Prevalence of β-lactam antibiotic resistance of Escherichia coli isolated from a neonatal intensive care unit

Prevalence of β-lactam antibiotic resistance of Escherichia coli isolated from a neonatal intensive care unit | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article Prevalence of β-lactam antibiotic resistance of Escherichia coli isolated from a neonatal intensive care unit Jian Zhou, Jingqian Zhou, Min Chen, Pan Lü, ChunMing Jiang This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-5116757/v1 This work is licensed under a CC BY 4.0 License Status: Published Journal Publication published 04 Feb, 2025 Read the published version in BMC Pediatrics → Version 1 posted 4 You are reading this latest preprint version Abstract Introduction: Escherichia coli ( E. coli ) causes infections in neonates admitted to neonatal intensive care units (NICUs). Although β-lactam antibiotics are commonly used for neonatal infectious diseases, E. coli has exhibited resistance to them. Therefore, we investigated the resistance of E. coli strains isolated from a NICU to β-lactam antibiotics. Methods: E. coli isolates were collected from patients admitted to a NICU from 2020–2023. The clinical characteristics of the patients were analyzed. The antimicrobial susceptibility was determined using the agar dilution method, and the distribution of β-lactamase genes was analyzed using PCR. Conjugation experiments were conducted to analyze the horizontal transferability of resistance genes on plasmids. Genomic DNA was extracted for whole genome sequencing, construction of plasmid physical maps, locating resistance genes, and analyzing flanking regions and the resistance gene-related sequences. Results: Throughout the study period, 110 distinct E. coli strains were collected. Among these, 62 cases presented strains with high minimum inhibitory concentrations (MIC) associated with conditions such as ventilator-associated pneumonia (35/62), catheter-associated urinary tract infection (14/62), necrotizing enterocolitis (7/62), skin infection (1/62), and neonatal septicemia (5/62). Resistance of E. coli i solates to seven β-lactam antibiotics ranged from 2.73–56.36%. In 62 strains (56.36%, 62/110), six genotypes (11 sub-genotypes) of 111 β-lactamase genes were identified. Conjugation experiments revealed two transconjugants carrying the bla KPC-2 gene and two carrying the bla OXA-1 gene, exhibiting resistance to carbapenems and other β-lactams. The plasmids of four strains were successfully conjugated and transferred to recipient E. coli C600. PCR of the transconjugant resistance genes revealed that two carried a bla KPC-2 gene with a MIC increased up to 32-fold relative to the recipients, and the other two carried a bla OXA-1 gene with a 32-fold increased MIC. For isolate ECK03 carrying bla KPC-2 , bla CTX-M-64 , bla CTX-M-65 , and bla TEM-1 , sequencing results showed that bla KPC-2 , bla CTX-M-64 , and bla TEM-1 were harbored on a 114-kb pECK03_KPC-2 plasmid, whereas two identical bla CTX-M-64 genes were harbored in E. coli isolate ECF13. Conclusion: These findings highlight the existence of E. coli β-lactam resistance within NICU populations, emphasizing the need for continual monitoring of β-lactamase isolates to facilitate effective antibiotic selection. Escherichia coli resistance antimicrobial susceptibility test β-lactamase gene neonatal ICU Figures Figure 1 Figure 2 Figure 3 Figure 4 1 Introduction Escherichia coli , a common inhabitant of human and animal intestines, is typically innocuous to the human body. However, a compromised immune system coupled with prolonged stimulation of the gastrointestinal tract can lead to extraintestinal infections, potentially resulting in severe acute abdomen and sepsis ( 1 ). Patients in the neonatal intensive care unit (NICU) have distinct characteristics, such as underdeveloped overall immune systems, relatively poor resistance, and weak adaptability to external environments. These factors increase the vulnerability of neonates to E. coli infections, thereby enhancing antibiotic resistance ( 2 ). Reports of cephalosporin resistance, including extended-spectrum β-lactamase (ESBL) and/or carbapenem resistance, are emerging in NICUs and other healthcare settings ( 3 ). E. coli demonstrates resistance to almost all classes, with heightened resistance observed in β-lactams, aminoglycosides, and fluoroquinolones. However, β-lactam antibiotics are commonly used for neonatal infectious diseases. A primary mechanism contributing to antibiotic resistance in neonatal E. coli infections is the production of β-lactamases ( 4 ). The expression of β-lactamases, including ESBLs, ampC enzymes, and carbapenem-hydrolyzing β-lactamases, is a prevalent and crucial mechanism employed by antibiotic-resistant organisms to counteract β-lactams ( 5 ). The AmpC beta-lactamase gene in E. coli can be classified as chromosome-associated (cAmpC) or plasmid-associated (pAmpC), enhancing resistance to penicillin and cephalosporin ( 6 ). Notable ampC types include CMY-, FOX-, DHA-, and MIR-type β-lactamases ( 7 ). In clinical settings, ESBL-β-lactamase strains of E. coli are relevant in clinical settings for β-lactams, including the third- and fourth-generation cephalosporins. The sulfhydryl reagent variable (SHV) family of class A β-lactamases and the TEM (Temoneira) type β-lactamases, belonging to ESBLs, were the first reported instances of such enzymes ( 8 ). Plasmid-encoded AmpC types of β-lactamases, with CMY-2 and DHA-1 being prevalent, have gained global prominence in medical clinical practice (9). Recently, carbapenemases, notably the VIM-, NDM-, IMP-, OXA-, and GES-types, have become a global concern in clinical medicine ( 10 ). Notably, the dissemination of β-lactamases, particularly ESBL genes, among E. coli strains is mainly driven by horizontal transmission, with ESBL genes commonly associated with insertion sequences (ISs), transposons, and integrons ( 11 ). In this study, we investigated the prevalence of β-lactam antibiotics and the characteristics of resistance gene-related sequences in E. coli from the NICU. We also conducted clinical and molecular epidemiological analyses of β-lactamase-harboring E. coli in the NICU of a county hospital. 2 Materials and methods 2.1 Bacterial strains and relevant patient demographics Non-duplicate samples of clinically isolated Pseudomonas aeruginosa were collected from the NICU of the First People's Hospital of Yongkang, Zhejiang, China, between 2020 and 2023. The bacteria were purified using the Vitek-60 Microorganism Auto Analysis System (BioMérieux Corporate, Craponne, France). Bacterial species of all isolated bacteria were identified through biochemical identification and genome 16s rRNA sequencing methods, referred to as EC in isolate numbers of E. coli . Patient clinical records were collected and analyzed. Ethical approval was obtained from the First People's Hospital of Yongkang’s Institutional Ethics Committee (approval number: ykyy2020-13). We confirm that we have explained the detailed information of this study, including its rights, potential benefits, and risks, to all guardians of minors. All subject guardians have read and expressed their understanding of the contents of the informed consent form, and have voluntarily allowed their children to participate in this study. 2.2 Antimicrobial susceptibility test The antimicrobial susceptibility of E . coli was determined using the agar dilution method and interpreted according to the Clinical and Laboratory Standards Institute guidelines (CLSI, 2022). E. coli ATCC 25922 was used as the quality control strain. 2.3 DNA extraction and sequencing E. coli was cultured in Luria–Bertani (LB) broth placed in a bacterial incubator at 37 ℃ for 18–24 h, and genomic DNA was extracted using an AxyPrep Bacterial Genomic DNA Miniprep kit (Axygen Scientific, Union City, CA, USA). For mixed genomic DNA sequencing, cultured LB broth from every 10 bacterial strains was used, and then DNA was extracted. A library with an average insert size of 400 bp was prepared using the Next Generation Sequencing (NGS) DNA library preparation kit and sequenced using the Illumina NovaSeq platform (paired-end run; 2×150 bp). Additionally, for whole genome sequencing of a particular strain, a 10–20-kb insert library was prepared and sequenced using the Oxford Nanopore Technology (ONT) sequencer (Personalbio Technology Co., Ltd., Shanghai, China). 2.4 Genome assembly and annotation Genome assembly of the mixed DNA sequencing data was performed using Megahit (https://github.com/voutcn/megahit). The complete genome of an E. coli isolate was assembled using Unicycle and Flye software to obtain the contig sequence. Pilon software (https://github.com/broadinstitute/pilon) was used to correct the third-generation contig results and the final concatenation with the second-generation data from Illumina sequencing. Using the antibiotic resistance genes of the CARD (http://arpcard.mcmaster.ca) to query sequences, a BLASTN search was performed against the assembled sequences of the DNA with a parameter E-value of 1e-6, consistency of amino acid sequences above 45%, thresholds of >70% nucleotide identity, and >80% alignment coverage. 2.5 Screening of the β-lactamase resistance determinants β-lactamase resistance genes, including Amber class A ( bla KPC-2 , bla SHV-28 , bla CTX-M-14 , bla CTX-M-15 , bla CTX-M-55 , bla CTX-M-64 , bla CTX-M-65 , bla TEM-1, and bla TEM-244 ), class C ( bla DHA-1 ), and class D ( bla OXA-1 ), were screened using polymerase chain reaction (PCR). The primers were designed using Primer Premier, compared with that in corresponding publications, and synthesized by Personalbio Technology Co., Ltd. (Shanghai, China) (Table 1). The PCR products were verified by sequencing and compared using BLASTN (http://blast.ncbi.nlm.nih.gov/Blast.cgi). Table 1. Primers used in this study. Primer (restriction endonuclease) Primer sequence (5′→ 3′) PCR product size (bp) Tm (℃) IMP-45-F ATGTTTTTGTTTTGTAGCATTACTG 714 60 IMP-45-R TTAATGTGCAGTGGTACTTTTTTTG KPC-2-F ATGTCACTGTATCGCCGTC 953 60 KPC-2-R CTCAGTGCTCTACAGAAAACC OXA-1-F GGCACCAGATTCAACTTTCAAG 564 58 OXA-1-R GACCCCAAGTTTCCTGTAAGTG SHV-28-F CCGATGAACGCTTTCCCATG 615 57 SHV-28-R GCGTATCCCGCAGATATATCAC CTX-M-14-F GAGTGTTGCTCTGTGGATAAC 741 60 CTX-M-14-R GTTACAGCCCTTCGGCGATG CTX-M-15-F CGTCTCTTCCAGAATAAGG 905 60 CTX-M-15-R GTTTCCCCATTCCGTTTCCGC CTX-M-55-F CAAAGAGAGTGCAACGGATG 695 60 CTX-M-55-R ATTGGAAAGCGTTCATCACC CTX-M-64-F AACTTGCCGAATTAGAGCGG 1055 56 CTX-M-64-R GTTGCTCTGTGGATAACTTGC CTX-M-65-F GCAACGGATGATGTTCGCG 832 58 CTX-M-65-R GCGGCTGGGTAAAATAGGTC TEM-1-F AATGATACCGCGAGACCCAC 716 58 TEM-1-R AATGATACCGCGAGACCCAC TEM-244-F GATGCTGAAGATCAGTTGGGTG 745 58 TEM-244-R GGCACCTATCTCAGCGATCTGTC DHA-1-F GGTAAAACTGAGATGACGGGC 1422 58 DHA-1-R CTCATCCTCCATAAAACAGCCC Underlined sequences are restriction endonuclease sites. 2.6 Plasmid conjugation experiments Multidrug-resistant E. coli was used as the donor strain, and rifampicin-resistant E. coli EC600 was used as the recipient strain for the conjugation experiment of biparental mating. The transconjugants were selected from Mueller–Hinton agar plates containing 600 μg/mL rifampin and 32 μg/mL cefoperazone/sulbactam or 8 μg/mL imipenem. Target resistance genes in the transformants and in the plasmid of transformant cells were verified using PCR, and the product was then sequenced. 2.7 Comparative genomic analysis of the β-lactamase gene-related sequences For the comparative genomic analysis of bla CTX-M-64 and bla KPC-2 gene-related fragments, sequences containing these bla genes were obtained from the NCBI nucleotide database using the bla CTX-M-64 gene (NG_049015) or bla KPC-2 gene (NG_049253) as a query for a BLASTN search. The results were retained only if the sequences contained complete bla CTX-M-64 (20 kb, approximately 12 kb upstream and 8 kb downstream of the gene) or bla KPC-2 (30 kb, approximately 21 kb upstream and 9 kb downstream of the gene). Multiple sequence alignments were performed by MAFFT24 (https://mafft.cbrc.jp/alignment/software/) using each of the bla gene-related fragments of this work as a reference, and the sequences were clustered with an identity of ≥80%. The sequence sharing the highest similarity with the other sequences in each cluster was chosen as the candidate for ortholog analysis. Orthologous groups of genes from the candidate sequences were identified using BLASTP and InParanoid (https://inparanoid.sbc.su.se/). 3 Results 3.1 Demographic and clinical characteristics A total of 110 E. coli isolates were obtained from 110 patients (67 preterm and 43 full-term neonates) in the NICU. Most of the patients were male (55.3%), with a mean age of 16.4 ± 11.9 days. Identified diseases included 41 cases of ventilator-associated pneumonia, 22 cases of necrotizing enterocolitis, three cases of skin infection, five cases of purulent meningitis, 20 cases of catheter-associated urinary tract infection, and 19 cases of neonatal septicemia. Clinical manifestations in infected newborns included fever in 67 cases, with an average peak of 38.6 ± 1.38 ℃ and an average duration of 5.6 days. Inflammatory indicators of infected children included peripheral blood white cell count (13.78±7.56 × 10 9 /L), C- reactive protein level (48.5 ± 20.2 mg/L), and hypoallergenic (2.4 ±0.8 ng/ml). 3.2 Antimicrobial susceptibility The minimum inhibitory concentration (MIC) of seven antibiotics (piperacillin-tazobactam, ceftazidime, cefepime, ceftriaxone, cefoperazone/sulbactam, imipenem, and ertapenem) was determined for the 110 E. coli isolates, revealing resistance to all seven β-lactams. Resistance rates ranged from 2.7% (imipenem) to 54.5% (ceftriaxone). High-MIC isolates were associated with ventilator-associated pneumonia (35 cases), catheter-associated urinary tract infection (14 cases), necrotizing enterocolitis (seven cases), skin infection (one case), and neonatal septicemia (five cases). Two strains from ventilator-associated pneumonia and one strain from catheter-associated urinary tract infection showed resistance to all β-lactams, including imipenem and ertapenem. Additionally, two strains of ventilator-associated pneumonia, two strains of catheter-associated urinary tract infection, and one strain of neonatal septicemia were resistant to cefoperazone/sulbactam (Table 2). Table 2. Antimicrobial susceptibility of the 110 E. coli isolates against seven antimicrobials tested Antimicrobial S (%) I (%) R (%) MIC50 (μg/mL) MIC90 (μg/mL) Range (μg/mL) Piperacillin-Tazobactam 88.16 3.64 (4) 8.20 (9/110) 0.5 16 0.5–128 Ceftazidime 57.28 17.27 (19) 25.45 (28) 4 16 0.5–64 Cefepime 46.37 25.45 (28) 28.18 (31) 1 16 0.25–64 Ceftriaxone 19.09 24.55 (27) 56.36 (62) 4 16 1–256 Ertapenem 92.72 4.55 (5) 2.73 (3) 0.125 8 0.0625–64 Imipenem 90.00 7.27 (8) 2.73 0.125 8 0.0625–64 Cefoperazone/sulbactam 83.64 10.00 (11) 6.36 (7) 1 8 0.0625–64 3.3 Distribution of β-lactamases A total of 111 resistance genes of six genotypes were identified in 62 strains from 110 E. coli isolates through bla gene screening. Bla CTX-M showed the highest positivity rate of 50.45% (56/111), which consisted of five sub-genotypes: bla CTX-M-14 (17.12%, 19/111), bla CTX-M-15 (13.51%, 15/111), bla CTX-M-55 (9.91%, 11/111), bla CTX-M-64 (5.41%, 6/111), and bla CTX-M-65 (4.50%, 5/111). Bla TEM was second (36.04%, 40/111), followed by bla TEM-1 (21.62%, 24/111), bla TEM-244 (14.41%, 16/111), and bla SHV-28 (7.21%, 8/111). The remaining three ( bla KPC-2 , bla OXA-1 , and bla DHA-1 ) showed lower positivity rates of 2.70% (3/111), 1.80% (2/111), and 1.80% (2/111), respectively. Approximately half the 62 β-lactamase gene- harboring isolates (54.84%, 24/62) carried two resistance genes, eight isolates carried three resistance genes, three isolates carried four resistance genes, and the remaining 27 carried one gene (Table 3). These 62 isolates revealed higher MICs to β-lactams. All strains had MIC values ≥ 32 μg/mL for ceftriaxone, with 50.00% (31/62) and 45.16% (28/62) had MIC levels ≥ 16 μg/mL for cefepime and ceftazidime. Only three strains (4.84%, 3/62) had MIC levels ≥ 8 μg/mL for ertapenem or imipenem (Table 3). None of the β-lactamase gene-negative strains showed resistance to the evaluated β-lactams. Table 3. MIC results of the isolates harboring three or four β-lactamase gene and transconjugants Isolate MIC (μg/ml) of Antimicrobial agent Resistance mechanism No. ceftazidime cefepime ertapenem imipenem Ceftriaxone piperacillin-tazobactam cefoperazone/sulbactam ECF13 64 64 0.25 0.5 64 64 64 bla CTX-M-64 bla CTX-M-64 bla CTX-M-14 bla TEM-1 ECK46 32 64 1 2 128 64 2 bla CTX-M-14 bla CTX-M-15 bla SHV-28 bla TEM-1 ECK42 64 64 32 32 64 32 32 bla CTX-M-64 bla KPC-2 bla TEM-244 ECK55 32 32 1 1 64 64 1 bla TEM-1 bla CTX-M-15 bla SHV-28 ECK67 32 32 32 32 128 128 1 bla CTX-M-55 bla KPC-2 bla CTX-M-64 ECK03 64 64 64 64 128 128 32 bla KPC-2 bla CTX-M-64 bla TEM-1 bla CTX-M-14 ECK01 64 32 2 4 32 128 32 bla CTX-M-55 bla TEM-244 bla SHV-28 ECK69 64 64 1 2 64 64 16 bla TEM-244 bla OXA-1 bla DHA-1 ECF04 32 32 4 8 128 128 32 bla CTX-M-65 bla TEM-1 bla DHA-1 ECF09 64 64 8 8 64 4 32 bla TEM-244 bla CTX-M-55 bla OXA-1 ECF81 32 32 0.25 0.5 64 8 1 bla CTX-M-65 bla TEM-244 bla CTX-M-15 ECF04/EC600 32 32 4 4 32 64 16 bla CTX-M-65 ECK03/EC600 64 32 64 64 128 128 32 bla KPC-2 bla CTX-M-64 bla TEM-1 ECK55/EC600 32 32 8 1 32 32 1 bla CTX-M-15 bla SHV-28 ECF09/EC600 32 32 8 32 32 8 16 bla CTX-M-55 -45 bla OXA-1 3.4 Resistant plasmid transferability Of the five strains resistant to the β-lactams, the conjugative plasmids of four strains were successfully transferred to E. coli EC600. PCR amplification revealed that two of the transconjugants carried a bla KPC-2 gene (including one co-expressing bla CTX-M-64 and bla TEM-1 genes), and the other two carried a bla OXA-1 gene. The four transconjugants with bla OXA-1 exhibited reduced susceptibility to carbapenems with MIC values up to 32-fold compared with that in the recipients. The two bla KPC-2 harboring transconjugants showed MIC values up to 64-fold compared with that in recipients against carbapenems (Table 3). The remaining strain harboring bla KPC-2 failed the conjugation experiments. 3.5 General features of the E. Coli genomes Whole genome sequencing and further molecular analysis were performed for two strains carrying different β-lactamases. The two isolates included E. coli ECK03 carrying bla KPC-2 , bla CTX-M-64 , bla CTX-M-65 , and bla TEM-1 , and E. coli ECF13 carrying bla CTX-M-14, bla TEM-1 , and two genes of bla CTX-M-64 . Sequencing results showed that only ECK03 carried one resistance plasmid. The bla KPC-2 of E. coli ECK03 was encoded in a plasmid (pECK03_KPC-2), while the two genes of bla CTX-M-64 of E. coli ECF13 were encoded on the chromosomes. ECK03 harbored four plasmids, a 114 kb plasmid named pECK03_KPC-2 harbored bla KPC-2 , bla CTX-M-64 , and bla TEM-1 , and another three plasmids free of the resistance gene). ECF13 contained two plasmids free of resistance genes. 3.6 Comparative genomic analysis of sequences harbored bla KPC-2 and bla CTX-M-64 Comparative analysis of sequence data from the NCBI database revealed four plasmids sharing maximum similarities with pECK03_KPC-2 in two E. coli species: p116753-KPC ( K. pneumoniae , MN891682.1), pKP19-3138-4 ( K. pneumoniae , CP090620.1), pOW1E2a ( E. coli , CP067246.1), and pKPC2_090374 ( K. pneumoniae , CP066536.1) (Figure 1). Plasmid pECK03_KPC-2 harbored three bla resistance genes: bla KPC-2 , bla CTX-M-64 , and bla TEM-1 (Figure 2). Two genes, bla CTX-M-64 from E. coli ECF13 with bla CTX-M-14, bla TEM-1 , and two bla CTX-M-64 genes, were found on the chromosomes. The sequence of the first bla CTX-M-64 possessed an identical gene environment to that of clone CP104274 of E. coli EC15103 containing the bla CTX-M-64 gene. However, the sequence of the second bla CTX-M-64 was not identified in the sequences of the highest similarity gene environment containing bla CTX-M-64 (Figures 3 and 4). 4 Discussion In this study, all isolates were from the NICU, and 62 of the 110 clinical isolates had high MICs. Due to the limitations of antibiotic use in pediatric patients, especially neonates, only β-lactam antibiotics are used against gram-negative bacteria. Therefore, the high drug resistance rate poses significant challenges for selecting antibiotics in clinical practice. In addition, 49 of the 62 (79.03%) isolates were from ventilator-associated pneumonia and catheter-associated urinary tract infections and were believed to be hospital-acquired infections. From the high MIC isolates, nine bla genotypes (11 sub-genotypes) of Ambler classes were identified, including class A ( bla KPC−2, bla SHV−28, bla CTX−M−14, bla CTX−M−15, bla CTX−M−55, bla CTX−M−64, bla CTX−M−65 , bla TEM−1, and bla TEM−244 ), class C ( bla DHA−1 ), and class D ( bla OXA−1 ), of which Ambler class A was the most prevalent with a total of 57 strains (91.94%, 57/62). In this study, bla CTX−M showed the highest positive rate of 50.45% (56/111), and bla TEM was second (36.04%, 40/111). These results suggest that all β-lactam antibiotics except carbapenem are prone to high resistance ( 12 ). As an ESBL variant, CTX-M β-lactamases are prevalent and widespread worldwide ( 13 ). In this study, three isolates from patients with ventilator-associated pneumonia (2.73%, 3/110) carrying bla KPC−2 revealed higher MIC values to ertapenem and imipenem (both 64 µg/mL). As a less frequent but much more powerful carbapenem-hydrolyzing enzyme in E. coli , KPC-2 belongs to the class A β-lactamase, and was frequently reported in Klebsiella pneumoniae samples from NICUs ( 14 ) but was rarely reported in NICU E. coli strains. In addition, the two strains carrying bla OXA−1 showed weak carbapenem resistance. Despite the presence of genes that are resistant to carbapenems, the resistance rate was not higher than that reported in other studies ( 15 ). Eight of the 62 isolates carried three resistance genes, and three strains carried four resistance genes. The resistance gene combinations of each strain were mainly composed of bla CTX−M and bla TEM, and often, two bla CTX−MS or two bla TEM s were present, and two identical bla CTX−M or bla TEM genes existed in individual strains, such as bla CTX−M−64 . No similar β-lactamase gene combinations co-occurring with identical bla CTX−M or bla TEM genes have been reported. In addition, bla KPC−2 and bla OXA−1 are all present in these multiple β-lactamase isolates. The co-occurrence of β-lactamase genes displays high rates of resistance to broad-spectrum cephalosporins owing to induced expression of the β-lactamase gene; an increasing number of organisms harboring plasmid-encoded ESBL and/or carbapenemase genes have been reported ( 16 ). The most common is the co-occurrence of two different β-lactamase genes; however, the co-occurrence of three or more β-lactamase genes is rare ( 17 ), with no report from NICUs. Whole genome sequencing of the isolate ECK03 displayed that the β-lactamase genes were related to mobile genetic elements and were mainly harbored on plasmids. The resistance genes of pECK03_KPC-2 clustered mainly in the multidrug-resistant (MDR) region. The MDR regions of pECK03_KPC-2 carrying bla KPC−2 , bla CTX−M−64, and bla TEM−1 were similar to those of plasmid MN891682 (19). Compared with MN891682 and other plasmids with similar MDR regions, the MDR region of pECK03_KPC-2 contained many more massive insertions of foreign resistance genetic contents and showed a higher degree of genomic plasticity. Two identical bla CTX−M−64 genes in ECF13 were located on different chromosomes and contained other resistance genes, such as bla CTX−M−14 and bla TEM−1 . This rare combination of MDR genes demonstrated stronger resistance than the other isolates. In conclusion, the result reveals a high ratio of β-lactam antibiotic resistance in E. coli strains from NICUs. Notably, among the 62 of 110 isolates that carried β-lactamase genes, bla CTX−M and bla TEM were the most common, and co-occurrence of two or more β-lactamase genes was prevalent. The resistance gene-carrying plasmid may be transferred, and the harbored β-lactamase gene maintains resistance to β-lactams in recipients. In contrast, the three co-carrying bla KPC−2 also exhibited reduced susceptibility to carbapenems. The identification of high β-lactam antibiotic resistance in NICU E. coli strains highlights the urgent need for targeted antibiotic stewardship and infection control measures in neonatal healthcare settings. Clinicians should carefully consider alternative treatment options, given the observed prevalence of specific resistance genes, such as blaCTX-M and blaTEM, to ensure effective management of E. coli infections in this vulnerable patient population. Declarations Conflict of interest The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. Ethics approval This study was approved by the Institutional Ethics Committee of the First People's Hospital of Yongkang, Zhejiang, China (approval number: ykyy2020-87). Informed consent was obtained from all participants involved in the study. Clinical Trial Registration Not applicable. Consent for publication Not applicable. Availability of data and material The datasets used and/or analysed during the current study are available from the corresponding author on reasonable request. Funding This work was supported by Medical and Health Science and Technology Project of Zhejiang Province (CN, no. 2020ZH068). Author Contribution J.Z was responsible for the study's conceptualization, methodology, project administration, formal analysis, data analysis, and also contributed to the review and editing of the manuscript.JQ.Z and M.C both conducted the investigation and curated the data. P.L was involved in testing. CM.J provided supervision, wrote the original draft, and contributed to the manuscript's review, editing, and validation. All authors have read and consented to the published version of the manuscript. Acknowledgments The authors would like to thank the participants, who made this study possible. Data Availability Sequence data that support the findings of this study have been deposited in national center for biobechnology ;Accession: SAMN37977427 ID: 37977427. References Longhi C, Maurizi L, Conte AL, Marazzato M, Comanducci A, Nicoletti M et al. Extraintestinal pathogenic Escherichia coli: beta-lactam antibiotic and heavy metal resistance. Antibiotics (Basel) (2022) 11:328. doi: 10.3390/antibiotics11030328 Flannery DD, Akinboyo IC, Mukhopadhyay S, Tribble AC, Song L, Chen F et al. Antibiotic Susceptibility of Escherichia coli Among Infants Admitted to Neonatal Seidel J, Haller S, Eckmanns T, Harder T. Routine screening for colonization by Gram-negative bacteria in neonates at intensive care units for the prediction of sepsis: systematic review and meta-analysis. J Hosp Infect (2018) 99:367-80. doi: 10.1016/j.jhin.2018.03.017 Lee YQ, Ahmad Kamar A, Velayuthan RD, Chong CW, Teh CSJ. Clonal relatedness in the acquisition of intestinal carriage and transmission of multidrug resistant (MDR) Klebsiella pneumoniae and Escherichia coli and its risk factors among preterm infants admitted to the neonatal intensive care unit (NICU). Pediatr Neonatol (2021) 62:129-37. doi: 10.1016/j.pedneo.2020.10.002 Blair JM, Webber MA, Baylay AJ, Ogbolu DO, Piddock LJ. Molecular mechanisms of antibiotic resistance. Nat Rev Microbiol (2015) 13:42-51. doi: 10.1038/nrmicro3380 Meini S, Tascini C, Cei M, Sozio E, Rossolini GM. AmpC β-lactamase-producing Enterobacterales: what a clinician should know. Infection (2019) 47:363-75. doi: 10.1007/s15010-019-01291-9 Michel-Briand Y. Resistance to the latest beta-lactams: mechanisms of acquisition and spread of resistance in Enterobacteriaceae. Bull Acad Natl Med (2007) 191:35-50; discussion 50-1 Daoud Z, Salem Sokhn E, Masri K, Matar GM, Doron S. Escherichia coli isolated from urinary tract infections of Lebanese patients between 2005 and 2012: epidemiology and profiles of resistance. Front Med (Lausanne) (2015) 2:26. doi: 10.3389/fmed.2015.000269 Hayer SS, Casanova-Higes A, Paladino E, Elnekave E, Nault A, Johnson T et al. Global distribution of extended spectrum cephalosporin and carbapenem resistance and associated resistance markers in Escherichia coli of swine origin – A systematic review and meta-analysis. Front Microbiol (2022) 13:853810. doi: 10.3389/fmicb.2022.853810 Schäfer F, Görner P, Woltemate S, Brandenberger C, Geffers R, Ziesing S et al. The resistance mechanism governs physiological adaptation of Escherichia coli to growth with sublethal concentrations of carbapenem. Front Microbiol (2021) 12:812544. doi: 10.3389/fmicb.2021.812544 Wedel E, Bernabe-Balas C, Ares-Arroyo M, Montero N, Santos-Lopez A, Mazel D et al. Insertion sequences determine plasmid adaptation to new bacterial Hosts. mBio (2023) 14:e0315822. doi: 10.1128/mbio.03158-22 Oteo J, Cercenado E, Fernández-Romero S, Saéz D, Padilla B, Zamora E et al. Extended-spectrum-β-lactamase-producing Escherichia coli as a cause of pediatric infections: report of a neonatal intensive care unit outbreak due to a CTX-M-14-producing strain. Antimicrob Agents Chemother (2012) 56:54-8. doi: 10.1128/AAC.05103-11 Giedraitiene A, Pereckaite L, Bredelyte-Gruodiene E, Virgailis M, Ciapiene I, Tatarunas V. CTX-M-producing Escherichia coli strains: resistance to temocillin, fosfomycin, nitrofurantoin and biofilm formation.Future Microbiol (2022),17:789-802. doi: 10.2217/fmb-2021-0202. Maida CM, Bonura C, Geraci DM, Graziano G, Carattoli A, Rizzo A et al. Outbreak of ST395 KPC-producing Klebsiella pneumoniae in a neonatal intensive care unit in Palermo, Italy. Infect Control Hosp Epidemiol (2018) 39:496-8. doi: 10.1017/ice.2017.267 Xu Q, Pan F, Sun Y, Wang C, Shi Y, Zhang T et al. Fecal carriage and molecular epidemiology of carbapenem-resistant Enterobacteriaceae from inpatient children in a pediatric hospital of shanghai. Infect Drug Resist (2020) 13:4405-15. doi: 10.2147/IDR.S275549 Zorgani A, Daw H, Sufya N, Bashein A, Elahmer O, Chouchani C. Co-occurrence of plasmid-mediated AmpC β-lactamase activity among Klebsiella pneumoniae and Escherichia coli. Open Microbiol J (2017) 11:195-202. doi: 10.2174/1874285801711010195 Ramoul A, Loucif L, Bakour S, Amiri S, Dekhil M, Rolain JM. Co-occurrence of blaNDM-1 with blaOXA-23 or blaOXA-58 in clinical multidrug-resistant Acinetobacter baumannii isolates in Algeria. J Glob Antimicrob Resist (2016) 6:136-41. doi: 10.1016/j.jgar.2016.05.003 Jeong S, Kim JO, Yoon EJ, Bae IK, Lee W, Lee H et al. Extensively drug-resistant Escherichia coli sequence Type 1642 carrying an IncX3 plasmid containing the blaKPC-2 gene associated with transposon Tn4401a. Ann Lab Med (2018) 38:17-22. doi: 10.3343/alm.2018.38.1.17 Additional Declarations No competing interests reported. Cite Share Download PDF Status: Published Journal Publication published 04 Feb, 2025 Read the published version in BMC Pediatrics → Version 1 posted Editorial decision: Revision requested 30 Sep, 2024 Editor assigned by journal 28 Sep, 2024 Submission checks completed at journal 28 Sep, 2024 First submitted to journal 19 Sep, 2024 You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-5116757","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":360446994,"identity":"6fdc09de-bd78-49cc-be3c-6c38109c9ee0","order_by":0,"name":"Jian Zhou","email":"","orcid":"","institution":"Zhejiang University School of Medicine","correspondingAuthor":false,"prefix":"","firstName":"Jian","middleName":"","lastName":"Zhou","suffix":""},{"id":360446995,"identity":"f72d2675-1505-46a0-8160-d79456f69ebe","order_by":1,"name":"Jingqian Zhou","email":"","orcid":"","institution":"Zhejiang University School of Medicine","correspondingAuthor":false,"prefix":"","firstName":"Jingqian","middleName":"","lastName":"Zhou","suffix":""},{"id":360446996,"identity":"d36734b6-f2ac-4f39-a3a2-04a3b0f979df","order_by":2,"name":"Min Chen","email":"","orcid":"","institution":"The First People's Hospital of Yongkang","correspondingAuthor":false,"prefix":"","firstName":"Min","middleName":"","lastName":"Chen","suffix":""},{"id":360446997,"identity":"1b223e2d-d3de-4f3d-9b92-91d3d34b4a4d","order_by":3,"name":"Pan Lü","email":"","orcid":"","institution":"The First People's Hospital of Yongkang","correspondingAuthor":false,"prefix":"","firstName":"Pan","middleName":"","lastName":"Lü","suffix":""},{"id":360446998,"identity":"a177eedb-2a19-4f22-82ab-e3ce278acb3c","order_by":4,"name":"ChunMing Jiang","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAA9ElEQVRIiWNgGAWjYBACxmYIzcPGzHzwAQMPiJ1AnBYZPna2ZAOitMCAjRw/j5kEhE1AC3M787OHX/4cBjqMwazyh8xhBn72HAOGnzvwOYzN3Fi2Dawl7YYEz2EGyZ43Boy9Z/D6xUxasgGs5dgNA6AWgxs5BsyMbfi0sH+TlgA7jLGtIAGoxZ6wFh4zyQ9sIC3MbAwHQLZIENZSJs3Ylg7UwsYs2cCTziNx5lnBwV48Wgz7j2+T/PHH2l6+//zHjz97rOX425M3PviJT0sDMKB5GKARytgDicwDuDUwMMiDFP5gqINyf+BTOwpGwSgYBSMVAABh1kZxc5KpWgAAAABJRU5ErkJggg==","orcid":"","institution":"Hangzhou First People’s Hospital","correspondingAuthor":true,"prefix":"","firstName":"ChunMing","middleName":"","lastName":"Jiang","suffix":""}],"badges":[],"createdAt":"2024-09-19 11:54:41","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-5116757/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-5116757/v1","draftVersion":[],"editorialEvents":[{"content":"https://doi.org/10.1186/s12887-025-05389-y","type":"published","date":"2025-02-04T15:58:19+00:00"}],"editorialNote":"","failedWorkflow":false,"files":[{"id":71683679,"identity":"f0b51195-e204-4027-8eb5-d3131f479147","added_by":"auto","created_at":"2024-12-17 16:56:30","extension":"jpg","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":137182,"visible":true,"origin":"","legend":"\u003cp\u003eComplete sequence of the pECK03_KPC-2 plasmid and comparative genomic analysis of the pPA1609-475 plasmid sequence with other sequences. The circles (from innermost to outermost) represent (i) the scale in kb; (ii) the cumulative GC skew; (iii) the GC content; (iv) the annotated coding sequences with selected genes indicated according to the gene function: resistance genes in red arrows, transposase genes, IS elements in bottle-green arrows, and hypothetical proteins in dark gray arrows; and (v) circles (from inside to outside) representing three homologous plasmids (p116753-KPC, MN891682.1; pKP19-3138-4, CP090620.1; pOW1E2a, CP067246.1; and pKPC2_090374, CP066536.1).\u003c/p\u003e","description":"","filename":"Figure1.jpg","url":"https://assets-eu.researchsquare.com/files/rs-5116757/v1/a5d5af6a7cb733300be01bc3.jpg"},{"id":71683678,"identity":"b651abae-a2ca-458c-a683-d01dcd256233","added_by":"auto","created_at":"2024-12-17 16:56:30","extension":"jpg","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":1294929,"visible":true,"origin":"","legend":"\u003cp\u003eComparative genomic analysis of the resistance gene region of pECK03_KPC-2 with other sequences from different bacteria. The homologous gene clusters among plasmids pECK03_KPC-2; p116753-KPC, MN891682.1; pOW1E2a, CP067246.1; and 2019XSD11-92, MN101857.1, with the resistance gene clusters in bottle red. Annotated coding sequences are denoted using arrows. Coding sequences are colored based on their assigned gene functions.\u003c/p\u003e","description":"","filename":"figure2envKPC2.jpg","url":"https://assets-eu.researchsquare.com/files/rs-5116757/v1/78ea1420de6314543de096eb.jpg"},{"id":71682676,"identity":"387e590e-c302-42e5-b4ec-14b0f921ae41","added_by":"auto","created_at":"2024-12-17 16:48:30","extension":"jpg","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":1249351,"visible":true,"origin":"","legend":"\u003cp\u003eComparative genomic analysis of the resistance gene region of ECF13 of the first \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX-M-64 \u003c/sub\u003ewith other sequences from different bacteria. The homologous gene clusters among ECF13; Ec15103, CP104274.1; DETEC-P793 chromosome, CP116115.1; and DETEC-P836 chromosome, CP116103, with the resistance gene clusters in bottle red. Annotated coding sequences are denoted using arrows. Coding sequences are colored based on their assigned gene functions.\u003c/p\u003e","description":"","filename":"Figure3Ec13ChroCTXM1.jpg","url":"https://assets-eu.researchsquare.com/files/rs-5116757/v1/7c7dd145bfd26161ab86272a.jpg"},{"id":71682675,"identity":"52683076-bee1-4786-935f-116ddbfa0166","added_by":"auto","created_at":"2024-12-17 16:48:30","extension":"jpg","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":1021979,"visible":true,"origin":"","legend":"\u003cp\u003eComparative genomic analysis of the resistance gene region of ECF13 of the second \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX-M-64 \u003c/sub\u003ewith other sequences from different bacteria. The homologous gene clusters among ECF13; EcPF40 chromosome, CP054214.1; SCU-204 chromosome, CP053251.2; and 09-02E, AP022650, with the resistance gene clusters in bottle red. Annotated coding sequences are denoted using arrows. Coding sequences are colored based on their assigned gene functions.\u003c/p\u003e","description":"","filename":"figure4Ec13chro2CTXM2.jpg","url":"https://assets-eu.researchsquare.com/files/rs-5116757/v1/09769394ffc68bcf4e6653ea.jpg"},{"id":75930788,"identity":"7c3c1e5b-ac05-4b48-82b1-813448795251","added_by":"auto","created_at":"2025-02-10 16:13:22","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":4855881,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-5116757/v1/34d33f1b-def1-4e4e-bb9d-0803317c9b62.pdf"}],"financialInterests":"No competing interests reported.","formattedTitle":"Prevalence of β-lactam antibiotic resistance of Escherichia coli isolated from a neonatal intensive care unit","fulltext":[{"header":"1 Introduction","content":"\u003cp\u003e \u003cem\u003eEscherichia coli\u003c/em\u003e, a common inhabitant of human and animal intestines, is typically innocuous to the human body. However, a compromised immune system coupled with prolonged stimulation of the gastrointestinal tract can lead to extraintestinal infections, potentially resulting in severe acute abdomen and sepsis (\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e). Patients in the neonatal intensive care unit (NICU) have distinct characteristics, such as underdeveloped overall immune systems, relatively poor resistance, and weak adaptability to external environments. These factors increase the vulnerability of neonates to \u003cem\u003eE. coli\u003c/em\u003e infections, thereby enhancing antibiotic resistance (\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e). Reports of cephalosporin resistance, including extended-spectrum β-lactamase (ESBL) and/or carbapenem resistance, are emerging in NICUs and other healthcare settings (\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e).\u003c/p\u003e \u003cp\u003e \u003cem\u003eE. coli\u003c/em\u003e demonstrates resistance to almost all classes, with heightened resistance observed in β-lactams, aminoglycosides, and fluoroquinolones. However, β-lactam antibiotics are commonly used for neonatal infectious diseases. A primary mechanism contributing to antibiotic resistance in neonatal \u003cem\u003eE. coli\u003c/em\u003e infections is the production of β-lactamases (\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e). The expression of β-lactamases, including ESBLs, ampC enzymes, and carbapenem-hydrolyzing β-lactamases, is a prevalent and crucial mechanism employed by antibiotic-resistant organisms to counteract β-lactams (\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eThe AmpC beta-lactamase gene in \u003cem\u003eE. coli\u003c/em\u003e can be classified as chromosome-associated (cAmpC) or plasmid-associated (pAmpC), enhancing resistance to penicillin and cephalosporin (\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e). Notable ampC types include CMY-, FOX-, DHA-, and MIR-type β-lactamases (\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e). In clinical settings, ESBL-β-lactamase strains of \u003cem\u003eE. coli\u003c/em\u003e are relevant in clinical settings for β-lactams, including the third- and fourth-generation cephalosporins. The sulfhydryl reagent variable (SHV) family of class A β-lactamases and the TEM (Temoneira) type β-lactamases, belonging to ESBLs, were the first reported instances of such enzymes (\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e). Plasmid-encoded AmpC types of β-lactamases, with CMY-2 and DHA-1 being prevalent, have gained global prominence in medical clinical practice (9). Recently, carbapenemases, notably the VIM-, NDM-, IMP-, OXA-, and GES-types, have become a global concern in clinical medicine (\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e10\u003c/span\u003e). Notably, the dissemination of β-lactamases, particularly ESBL genes, among \u003cem\u003eE. coli\u003c/em\u003e strains is mainly driven by horizontal transmission, with ESBL genes commonly associated with insertion sequences (ISs), transposons, and integrons (\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e11\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eIn this study, we investigated the prevalence of β-lactam antibiotics and the characteristics of resistance gene-related sequences in \u003cem\u003eE. coli\u003c/em\u003e from the NICU. We also conducted clinical and molecular epidemiological analyses of β-lactamase-harboring \u003cem\u003eE. coli\u003c/em\u003e in the NICU of a county hospital.\u003c/p\u003e"},{"header":"2 Materials and methods","content":"\u003ch2\u003e2.1 Bacterial strains and relevant patient demographics\u003c/h2\u003e\n\u003cp\u003eNon-duplicate samples of clinically isolated \u003cem\u003ePseudomonas aeruginosa\u003c/em\u003e were collected from the NICU of the First People\u0026apos;s Hospital of Yongkang, Zhejiang, China, between 2020 and 2023. The bacteria were purified using the Vitek-60 Microorganism Auto Analysis System (BioM\u0026eacute;rieux Corporate, Craponne, France). Bacterial species of all isolated bacteria were identified through biochemical identification and genome 16s rRNA sequencing methods, referred to as EC in isolate numbers of \u003cem\u003eE. coli\u003c/em\u003e. Patient clinical records were collected and analyzed. Ethical approval was obtained from the First People\u0026apos;s Hospital of Yongkang\u0026rsquo;s Institutional Ethics Committee (approval number: ykyy2020-13). We confirm that we have explained the detailed information of this study, including its rights, potential benefits, and risks, to all guardians of minors. All subject guardians have read and expressed their understanding of the contents of the informed consent form, and have voluntarily allowed their children to participate in this study.\u003c/p\u003e\n\u003ch2\u003e2.2 Antimicrobial susceptibility test\u003c/h2\u003e\n\u003cp\u003eThe\u0026nbsp;antimicrobial susceptibility\u0026nbsp;of \u003cem\u003eE\u003c/em\u003e\u003cem\u003e.\u0026nbsp;\u003c/em\u003e\u003cem\u003ecoli\u003c/em\u003e was determined using the agar dilution method and interpreted according to the Clinical and Laboratory Standards Institute guidelines (CLSI, 2022). \u003cem\u003eE. coli\u003c/em\u003e ATCC 25922 was used as the quality control strain.\u003c/p\u003e\n\u003ch2\u003e2.3 DNA extraction and sequencing\u003c/h2\u003e\n\u003cp\u003e\u003cem\u003eE. coli\u003c/em\u003e was cultured in Luria\u0026ndash;Bertani (LB) broth\u0026nbsp;placed in a bacterial incubator\u0026nbsp;at 37 ℃ for 18\u0026ndash;24 h, and genomic DNA was extracted using an AxyPrep Bacterial Genomic DNA Miniprep kit (Axygen Scientific, Union City, CA, USA). For mixed genomic DNA sequencing, cultured LB broth from every 10 bacterial strains was used, and then DNA was extracted. A library with an average insert size of 400 bp was prepared using the Next Generation Sequencing (NGS) DNA library preparation kit and sequenced using the Illumina NovaSeq platform (paired-end run; 2\u0026times;150 bp). Additionally, for whole genome sequencing of a particular strain, a 10\u0026ndash;20-kb insert library was prepared and sequenced using the Oxford Nanopore Technology (ONT) sequencer (Personalbio Technology Co., Ltd., Shanghai, China).\u003c/p\u003e\n\u003ch2\u003e2.4 Genome assembly and annotation\u003c/h2\u003e\n\u003cp\u003eGenome assembly of the mixed DNA\u0026nbsp;sequencing data\u0026nbsp;was performed using Megahit\u0026nbsp;(https://github.com/voutcn/megahit).\u0026nbsp;The complete genome of an \u003cem\u003eE. coli\u003c/em\u003e isolate was assembled using Unicycle and Flye software to obtain the contig sequence. Pilon software\u0026nbsp;(https://github.com/broadinstitute/pilon)\u0026nbsp;was used to correct the third-generation contig results and the final concatenation with the second-generation data from Illumina sequencing. Using the antibiotic resistance genes of the CARD\u0026nbsp;(http://arpcard.mcmaster.ca)\u0026nbsp;to query sequences, a BLASTN search was performed against the assembled sequences of the DNA with a parameter E-value of 1e-6, consistency of amino acid sequences above 45%, thresholds of \u0026gt;70% nucleotide identity, and \u0026gt;80% alignment coverage.\u003c/p\u003e\n\u003ch2\u003e2.5 Screening of the \u0026beta;-lactamase resistance determinants\u003c/h2\u003e\n\u003cp\u003e\u0026beta;-lactamase\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003eresistance genes, including Amber class A (\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eKPC-2\u003cem\u003e,\u003c/em\u003e\u0026nbsp;\u003c/sub\u003e\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eSHV-28\u003cem\u003e,\u003c/em\u003e\u0026nbsp;\u003c/sub\u003e\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX-M-14\u003c/sub\u003e, \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX-M-15\u003cem\u003e,\u003c/em\u003e\u0026nbsp;\u003c/sub\u003e\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX-M-55\u003cem\u003e,\u0026nbsp;\u003c/em\u003e\u003c/sub\u003e\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX-M-64\u003cem\u003e,\u0026nbsp;\u003c/em\u003e\u003c/sub\u003e\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX-M-65\u003c/sub\u003e,\u003csub\u003e\u0026nbsp;\u003c/sub\u003e\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eTEM-1,\u0026nbsp;\u003c/sub\u003eand \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eTEM-244\u003c/sub\u003e), class C (\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eDHA-1\u003c/sub\u003e), and class D (\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eOXA-1\u003c/sub\u003e), were screened using polymerase chain reaction (PCR). The primers were designed using Primer Premier, compared with that in corresponding publications, and synthesized by Personalbio Technology Co., Ltd. (Shanghai, China) (Table 1). The PCR products were verified by sequencing and compared using BLASTN\u0026nbsp;(http://blast.ncbi.nlm.nih.gov/Blast.cgi).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eTable 1.\u003c/strong\u003e Primers used in this study.\u003c/p\u003e\n\u003ctable border=\"0\" cellspacing=\"0\" cellpadding=\"0\" align=\"\" width=\"579\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 129px;\"\u003e\u003cstrong\u003ePrimer (restriction endonuclease)\u003c/strong\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd style=\"width: 315px;\"\u003e\u003cstrong\u003ePrimer sequence (5\u0026prime;\u0026rarr; 3\u0026prime;)\u0026nbsp;\u003c/strong\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd style=\"width: 77px;\"\u003e\u003cstrong\u003ePCR product size (bp)\u003c/strong\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd style=\"width: 57px;\"\u003e\u003cstrong\u003eTm (℃)\u003c/strong\u003e\u003cbr\u003e\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 129px;\"\u003eIMP-45-F\u0026nbsp;\u003cbr\u003e\u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 315px;\"\u003eATGTTTTTGTTTTGTAGCATTACTG\u003cbr\u003e\u003c/td\u003e\n \u003ctd rowspan=\"2\" style=\"width: 77px;\"\u003e714\u003cbr\u003e\u003c/td\u003e\n \u003ctd rowspan=\"2\" style=\"width: 57px;\"\u003e60\u003cbr\u003e\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 129px;\"\u003eIMP-45-R\u0026nbsp;\u003cbr\u003e\u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 315px;\"\u003eTTAATGTGCAGTGGTACTTTTTTTG\u003cbr\u003e\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 129px;\"\u003eKPC-2-F\u0026nbsp;\u003cbr\u003e\u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 315px;\"\u003eATGTCACTGTATCGCCGTC\u003cbr\u003e\u003c/td\u003e\n \u003ctd rowspan=\"2\" style=\"width: 77px;\"\u003e953\u003cbr\u003e\u003c/td\u003e\n \u003ctd rowspan=\"2\" style=\"width: 57px;\"\u003e60\u003cbr\u003e\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 129px;\"\u003eKPC-2-R\u0026nbsp;\u003cbr\u003e\u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 315px;\"\u003eCTCAGTGCTCTACAGAAAACC\u003cbr\u003e\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 129px;\"\u003eOXA-1-F\u0026nbsp;\u003cbr\u003e\u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 315px;\"\u003eGGCACCAGATTCAACTTTCAAG\u003cbr\u003e\u003c/td\u003e\n \u003ctd rowspan=\"2\" style=\"width: 77px;\"\u003e564\u003cbr\u003e\u003c/td\u003e\n \u003ctd rowspan=\"2\" style=\"width: 57px;\"\u003e58\u003cbr\u003e\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 129px;\"\u003eOXA-1-R\u0026nbsp;\u003cbr\u003e\u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 315px;\"\u003eGACCCCAAGTTTCCTGTAAGTG\u003cbr\u003e\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 129px;\"\u003eSHV-28-F\u003cbr\u003e\u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 315px;\"\u003eCCGATGAACGCTTTCCCATG\u003cbr\u003e\u003c/td\u003e\n \u003ctd rowspan=\"2\" style=\"width: 77px;\"\u003e615\u003cbr\u003e\u003c/td\u003e\n \u003ctd rowspan=\"2\" style=\"width: 57px;\"\u003e57\u003cbr\u003e\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 129px;\"\u003eSHV-28-R\u0026nbsp;\u003cbr\u003e\u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 315px;\"\u003eGCGTATCCCGCAGATATATCAC\u003cbr\u003e\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 129px;\"\u003eCTX-M-14-F\u0026nbsp;\u003cbr\u003e\u003c/td\u003e\n \u003ctd style=\"width: 315px;\"\u003eGAGTGTTGCTCTGTGGATAAC\u003cbr\u003e\u003c/td\u003e\n \u003ctd rowspan=\"2\" style=\"width: 77px;\"\u003e741\u003cbr\u003e\u003c/td\u003e\n \u003ctd rowspan=\"2\" style=\"width: 57px;\"\u003e60\u003cbr\u003e\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 129px;\"\u003eCTX-M-14-R\u0026nbsp;\u003cbr\u003e\u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 315px;\"\u003eGTTACAGCCCTTCGGCGATG\u003cbr\u003e\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 129px;\"\u003eCTX-M-15-F\u0026nbsp;\u003cbr\u003e\u003c/td\u003e\n \u003ctd style=\"width: 315px;\"\u003eCGTCTCTTCCAGAATAAGG\u003cbr\u003e\u003c/td\u003e\n \u003ctd rowspan=\"2\" style=\"width: 77px;\"\u003e905\u003cbr\u003e\u003c/td\u003e\n \u003ctd rowspan=\"2\" style=\"width: 57px;\"\u003e60\u003cbr\u003e\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 129px;\"\u003eCTX-M-15-R\u003cbr\u003e\u003c/td\u003e\n \u003ctd style=\"width: 315px;\"\u003eGTTTCCCCATTCCGTTTCCGC\u003cbr\u003e\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 129px;\"\u003eCTX-M-55-F\u003cbr\u003e\u003c/td\u003e\n \u003ctd style=\"width: 315px;\"\u003eCAAAGAGAGTGCAACGGATG\u003cbr\u003e\u003c/td\u003e\n \u003ctd rowspan=\"2\" style=\"width: 77px;\"\u003e695\u003cbr\u003e\u003c/td\u003e\n \u003ctd rowspan=\"2\" style=\"width: 57px;\"\u003e60\u003cbr\u003e\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 129px;\"\u003eCTX-M-55-R\u003cbr\u003e\u003c/td\u003e\n \u003ctd style=\"width: 315px;\"\u003eATTGGAAAGCGTTCATCACC\u003cbr\u003e\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 129px;\"\u003eCTX-M-64-F\u003cbr\u003e\u003c/td\u003e\n \u003ctd style=\"width: 315px;\"\u003eAACTTGCCGAATTAGAGCGG\u003cbr\u003e\u003c/td\u003e\n \u003ctd rowspan=\"2\" style=\"width: 77px;\"\u003e1055\u003cbr\u003e\u003c/td\u003e\n \u003ctd rowspan=\"2\" style=\"width: 57px;\"\u003e56\u003cbr\u003e\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 129px;\"\u003eCTX-M-64-R\u003cbr\u003e\u003c/td\u003e\n \u003ctd style=\"width: 315px;\"\u003eGTTGCTCTGTGGATAACTTGC\u003cbr\u003e\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 129px;\"\u003eCTX-M-65-F\u003cbr\u003e\u003c/td\u003e\n \u003ctd style=\"width: 315px;\"\u003eGCAACGGATGATGTTCGCG\u003cbr\u003e\u003c/td\u003e\n \u003ctd rowspan=\"2\" style=\"width: 77px;\"\u003e832\u003cbr\u003e\u003c/td\u003e\n \u003ctd rowspan=\"2\" style=\"width: 57px;\"\u003e58\u003cbr\u003e\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 129px;\"\u003eCTX-M-65-R\u003cbr\u003e\u003c/td\u003e\n \u003ctd style=\"width: 315px;\"\u003eGCGGCTGGGTAAAATAGGTC\u003cbr\u003e\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 129px;\"\u003eTEM-1-F\u003cbr\u003e\u003c/td\u003e\n \u003ctd style=\"width: 315px;\"\u003eAATGATACCGCGAGACCCAC\u003cbr\u003e\u003c/td\u003e\n \u003ctd rowspan=\"2\" style=\"width: 77px;\"\u003e716\u003cbr\u003e\u003c/td\u003e\n \u003ctd rowspan=\"2\" style=\"width: 57px;\"\u003e58\u003cbr\u003e\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 129px;\"\u003eTEM-1-R\u003cbr\u003e\u003c/td\u003e\n \u003ctd style=\"width: 315px;\"\u003eAATGATACCGCGAGACCCAC\u003cbr\u003e\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 129px;\"\u003eTEM-244-F\u003cbr\u003e\u003c/td\u003e\n \u003ctd style=\"width: 315px;\"\u003eGATGCTGAAGATCAGTTGGGTG\u003cbr\u003e\u003c/td\u003e\n \u003ctd rowspan=\"2\" style=\"width: 77px;\"\u003e745\u003cbr\u003e\u003c/td\u003e\n \u003ctd rowspan=\"2\" style=\"width: 57px;\"\u003e58\u003cbr\u003e\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 129px;\"\u003eTEM-244-R\u003cbr\u003e\u003c/td\u003e\n \u003ctd style=\"width: 315px;\"\u003eGGCACCTATCTCAGCGATCTGTC\u003cbr\u003e\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 129px;\"\u003eDHA-1-F\u0026nbsp;\u003cbr\u003e\u003c/td\u003e\n \u003ctd style=\"width: 315px;\"\u003eGGTAAAACTGAGATGACGGGC\u003cbr\u003e\u003c/td\u003e\n \u003ctd rowspan=\"2\" style=\"width: 77px;\"\u003e1422\u003cbr\u003e\u003c/td\u003e\n \u003ctd rowspan=\"2\" style=\"width: 57px;\"\u003e58\u003cbr\u003e\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 129px;\"\u003eDHA-1-R\u0026nbsp;\u003cbr\u003e\u003c/td\u003e\n \u003ctd style=\"width: 315px;\"\u003eCTCATCCTCCATAAAACAGCCC\u003cbr\u003e\u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003eUnderlined sequences are restriction endonuclease sites.\u003c/p\u003e\n\u003ch2\u003e2.6 Plasmid conjugation experiments\u003c/h2\u003e\n\u003cp\u003eMultidrug-resistant \u003cem\u003eE.\u003c/em\u003e\u003cem\u003e\u0026nbsp;coli\u003c/em\u003e was used as the donor strain, and rifampicin-resistant E.\u003cem\u003e\u0026nbsp;coli\u003c/em\u003e EC600 was used as the recipient strain for the conjugation experiment of biparental mating. The transconjugants were selected from Mueller\u0026ndash;Hinton agar plates containing 600 \u0026mu;g/mL rifampin and 32 \u0026mu;g/mL cefoperazone/sulbactam or 8 \u0026mu;g/mL imipenem. Target resistance genes in the transformants and in the plasmid of transformant cells were verified using PCR, and the product was then sequenced.\u003c/p\u003e\n\u003ch2\u003e2.7 Comparative genomic\u0026nbsp;analysis of the \u0026beta;-lactamase gene-related sequences\u003c/h2\u003e\n\u003cp\u003eFor the comparative genomic analysis of \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX-M-64\u003c/sub\u003e and \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eKPC-2\u003c/sub\u003e gene-related fragments, sequences containing these \u003cem\u003ebla\u003c/em\u003e genes were obtained from the NCBI nucleotide database using the \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX-M-64\u0026nbsp;\u003c/sub\u003egene (NG_049015) or \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eKPC-2\u003c/sub\u003e gene (NG_049253) as a query for a BLASTN search. The results were retained only if the sequences contained complete \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX-M-64\u0026nbsp;\u003c/sub\u003e(20 kb, approximately 12 kb upstream and 8 kb downstream of the gene) or \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eKPC-2\u003c/sub\u003e (30 kb, approximately 21 kb upstream and 9 kb downstream of the gene). Multiple sequence alignments were performed by MAFFT24\u0026nbsp;(https://mafft.cbrc.jp/alignment/software/)\u0026nbsp;using each of the\u003cem\u003e\u0026nbsp;bla\u003c/em\u003e gene-related fragments of this work as a reference, and the sequences were clustered with an identity of \u0026ge;80%. The sequence sharing the highest similarity with the other sequences in each cluster was chosen as the candidate for ortholog analysis. Orthologous groups of genes from the candidate sequences were identified using BLASTP and InParanoid (https://inparanoid.sbc.su.se/).\u003c/p\u003e"},{"header":"3 Results","content":"\u003ch2\u003e3.1 Demographic and clinical characteristics\u003c/h2\u003e\n\u003cp\u003eA total of 110 \u003cem\u003eE. coli\u003c/em\u003e isolates were obtained from 110 patients (67 preterm and 43 full-term neonates) in the NICU. Most of the patients were male (55.3%), with a mean age of 16.4 \u0026plusmn; 11.9 days. Identified diseases included 41 cases of ventilator-associated pneumonia, 22 cases of necrotizing enterocolitis, three cases of skin infection, five cases of purulent meningitis, 20 cases of catheter-associated urinary tract infection, and 19 cases of neonatal septicemia. Clinical manifestations in infected newborns included fever in 67 cases, with an average peak of 38.6 \u0026plusmn; 1.38 ℃ and an average duration of 5.6 days. Inflammatory indicators of infected children included peripheral blood white cell count (13.78\u0026plusmn;7.56 \u0026times; 10\u003csup\u003e9\u003c/sup\u003e/L), C- reactive protein level (48.5 \u0026plusmn; 20.2 mg/L), and hypoallergenic (2.4 \u0026plusmn;0.8 ng/ml).\u003c/p\u003e\n\u003ch2\u003e3.2 Antimicrobial susceptibility\u003c/h2\u003e\n\u003cp\u003eThe minimum inhibitory concentration (MIC) of seven antibiotics (piperacillin-tazobactam, ceftazidime, cefepime, ceftriaxone, cefoperazone/sulbactam, imipenem, and ertapenem) was determined for the 110 \u003cem\u003eE. coli\u003c/em\u003e isolates, revealing resistance to all seven \u0026beta;-lactams. Resistance rates ranged from 2.7% (imipenem) to 54.5% (ceftriaxone). High-MIC isolates were associated with ventilator-associated pneumonia (35 cases), catheter-associated urinary tract infection (14 cases), necrotizing enterocolitis (seven cases), skin infection (one case), and neonatal septicemia (five cases). Two strains from ventilator-associated pneumonia and one strain from catheter-associated urinary tract infection showed resistance to all \u0026beta;-lactams, including imipenem and ertapenem. Additionally, two strains of ventilator-associated pneumonia, two strains of catheter-associated urinary tract infection, and one strain of neonatal septicemia were resistant to cefoperazone/sulbactam (Table 2).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eTable 2.\u003c/strong\u003e Antimicrobial susceptibility of the 110 \u003cem\u003eE. coli\u003c/em\u003e isolates against seven antimicrobials tested\u003c/p\u003e\n\u003ctable border=\"0\" cellspacing=\"0\" cellpadding=\"0\" width=\"593\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 159px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eAntimicrobial\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 65px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eS (%)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 61px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eI (%)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 72px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eR (%)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 72px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eMIC50 (\u0026mu;g/mL)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 70px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eMIC90 (\u0026mu;g/mL)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 94px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eRange (\u0026mu;g/mL)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 159px;\"\u003e\n \u003cp\u003ePiperacillin-Tazobactam\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 65px;\"\u003e\n \u003cp\u003e88.16\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 61px;\"\u003e\n \u003cp\u003e3.64 (4)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 72px;\"\u003e\n \u003cp\u003e8.20 (9/110)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 72px;\"\u003e\n \u003cp\u003e0.5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 70px;\"\u003e\n \u003cp\u003e16\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 94px;\"\u003e\n \u003cp\u003e0.5\u0026ndash;128\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 159px;\"\u003e\n \u003cp\u003eCeftazidime\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 65px;\"\u003e\n \u003cp\u003e57.28\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 61px;\"\u003e\n \u003cp\u003e17.27 (19)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 72px;\"\u003e\n \u003cp\u003e25.45 (28)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 72px;\"\u003e\n \u003cp\u003e4\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 70px;\"\u003e\n \u003cp\u003e16\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 94px;\"\u003e\n \u003cp\u003e0.5\u0026ndash;64\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 159px;\"\u003e\n \u003cp\u003eCefepime\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 65px;\"\u003e\n \u003cp\u003e46.37\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 61px;\"\u003e\n \u003cp\u003e25.45 (28)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 72px;\"\u003e\n \u003cp\u003e28.18 (31)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 72px;\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 70px;\"\u003e\n \u003cp\u003e16\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 94px;\"\u003e\n \u003cp\u003e0.25\u0026ndash;64\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 159px;\"\u003e\n \u003cp\u003eCeftriaxone\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 65px;\"\u003e\n \u003cp\u003e19.09\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 61px;\"\u003e\n \u003cp\u003e24.55 (27)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 72px;\"\u003e\n \u003cp\u003e56.36 (62)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 72px;\"\u003e\n \u003cp\u003e4\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 70px;\"\u003e\n \u003cp\u003e16\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 94px;\"\u003e\n \u003cp\u003e1\u0026ndash;256\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 159px;\"\u003e\n \u003cp\u003eErtapenem\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 65px;\"\u003e\n \u003cp\u003e92.72\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 61px;\"\u003e\n \u003cp\u003e4.55 (5)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 72px;\"\u003e\n \u003cp\u003e2.73 (3)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 72px;\"\u003e\n \u003cp\u003e0.125\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 70px;\"\u003e\n \u003cp\u003e8\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 94px;\"\u003e\n \u003cp\u003e0.0625\u0026ndash;64\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 159px;\"\u003e\n \u003cp\u003eImipenem\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 65px;\"\u003e\n \u003cp\u003e90.00\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 61px;\"\u003e\n \u003cp\u003e7.27 (8)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 72px;\"\u003e\n \u003cp\u003e2.73\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 72px;\"\u003e\n \u003cp\u003e0.125\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 70px;\"\u003e\n \u003cp\u003e8\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 94px;\"\u003e\n \u003cp\u003e0.0625\u0026ndash;64\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 159px;\"\u003e\n \u003cp\u003eCefoperazone/sulbactam\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 65px;\"\u003e\n \u003cp\u003e83.64\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 61px;\"\u003e\n \u003cp\u003e10.00 (11)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 72px;\"\u003e\n \u003cp\u003e6.36 (7)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 72px;\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 70px;\"\u003e\n \u003cp\u003e8\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 94px;\"\u003e\n \u003cp\u003e0.0625\u0026ndash;64\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003ch2\u003e3.3 Distribution of \u0026beta;-lactamases\u003c/h2\u003e\n\u003cp\u003eA total of 111 resistance genes of six genotypes were identified in 62 strains from 110 \u003cem\u003eE. coli\u003c/em\u003e isolates through \u003cem\u003ebla\u003c/em\u003e gene screening. \u003cem\u003eBla\u003c/em\u003e\u003csub\u003eCTX-M\u003c/sub\u003e showed the highest positivity rate of 50.45% (56/111), which consisted of five sub-genotypes: \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX-M-14\u0026nbsp;\u003c/sub\u003e(17.12%, 19/111),\u003cem\u003e\u0026nbsp;bla\u003c/em\u003e\u003csub\u003eCTX-M-15\u0026nbsp;\u003c/sub\u003e(13.51%, 15/111),\u003cem\u003e\u0026nbsp;bla\u003c/em\u003e\u003csub\u003eCTX-M-55\u0026nbsp;\u003c/sub\u003e(9.91%, 11/111),\u003cem\u003e\u0026nbsp;bla\u003c/em\u003e\u003csub\u003eCTX-M-64\u0026nbsp;\u003c/sub\u003e(5.41%, 6/111), and \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX-M-65\u003c/sub\u003e (4.50%, 5/111). \u003cem\u003eBla\u003c/em\u003e\u003csub\u003eTEM\u003c/sub\u003e was second (36.04%, 40/111), followed by \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eTEM-1\u0026nbsp;\u003c/sub\u003e(21.62%, 24/111), \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eTEM-244\u0026nbsp;\u003c/sub\u003e(14.41%, 16/111), and \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eSHV-28\u0026nbsp;\u003c/sub\u003e(7.21%, 8/111). The remaining three (\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eKPC-2\u003c/sub\u003e, \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eOXA-1\u003c/sub\u003e,\u003csub\u003e\u0026nbsp;\u003c/sub\u003eand \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eDHA-1\u003c/sub\u003e) showed lower positivity rates of 2.70% (3/111), 1.80% (2/111), and 1.80% (2/111), respectively. Approximately half the 62 \u0026beta;-lactamase gene- harboring isolates (54.84%, 24/62) carried two resistance genes, eight isolates carried three resistance genes, three isolates carried four resistance genes, and the remaining 27 carried one gene (Table 3). These 62 isolates revealed higher MICs to \u0026beta;-lactams. All strains had MIC values \u0026ge; 32 \u0026mu;g/mL for ceftriaxone, with 50.00% (31/62) and 45.16% (28/62) had MIC levels \u0026ge; 16 \u0026mu;g/mL for cefepime and ceftazidime. Only three strains (4.84%, 3/62) had MIC levels \u0026ge; 8 \u0026mu;g/mL for ertapenem or imipenem (Table 3). None of the \u0026beta;-lactamase gene-negative strains showed resistance to the evaluated \u0026beta;-lactams.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eTable 3.\u003c/strong\u003e MIC results of the isolates harboring three or four \u0026beta;-lactamase gene and transconjugants\u003c/p\u003e\n\u003ctable border=\"0\" cellspacing=\"0\" cellpadding=\"0\" align=\"\" width=\"698\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eIsolate\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"7\" valign=\"top\"\u003e\n \u003cp\u003eMIC (\u0026mu;g/ml) of Antimicrobial agent\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"3\" rowspan=\"2\" valign=\"top\"\u003e\n \u003cp\u003eResistance mechanism\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eNo.\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eceftazidime\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003ecefepime\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eertapenem\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eimipenem\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eCeftriaxone\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003epiperacillin-tazobactam\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003ecefoperazone/sulbactam\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eECF13\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e64\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e64\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e0.25\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e0.5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e64\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e64\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e64\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX-M-64\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX-M-64\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX-M-14\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eTEM-1\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eECK46\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e32\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e64\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e128\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e64\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX-M-14\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX-M-15\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eSHV-28\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eTEM-1\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eECK42\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e64\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e64\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e32\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e32\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e64\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e32\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e32\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX-M-64\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eKPC-2\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eTEM-244\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eECK55\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e32\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e32\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e64\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e64\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eTEM-1\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX-M-15\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eSHV-28\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eECK67\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e32\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e32\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e32\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e32\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e128\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e128\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX-M-55\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eKPC-2\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX-M-64\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eECK03\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e64\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e64\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e64\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e64\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e128\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e128\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e32\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eKPC-2\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX-M-64\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eTEM-1\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX-M-14\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eECK01\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e64\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e32\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e4\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e32\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e128\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e32\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX-M-55\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eTEM-244\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eSHV-28\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eECK69\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e64\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e64\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e64\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e64\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e16\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eTEM-244\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eOXA-1\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eDHA-1\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eECF04\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e32\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e32\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e4\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e8\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e128\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e128\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e32\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX-M-65\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eTEM-1\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eDHA-1\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eECF09\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e64\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e64\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e8\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e8\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e64\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e4\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e32\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eTEM-244\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX-M-55\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eOXA-1\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eECF81\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e32\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e32\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e0.25\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e0.5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e64\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e8\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX-M-65\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eTEM-244\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX-M-15\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eECF04/EC600\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e32\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e32\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e4\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e4\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e32\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e64\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e16\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX-M-65\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eECK03/EC600\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e64\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e32\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e64\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e64\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e128\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e128\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e32\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eKPC-2\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX-M-64\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eTEM-1\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eECK55/EC600\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e32\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e32\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e8\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e32\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e32\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX-M-15\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eSHV-28\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eECF09/EC600\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e32\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e32\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e8\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e32\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e32\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e8\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e16\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX-M-55\u003c/sub\u003e-45\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eOXA-1\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003ch2\u003e3.4 Resistant plasmid transferability\u003c/h2\u003e\n\u003cp\u003eOf the five strains resistant to the \u0026beta;-lactams, the conjugative plasmids of four strains were successfully transferred to \u003cem\u003eE. coli\u003c/em\u003e EC600. PCR amplification revealed that two of the transconjugants carried a \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eKPC-2\u0026nbsp;\u003c/sub\u003egene (including one co-expressing \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX-M-64\u003c/sub\u003e and \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eTEM-1\u0026nbsp;\u003c/sub\u003egenes), and the other two carried a \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eOXA-1\u0026nbsp;\u003c/sub\u003egene. The four transconjugants with \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eOXA-1\u0026nbsp;\u003c/sub\u003eexhibited reduced susceptibility to carbapenems with MIC values up to 32-fold compared with that in the recipients. The two \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eKPC-2\u0026nbsp;\u003c/sub\u003eharboring transconjugants showed MIC values up to 64-fold compared with that in recipients against carbapenems (Table 3). The remaining strain harboring \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eKPC-2\u0026nbsp;\u003c/sub\u003efailed the conjugation experiments.\u003c/p\u003e\n\u003ch2\u003e3.5 General features of the E. Coli genomes\u003c/h2\u003e\n\u003cp\u003eWhole genome sequencing and further molecular analysis were performed for two strains carrying different \u0026beta;-lactamases. The two isolates included \u003cem\u003eE. coli\u003c/em\u003e ECK03 carrying \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eKPC-2\u003c/sub\u003e, \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX-M-64\u003c/sub\u003e, \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX-M-65\u003c/sub\u003e,\u003csub\u003e\u0026nbsp;\u003c/sub\u003eand\u003csub\u003e\u0026nbsp;\u003c/sub\u003e\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eTEM-1\u003c/sub\u003e, and \u003cem\u003eE. coli\u003c/em\u003e ECF13 carrying \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX-M-14,\u0026nbsp;\u003c/sub\u003e\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eTEM-1\u003c/sub\u003e, and two genes of \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX-M-64\u003c/sub\u003e. Sequencing results showed that only ECK03 carried one resistance plasmid. The \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eKPC-2\u0026nbsp;\u003c/sub\u003eof \u003cem\u003eE. coli\u0026nbsp;\u003c/em\u003eECK03 was encoded in a plasmid (pECK03_KPC-2), while the two genes of \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX-M-64\u0026nbsp;\u003c/sub\u003eof \u003cem\u003eE. coli\u0026nbsp;\u003c/em\u003eECF13 were encoded on the chromosomes. ECK03 harbored four plasmids, a 114 kb plasmid named pECK03_KPC-2 harbored \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eKPC-2\u003c/sub\u003e, \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX-M-64\u003c/sub\u003e,\u003csub\u003e\u0026nbsp;\u003c/sub\u003eand\u003csub\u003e\u0026nbsp;\u003c/sub\u003e\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eTEM-1\u003c/sub\u003e, and another three plasmids free of the resistance gene). ECF13 contained two plasmids free of resistance genes.\u003c/p\u003e\n\u003ch2\u003e3.6 Comparative genomic analysis of sequences harbored\u003cem\u003e\u0026nbsp;bla\u003c/em\u003e\u003csub\u003eKPC-2\u003c/sub\u003e and \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX-M-64\u003c/sub\u003e\u003c/h2\u003e\n\u003cp\u003eComparative analysis of sequence data from the NCBI database revealed four plasmids sharing maximum similarities with pECK03_KPC-2 in two \u003cem\u003eE. coli\u003c/em\u003e species: p116753-KPC (\u003cem\u003eK. pneumoniae\u003c/em\u003e, MN891682.1), pKP19-3138-4 (\u003cem\u003eK. pneumoniae\u003c/em\u003e, CP090620.1), pOW1E2a (\u003cem\u003eE.\u003c/em\u003e \u003cem\u003ecoli\u003c/em\u003e, CP067246.1), and pKPC2_090374 (\u003cem\u003eK. pneumoniae\u003c/em\u003e, CP066536.1) (Figure 1). Plasmid pECK03_KPC-2 harbored three \u003cem\u003ebla\u003c/em\u003e resistance genes: \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eKPC-2\u003c/sub\u003e, \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX-M-64\u003c/sub\u003e,\u003csub\u003e\u0026nbsp;\u003c/sub\u003eand\u003csub\u003e\u0026nbsp;\u003c/sub\u003e\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eTEM-1\u003c/sub\u003e (Figure 2). Two genes, \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX-M-64\u0026nbsp;\u003c/sub\u003efrom \u003cem\u003eE. coli\u003c/em\u003e ECF13 with \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX-M-14,\u0026nbsp;\u003c/sub\u003e\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eTEM-1\u003c/sub\u003e, and two \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX-M-64\u0026nbsp;\u003c/sub\u003egenes,\u003csub\u003e\u0026nbsp;\u003c/sub\u003ewere found on the chromosomes. The sequence of the first \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX-M-64\u0026nbsp;\u003c/sub\u003epossessed an identical gene environment to that of clone CP104274 of \u003cem\u003eE. coli\u003c/em\u003e EC15103\u003cem\u003e\u0026nbsp;\u003c/em\u003econtaining the \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX-M-64\u0026nbsp;\u003c/sub\u003egene. However, the sequence of the second \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX-M-64\u003c/sub\u003e was not identified in the sequences of the highest similarity gene environment containing \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX-M-64\u003c/sub\u003e (Figures 3 and 4).\u0026nbsp;\u003c/p\u003e"},{"header":"4 Discussion","content":"\u003cp\u003eIn this study, all isolates were from the NICU, and 62 of the 110 clinical isolates had high MICs. Due to the limitations of antibiotic use in pediatric patients, especially neonates, only β-lactam antibiotics are used against gram-negative bacteria. Therefore, the high drug resistance rate poses significant challenges for selecting antibiotics in clinical practice. In addition, 49 of the 62 (79.03%) isolates were from ventilator-associated pneumonia and catheter-associated urinary tract infections and were believed to be hospital-acquired infections. From the high MIC isolates, nine \u003cem\u003ebla\u003c/em\u003e genotypes (11 sub-genotypes) of Ambler classes were identified, including class A (\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eKPC\u0026minus;2,\u003c/sub\u003e \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eSHV\u0026minus;28,\u003c/sub\u003e \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX\u0026minus;M\u0026minus;14,\u003c/sub\u003e \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX\u0026minus;M\u0026minus;15,\u003c/sub\u003e \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX\u0026minus;M\u0026minus;55,\u003c/sub\u003e \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX\u0026minus;M\u0026minus;64,\u003c/sub\u003e \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX\u0026minus;M\u0026minus;65\u003c/sub\u003e, \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eTEM\u0026minus;1,\u003c/sub\u003e and \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eTEM\u0026minus;244\u003c/sub\u003e), class C (\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eDHA\u0026minus;1\u003c/sub\u003e), and class D (\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eOXA\u0026minus;1\u003c/sub\u003e), of which Ambler class A was the most prevalent with a total of 57 strains (91.94%, 57/62). In this study, \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX\u0026minus;M\u003c/sub\u003e showed the highest positive rate of 50.45% (56/111), and \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eTEM\u003c/sub\u003e was second (36.04%, 40/111). These results suggest that all β-lactam antibiotics except carbapenem are prone to high resistance (\u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e12\u003c/span\u003e). As an ESBL variant, CTX-M β-lactamases are prevalent and widespread worldwide (\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e13\u003c/span\u003e). In this study, three isolates from patients with ventilator-associated pneumonia (2.73%, 3/110) carrying \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eKPC\u0026minus;2\u003c/sub\u003e revealed higher MIC values to ertapenem and imipenem (both 64 \u0026micro;g/mL). As a less frequent but much more powerful carbapenem-hydrolyzing enzyme in \u003cem\u003eE. coli\u003c/em\u003e, KPC-2 belongs to the class A β-lactamase, and was frequently reported in \u003cem\u003eKlebsiella pneumoniae\u003c/em\u003e samples from NICUs (\u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e14\u003c/span\u003e) but was rarely reported in NICU \u003cem\u003eE. coli\u003c/em\u003e strains. In addition, the two strains carrying \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eOXA\u0026minus;1\u003c/sub\u003e showed weak carbapenem resistance. Despite the presence of genes that are resistant to carbapenems, the resistance rate was not higher than that reported in other studies (\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e15\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eEight of the 62 isolates carried three resistance genes, and three strains carried four resistance genes. The resistance gene combinations of each strain were mainly composed of \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX\u0026minus;M\u003c/sub\u003e and \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eTEM,\u003c/sub\u003e and often, two \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX\u0026minus;MS\u003c/sub\u003e or two \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eTEM\u003c/sub\u003es were present, and two identical \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX\u0026minus;M\u003c/sub\u003eor \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eTEM\u003c/sub\u003e genes existed in individual strains, such as \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX\u0026minus;M\u0026minus;64\u003c/sub\u003e. No similar β-lactamase gene combinations co-occurring with identical \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX\u0026minus;M\u003c/sub\u003e or \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eTEM\u003c/sub\u003e genes have been reported. In addition, \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eKPC\u0026minus;2\u003c/sub\u003e and \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eOXA\u0026minus;1\u003c/sub\u003e are all present in these multiple β-lactamase isolates. The co-occurrence of β-lactamase genes displays high rates of resistance to broad-spectrum cephalosporins owing to induced expression of the β-lactamase gene; an increasing number of organisms harboring plasmid-encoded ESBL and/or carbapenemase genes have been reported (\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e16\u003c/span\u003e). The most common is the co-occurrence of two different β-lactamase genes; however, the co-occurrence of three or more β-lactamase genes is rare (\u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e17\u003c/span\u003e), with no report from NICUs.\u003c/p\u003e \u003cp\u003eWhole genome sequencing of the isolate ECK03 displayed that the β-lactamase genes were related to mobile genetic elements and were mainly harbored on plasmids. The resistance genes of pECK03_KPC-2 clustered mainly in the multidrug-resistant (MDR) region. The MDR regions of pECK03_KPC-2 carrying \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eKPC\u0026minus;2\u003c/sub\u003e, \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX\u0026minus;M\u0026minus;64,\u003c/sub\u003e and \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eTEM\u0026minus;1\u003c/sub\u003e were similar to those of plasmid MN891682 (19). Compared with MN891682 and other plasmids with similar MDR regions, the MDR region of pECK03_KPC-2 contained many more massive insertions of foreign resistance genetic contents and showed a higher degree of genomic plasticity. Two identical \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX\u0026minus;M\u0026minus;64\u003c/sub\u003e genes in ECF13 were located on different chromosomes and contained other resistance genes, such as \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX\u0026minus;M\u0026minus;14\u003c/sub\u003e and \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eTEM\u0026minus;1\u003c/sub\u003e. This rare combination of MDR genes demonstrated stronger resistance than the other isolates.\u003c/p\u003e \u003cp\u003eIn conclusion, the result reveals a high ratio of β-lactam antibiotic resistance in \u003cem\u003eE. coli\u003c/em\u003e strains from NICUs. Notably, among the 62 of 110 isolates that carried β-lactamase genes, \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX\u0026minus;M\u003c/sub\u003e and \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eTEM\u003c/sub\u003e were the most common, and co-occurrence of two or more β-lactamase genes was prevalent. The resistance gene-carrying plasmid may be transferred, and the harbored β-lactamase gene maintains resistance to β-lactams in recipients. In contrast, the three co-carrying \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eKPC\u0026minus;2\u003c/sub\u003e also exhibited reduced susceptibility to carbapenems. The identification of high β-lactam antibiotic resistance in NICU E. coli strains highlights the urgent need for targeted antibiotic stewardship and infection control measures in neonatal healthcare settings. Clinicians should carefully consider alternative treatment options, given the observed prevalence of specific resistance genes, such as blaCTX-M and blaTEM, to ensure effective management of E. coli infections in this vulnerable patient population.\u003c/p\u003e"},{"header":"Declarations","content":"\u003ch2\u003eConflict of interest\u003c/h2\u003e\n\u003cp\u003eThe authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.\u003c/p\u003e\n\u003ch2\u003eEthics approval\u003c/h2\u003e\n\u003cp\u003eThis study was approved by the Institutional Ethics Committee of the First People\u0026apos;s Hospital of Yongkang, Zhejiang, China (approval number: ykyy2020-87). Informed consent was obtained from all participants involved in the study.\u003c/p\u003e\n\u003ch2\u003eClinical Trial Registration\u003c/h2\u003e\n\u003cp\u003eNot applicable.\u003c/p\u003e\n\u003ch2\u003eConsent for publication\u003c/h2\u003e\n\u003cp\u003eNot applicable.\u003c/p\u003e\n\u003ch2\u003eAvailability of data and material\u003c/h2\u003e\n\u003cp\u003eThe datasets used and/or analysed during the current study are available from the corresponding author on reasonable request.\u003c/p\u003e\n\u003ch2\u003eFunding\u003c/h2\u003e\n\u003cp\u003eThis work was supported by Medical and Health Science and Technology Project of Zhejiang Province (CN, no. 2020ZH068).\u003c/p\u003e\n\u003ch2\u003eAuthor Contribution\u003c/h2\u003e\n\u003cp\u003eJ.Z was responsible for the study\u0026apos;s conceptualization, methodology, project administration, formal analysis, data analysis, and also contributed to the review and editing of the manuscript.JQ.Z and M.C both conducted the investigation and curated the data. P.L was involved in testing. CM.J provided supervision, wrote the original draft, and contributed to the manuscript\u0026apos;s review, editing, and validation. All authors have read and consented to the published version of the manuscript.\u003c/p\u003e\n\u003ch2\u003eAcknowledgments\u003c/h2\u003e\n\u003cp\u003eThe authors would like to thank the participants, who made this study possible.\u003c/p\u003e\n\u003ch2\u003eData Availability\u003c/h2\u003e\n\u003cp\u003eSequence data that support the findings of this study have been deposited in national center for biobechnology ;Accession: SAMN37977427 ID: 37977427.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\n\u003cli\u003eLonghi C, Maurizi L, Conte AL, Marazzato M, Comanducci A, Nicoletti M et al. Extraintestinal pathogenic Escherichia coli: beta-lactam antibiotic and heavy metal resistance. Antibiotics (Basel) (2022) 11:328. doi: 10.3390/antibiotics11030328\u003c/li\u003e\n\u003cli\u003eFlannery DD, Akinboyo IC, Mukhopadhyay S, Tribble AC, Song L, Chen F et al. Antibiotic Susceptibility of Escherichia coli Among Infants Admitted to Neonatal\u003c/li\u003e\n\u003cli\u003eSeidel J, Haller S, Eckmanns T, Harder T. Routine screening for colonization by Gram-negative bacteria in neonates at intensive care units for the prediction of sepsis: systematic review and meta-analysis. J Hosp Infect (2018) 99:367-80. doi: 10.1016/j.jhin.2018.03.017\u003c/li\u003e\n\u003cli\u003eLee YQ, Ahmad Kamar A, Velayuthan RD, Chong CW, Teh CSJ. Clonal relatedness in the acquisition of intestinal carriage and transmission of multidrug resistant (MDR) Klebsiella pneumoniae and Escherichia coli and its risk factors among preterm infants admitted to the neonatal intensive care unit (NICU). Pediatr Neonatol (2021) 62:129-37. doi: 10.1016/j.pedneo.2020.10.002\u003c/li\u003e\n\u003cli\u003eBlair JM, Webber MA, Baylay AJ, Ogbolu DO, Piddock LJ. Molecular mechanisms of antibiotic resistance. Nat Rev Microbiol (2015) 13:42-51. doi: 10.1038/nrmicro3380\u003c/li\u003e\n\u003cli\u003eMeini S, Tascini C, Cei M, Sozio E, Rossolini GM. AmpC \u0026beta;-lactamase-producing Enterobacterales: what a clinician should know. Infection (2019) 47:363-75. doi: 10.1007/s15010-019-01291-9\u003c/li\u003e\n\u003cli\u003eMichel-Briand Y. Resistance to the latest beta-lactams: mechanisms of acquisition and spread of resistance in Enterobacteriaceae. Bull Acad Natl Med (2007) 191:35-50; discussion 50-1\u003c/li\u003e\n\u003cli\u003eDaoud Z, Salem Sokhn E, Masri K, Matar GM, Doron S. Escherichia coli isolated from urinary tract infections of Lebanese patients between 2005 and 2012: epidemiology and profiles of resistance. Front Med (Lausanne) (2015) 2:26. doi: 10.3389/fmed.2015.000269 \u003c/li\u003e\n\u003cli\u003eHayer SS, Casanova-Higes A, Paladino E, Elnekave E, Nault A, Johnson T et al. Global distribution of extended spectrum cephalosporin and carbapenem resistance and associated resistance markers in Escherichia coli of swine origin \u0026ndash; A systematic review and meta-analysis. Front Microbiol (2022) 13:853810. doi: 10.3389/fmicb.2022.853810\u003c/li\u003e\n\u003cli\u003eSch\u0026auml;fer F, G\u0026ouml;rner P, Woltemate S, Brandenberger C, Geffers R, Ziesing S et al. The resistance mechanism governs physiological adaptation of Escherichia coli to growth with sublethal concentrations of carbapenem. Front Microbiol (2021) 12:812544. doi: 10.3389/fmicb.2021.812544\u003c/li\u003e\n\u003cli\u003eWedel E, Bernabe-Balas C, Ares-Arroyo M, Montero N, Santos-Lopez A, Mazel D et al. Insertion sequences determine plasmid adaptation to new bacterial Hosts. mBio (2023) 14:e0315822. doi: 10.1128/mbio.03158-22\u003c/li\u003e\n\u003cli\u003eOteo J, Cercenado E, Fern\u0026aacute;ndez-Romero S, Sa\u0026eacute;z D, Padilla B, Zamora E et al. Extended-spectrum-\u0026beta;-lactamase-producing Escherichia coli as a cause of pediatric infections: report of a neonatal intensive care unit outbreak due to a CTX-M-14-producing strain. Antimicrob Agents Chemother (2012) 56:54-8. doi: 10.1128/AAC.05103-11\u003c/li\u003e\n\u003cli\u003eGiedraitiene A, Pereckaite L, Bredelyte-Gruodiene E, Virgailis M, Ciapiene I, Tatarunas V. CTX-M-producing Escherichia coli strains: resistance to temocillin, fosfomycin, nitrofurantoin and biofilm formation.Future Microbiol (2022),17:789-802. doi: 10.2217/fmb-2021-0202.\u003c/li\u003e\n\u003cli\u003eMaida CM, Bonura C, Geraci DM, Graziano G, Carattoli A, Rizzo A et al. Outbreak of ST395 KPC-producing Klebsiella pneumoniae in a neonatal intensive care unit in Palermo, Italy. Infect Control Hosp Epidemiol (2018) 39:496-8. doi: 10.1017/ice.2017.267\u003c/li\u003e\n\u003cli\u003eXu Q, Pan F, Sun Y, Wang C, Shi Y, Zhang T et al. Fecal carriage and molecular epidemiology of carbapenem-resistant Enterobacteriaceae from inpatient children in a pediatric hospital of shanghai. Infect Drug Resist (2020) 13:4405-15. doi: 10.2147/IDR.S275549\u003c/li\u003e\n\u003cli\u003eZorgani A, Daw H, Sufya N, Bashein A, Elahmer O, Chouchani C. Co-occurrence of plasmid-mediated AmpC \u0026beta;-lactamase activity among Klebsiella pneumoniae and Escherichia coli. Open Microbiol J (2017) 11:195-202. doi: 10.2174/1874285801711010195\u003c/li\u003e\n\u003cli\u003eRamoul A, Loucif L, Bakour S, Amiri S, Dekhil M, Rolain JM. Co-occurrence of blaNDM-1 with blaOXA-23 or blaOXA-58 in clinical multidrug-resistant Acinetobacter baumannii isolates in Algeria. J Glob Antimicrob Resist (2016) 6:136-41. doi: 10.1016/j.jgar.2016.05.003\u003c/li\u003e\n\u003cli\u003eJeong S, Kim JO, Yoon EJ, Bae IK, Lee W, Lee H et al. Extensively drug-resistant Escherichia coli sequence Type 1642 carrying an IncX3 plasmid containing the blaKPC-2 gene associated with transposon Tn4401a. Ann Lab Med (2018) 38:17-22. doi: 10.3343/alm.2018.38.1.17\u003c/li\u003e\n\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":false,"highlight":"","institution":"","isAcceptedByJournal":true,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"bmc-pediatrics","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"bped","sideBox":"Learn more about [BMC Pediatrics](http://bmcpediatr.biomedcentral.com/)","snPcode":"","submissionUrl":"https://www.editorialmanager.com/bped/default.aspx","title":"BMC Pediatrics","twitterHandle":"BMC_series","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"em","reportingPortfolio":"BMC Series","inReviewEnabled":true,"inReviewRevisionsEnabled":true},"keywords":"Escherichia coli, resistance, antimicrobial susceptibility test, β-lactamase gene, neonatal ICU","lastPublishedDoi":"10.21203/rs.3.rs-5116757/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-5116757/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003e\u003cstrong\u003eIntroduction: \u003c/strong\u003e\u003cem\u003eEscherichia coli\u003c/em\u003e (\u003cem\u003eE. coli\u003c/em\u003e) causes infections in neonates admitted to neonatal intensive care units (NICUs). Although β-lactam antibiotics are commonly used for neonatal infectious diseases,\u003cem\u003e E. coli \u003c/em\u003ehas\u003cem\u003e \u003c/em\u003eexhibited resistance to them. Therefore, we investigated the resistance of \u003cem\u003eE. coli\u003c/em\u003e strains isolated from a NICU to β-lactam antibiotics.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eMethods: \u003c/strong\u003e\u003cem\u003eE. coli\u003c/em\u003e isolates were collected from patients admitted to a NICU from 2020–2023. The clinical characteristics of the patients were analyzed. The \u003cu\u003eantimicrobial susceptibility\u003c/u\u003e was determined using the agar dilution method, and the distribution of β-lactamase genes was analyzed using PCR. Conjugation experiments were conducted to analyze the horizontal transferability of resistance genes on plasmids. Genomic DNA was extracted for whole genome sequencing, construction of plasmid physical maps, locating resistance genes, and analyzing flanking regions and the resistance gene-related sequences.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eResults:\u003c/strong\u003e Throughout the study period, 110 distinct \u003cem\u003eE. coli \u003c/em\u003estrains were collected. Among these, 62 cases presented strains with high minimum inhibitory concentrations (MIC) associated with conditions such as ventilator-associated pneumonia (35/62), catheter-associated urinary tract infection (14/62), necrotizing enterocolitis (7/62), skin infection (1/62), and neonatal septicemia (5/62). Resistance of \u003cem\u003eE. coli\u003c/em\u003e \u003cem\u003ei\u003c/em\u003esolates to seven β-lactam antibiotics ranged from 2.73–56.36%. In 62 strains (56.36%, 62/110), six genotypes (11 sub-genotypes) of 111 β-lactamase genes were identified. Conjugation experiments revealed two transconjugants carrying the \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eKPC-2 \u003c/sub\u003egene and two carrying the \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eOXA-1 \u003c/sub\u003egene, exhibiting resistance to carbapenems and other β-lactams. The plasmids of four strains were successfully conjugated and transferred to recipient \u003cem\u003eE. coli\u003c/em\u003e C600. PCR of the transconjugant resistance genes revealed that two carried a \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eKPC-2 \u003c/sub\u003egene with a MIC increased up to 32-fold relative to the recipients, and the other two carried a \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eOXA-1 \u003c/sub\u003egene with a 32-fold increased MIC. For isolate ECK03 carrying \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eKPC-2\u003c/sub\u003e, \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX-M-64\u003c/sub\u003e, \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX-M-65\u003c/sub\u003e,\u003csub\u003e \u003c/sub\u003eand\u003csub\u003e \u003c/sub\u003e\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eTEM-1\u003c/sub\u003e, sequencing results showed that \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eKPC-2\u003c/sub\u003e, \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX-M-64\u003c/sub\u003e,\u003csub\u003e \u003c/sub\u003eand\u003csub\u003e \u003c/sub\u003e\u003cem\u003ebla\u003c/em\u003e\u003csub\u003eTEM-1 \u003c/sub\u003ewere harbored on a 114-kb pECK03_KPC-2 plasmid, whereas two identical \u003cem\u003ebla\u003c/em\u003e\u003csub\u003eCTX-M-64 \u003c/sub\u003egenes were harbored in \u003cem\u003eE. coli\u003c/em\u003e isolate ECF13.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConclusion: \u003c/strong\u003eThese findings highlight the existence of \u003cem\u003eE. coli\u003c/em\u003e β-lactam resistance within NICU populations, emphasizing the need for continual monitoring of β-lactamase isolates to facilitate effective antibiotic selection.\u003c/p\u003e","manuscriptTitle":"Prevalence of β-lactam antibiotic resistance of Escherichia coli isolated from a neonatal intensive care unit","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2024-12-17 16:48:25","doi":"10.21203/rs.3.rs-5116757/v1","editorialEvents":[{"type":"communityComments","content":0},{"type":"decision","content":"Revision requested","date":"2024-09-30T06:23:29+00:00","index":"","fulltext":""},{"type":"editorAssigned","content":"","date":"2024-09-28T04:57:30+00:00","index":"","fulltext":""},{"type":"checksComplete","content":"","date":"2024-09-28T04:56:34+00:00","index":"","fulltext":""},{"type":"submitted","content":"BMC Pediatrics","date":"2024-09-19T11:53:23+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"bmc-pediatrics","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"bped","sideBox":"Learn more about [BMC Pediatrics](http://bmcpediatr.biomedcentral.com/)","snPcode":"","submissionUrl":"https://www.editorialmanager.com/bped/default.aspx","title":"BMC Pediatrics","twitterHandle":"BMC_series","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"em","reportingPortfolio":"BMC Series","inReviewEnabled":true,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"766f6fb7-2093-4d46-9142-27325b125aed","owner":[],"postedDate":"December 17th, 2024","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"published-in-journal","subjectAreas":[],"tags":[],"updatedAt":"2025-02-10T16:06:13+00:00","versionOfRecord":{"articleIdentity":"rs-5116757","link":"https://doi.org/10.1186/s12887-025-05389-y","journal":{"identity":"bmc-pediatrics","isVorOnly":false,"title":"BMC Pediatrics"},"publishedOn":"2025-02-04 15:58:19","publishedOnDateReadable":"February 4th, 2025"},"versionCreatedAt":"2024-12-17 16:48:25","video":"","vorDoi":"10.1186/s12887-025-05389-y","vorDoiUrl":"https://doi.org/10.1186/s12887-025-05389-y","workflowStages":[]},"version":"v1","identity":"rs-5116757","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-5116757","identity":"rs-5116757","version":["v1"]},"buildId":"qtupq5eGEP_6zYnWcrvyt","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}