过携带多种耐药质粒的同源分离株的取代，包括携带 pIncX4，这可能是由于在携带时间延长的患者中使用庆大霉素治疗时的共选择。在后续培养中，这些分离株的携带情况仍然存在。共选择和生长特性的优势表明，与染色体 中的突变相比， 介导的耐药性在 colR-KPC3-KP 中具有更少的适应度代价，这有助于这种高度耐药的医院适应的流行病学谱系的成功。我们的研究表明，携带 的耐粘菌素 分离株在人类中长期定植是成功的，通过获得新的耐药基因，强烈的抗生素治疗有助于维持这种微生物。携带 的分离株比染色体上具有 突变的同源分离株具有更少的适应度代价。共选择和降低的适应度代价可能解释了为什么携带 突变的分离株被其他分离株所取代，以及为什么尽管进行了抗生素治疗，该微生物仍能持续存在。
