Whole-Genome Assessment of Clinical Acinetobacter baumannii Isolates Uncovers Potentially Novel Factors Influencing Carbapenem Resistance

Carbapenems—one of the important last-line antibiotics for the treatment of gram-negative infections—are becoming ineffective for treating Acinetobacter baumannii infections. Studies have identified multiple genes (and mechanisms) responsible for carbapenem resistance. In some A. baumannii strains, the presence/absence of putative resistance genes is not consistent with their resistance phenotype—indicating the genomic factors underlying carbapenem resistance in A. baumannii are not fully understood. Here, we describe a large-scale whole-genome genotype-phenotype association study with 349 A. baumannii isolates that extends beyond the presence/absence of individual antimicrobial resistance genes and includes the genomic positions and pairwise interactions of genes. Ten known resistance genes exhibited statistically significant associations with resistance to imipenem, a type of carbapenem: blaOXA-23, qacEdelta1, sul1, mphE, msrE, ant(3”)-II, aacC1, yafP, aphA6, and xerD. A review of the strains without any of these 10 genes uncovered a clade of isolates with diverse imipenem resistance phenotypes. Finer resolution evaluation of this clade revealed the presence of a 38.6 kbp conserved chromosomal region found exclusively in imipenem-susceptible isolates. This region appears to host several HTH-type DNA binding transcriptional regulators and transporter genes. Imipenem-susceptible isolates from this clade also carried two mutually exclusive plasmids that contain genes previously known to be specific to imipenem-susceptible isolates. Our analysis demonstrates the utility of using whole genomes for genotype-phenotype correlations in the context of antibiotic resistance and provides several new hypotheses for future research.

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Imipenem Resistance Mediated by blaOXA-913 Gene in Pseudomonas aeruginosa

Antibiotics ◽

10.3390/antibiotics10101188 ◽

2021 ◽

Vol 10 (10) ◽

pp. 1188

Author(s):

Dong-Chan Moon ◽

Abraham Fikru Mechesso ◽

Hee-Young Kang ◽

Su-Jeong Kim ◽

Ji-Hyun Choi ◽

...

Keyword(s):

Pseudomonas Aeruginosa ◽

Whole Genome Sequencing ◽

Genome Sequencing ◽

Resistance Genes ◽

Carbapenem Resistance ◽

Rrna Genes ◽

Whole Genome ◽

Translation Initiation Region ◽

Broth Microdilution Method ◽

Imipenem Resistance

Treatment of infectious diseases caused by carbapenem-resistant Pseudomonas aeruginosa is becoming a greater challenge. This study aimed to identify the imipenem resistance mechanism in P. aeruginosa isolated from a dog. Minimum Inhibitory Concentration (MIC) was determined by the broth microdilution method according to the Clinical and Laboratory Standards Institute recommendations. We performed polymerase chain reaction and whole-genome sequencing to detect carbapenem resistance genes. Genomic DNA of P. aeruginosa K19PSE24 was sequenced via the combined analysis of 20-kb PacBio SMRTbell and PacBio RS II. Peptide-Peptide Nucleic Acid conjugates (P-PNAs) targeting the translation initiation region of blaOXA-913 were synthesized. The isolate (K19PSE24) was resistant to imipenem and piperacillin/tazobactam yet was susceptible to most of the tested antimicrobials. Whole-genome sequencing revealed that the K19PSE24 genome comprised a single contig amounting to 6,815,777 base pairs, with 65 tRNA and 12 rRNA genes. K19PSE24 belonged to sequence type 313 and carried the genes aph(3)-IIb, fosA, catB7, crpP, and blaOXA-913 (an allele deposited in GenBank but not described in the literature). K19PSE24 also carried genes encoding for virulence factors (exoenzyme T, exotoxin A, and elastase B) that are associated with adhesion, invasion, and tissue lysis. Nevertheless, we did not detect any of the previously reported carbapenem resistance genes. This is the first report of the blaOXA-913 gene in imipenem-resistant P. aeruginosa in the literature. Notably, no viable colonies were found after co-treatment with imipenem (2 µg/mL) and either of the P-PNAs (12.5 µM or 25 µM). The imipenem resistance in K19PSE24 was primarily due to blaOXA-913 gene carriage.

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Identification of Antibiotic Resistance Genes in Multi-Drug Resistant Acinetobacter Baumannii Clinical Isolates of Iraqi Patients (Zq Strains), Using Whole-Genome Sequencing

International Journal of Pharmaceutical Quality Assurance ◽

10.25258/ijpqa.10.4.20 ◽

2019 ◽

Vol 10 (04) ◽

pp. 670-680

Author(s):

Zainab J Qasim ◽

Haider Sabah Kadhim ◽

Ahmed Sahib Abdulamir

Keyword(s):

Antimicrobial Resistance ◽

Acinetobacter Baumannii ◽

Genome Sequence ◽

Resistance Genes ◽

Genetic Material ◽

Whole Genome ◽

Drug Resistant ◽

Intrinsic Resistance ◽

Antimicrobial Resistance Genes ◽

Recent Emergence

Background: The recent emergence of multidrug resistance (MDR) in Acinetobacter baumannii (A. baumannii ) has raised concern in health care settings in Iraq. This is the first report of the whole genome sequence of A. baumannii ZQ isolated from Iraqi patients. To better comprehend the repertoire of MDR genetic elements and organization, we compared the genome sequences of eight extended drug-resistant (XDR) and two less drug-resistant A. baumannii ZQ strains with that of other completely sequenced A. baumannii from divergent worldwide distributed isolates.Results: In consistence with their phenotypic antimicrobial resistance profiles, ZQ genomes harbors high to moderate numbers of genetic determinants, including β-lactamases, aminoglycoside-modifying enzymes, efflux pumps, modifications of target sites. Several strains showed nearly identical genome sequence, frequent structural variation was detected even between the closely related strains.Conclusion: In general, the shorter the genetic distance among strains, the less insertion/deletion events proceed. However, frequent genomic changes was observed even inside the closely related strains of A. baumannii. Antimicrobial resistance genes are likely to be the target accumulating such variations, suggesting that the resistance elements respond actively to the selection pressure in the hospital setting. Besides the lateral acquisition of genetic material from resistant bacterial strains, the drastic issues is associated with continuous presence of intrinsic resistance genes in the genome of A. baumannii, which are ready to be boosted by exposure to sub-inhibitory levels of the antibiotics in the environment and might also play an important role in the evolution of resistance to the new derivatives of different antibiotic classes.

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WGS based analysis of acquired antimicrobial resistance in human and non-human Acinetobacter baumannii isolates from a German perspective

BMC Microbiology ◽

10.1186/s12866-021-02270-7 ◽

2021 ◽

Vol 21 (1) ◽

Author(s):

Gamal Wareth ◽

Christian Brandt ◽

Lisa D. Sprague ◽

Heinrich Neubauer ◽

Mathias W. Pletz

Keyword(s):

Antimicrobial Resistance ◽

Acinetobacter Baumannii ◽

Genome Sequencing ◽

Resistance Genes ◽

In Silico ◽

Acquired Resistance ◽

Whole Genome ◽

Chromosomal Gene ◽

Milk Samples ◽

Carbapenemase Gene

Abstract Background Acinetobacter baumannii ability to develop and acquire resistance makes it one of the most critical nosocomial pathogens globally. Whole-genome sequencing (WGS) was applied to identify the acquired or mutational variants of antimicrobial resistance (AMR) genes in 85 German A. baumannii strains utilizing Illumina technology. Additionally, the whole genome of 104 German isolates deposited in the NCBI database was investigated. Results In-silico analysis of WGS data revealed wide varieties of acquired AMR genes mediating resistance mostly to aminoglycosides, cephalosporins, carbapenems, sulfonamides, tetracyclines and macrolides. In the 189 analyzed genomes, the ant (3″)-IIa conferring resistance to aminoglycosides was the most frequent (55%), followed by blaADC.25 (38.6%) conferring resistance to cephalosporin, blaOXA-23 (29%) and the blaOXA-66 variant of the intrinsic blaOXA-51-likes (26.5%) conferring resistance to carbapenems, the sul2 (26%) conferring resistance to sulfonamides, the tet. B (19.5%) conferring resistance to tetracycline, and mph. E and msr. E (19%) conferring resistance to macrolides. blaTEM variants conferring resistance to cephalosporins were found in 12% of genomes. Thirteen variants of the intrinsic blaOXA-51 carbapenemase gene, blaOXA-510 and blaADC-25 genes were found in isolates obtained from dried milk samples. Conclusion The presence of strains harboring acquired AMR genes in dried milk raises safety concerns and highlights the need for changes in producing dried milk. Acquired resistance genes and chromosomal gene mutation are successful routes for disseminating AMR determinants among A. baumannii. Identification of chromosomal and plasmid-encoded AMR in the genome of A. baumannii may help understand the mechanism behind the genetic mobilization and spread of AMR genes.

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2429. Whole-genome Sequencing of Healthcare-Onset C. difficile Infection (HO-CDI) Cases Shows Widespread Presence of Antimicrobial Resistance Genes

Open Forum Infectious Diseases ◽

10.1093/ofid/ofz360.2107 ◽

2019 ◽

Vol 6 (Supplement_2) ◽

pp. S839-S840

Author(s):

Jae Hyun Shin ◽

Deiziane Viana da Silva Costa ◽

Hardik Parikh ◽

Katie E Barry ◽

Amy J Mathers ◽

...

Keyword(s):

Antimicrobial Resistance ◽

Resistance Genes ◽

Susceptibility Testing ◽

De Novo ◽

Antimicrobial Treatment ◽

Small Sample ◽

Binary Toxin ◽

Future Research ◽

Toxin Gene ◽

Antimicrobial Resistance Genes

Abstract Background Clostridioides difficile infection (CDI) is the most common pathogen to cause healthcare-associated infections. Unlike some other bacterial pathogens antimicrobial treatment is seldom based on culture with susceptibility testing with infrequent surveillance for antimicrobial resistance. We have evaluated healthcare-onset CDI (HO-CDI) for both transmission and antimicrobial resistance emergence. Methods We identified cases of HO-CDI diagnosed by PCR within a 3 month period (October 1/2018–December 31/2018) at University of Virginia Health System with overlapping stays in the same inpatient units with other HO-CDI. Chart review of all cases was performed. C. difficile was cultured from stool, then DNA was extracted and underwent sequencing on Illumina Miseq platform. Antimicrobial resistance genes were screened using NCBI’s AMRFinder tool from the de-novo assembled contigs using SPAdes. All the C. difficile isolates underwent antibiotic susceptibility testing. Results Eleven patients were identified with overlapping stays from 5 units. Mean age was 64 years and 63.6% were female. 36.3% of cases were severe CDI with one case of fulminant CDI. There was one recurrence within 90 days (9.1%). Patients were treated with PO vancomycin (72.7%) or IV metronidazole and PO vancomycin (27.3%), none were treated with metronidazole alone. None of the hospital strains were genetically related. There were two isolates with binary toxin gene (cdtB), one ribotype 027 (CD196) and one ribotype 078 (M120). Ninety-one percent of isolates had vanG-like gene cluster and vanZ1 originally identified in Enterococcus sp. erm(B), tet(M), and cfr(C) genes were also detected in several strains. All isolates were susceptible to vancomycin, metronidazole, and tigecycline. There was one strain with moxifloxacin resistance associated with the presence of erm(B) gene. None of the isolates were susceptible to clindamycin. Conclusion There were no widely circulating clones or direct transmissions found in this small sample of HO-CDI cases at our hospital. Like others have we demonstrate carriage of many vanG/Z genes without conferring phenotypic resistance to vancomycin. The origin and function of Van genes in C. difficile could be an area of future research. Disclosures All authors: No reported disclosures.

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Accumulation of Antibiotic Resistance Genes in Carbapenem-Resistant Acinetobacter baumannii Isolates Belonging to Lineage 2, Global Clone 1, from Outbreaks in 2012–2013 at a Tehran Burns Hospital

mSphere ◽

10.1128/msphere.00164-20 ◽

2020 ◽

Vol 5 (2) ◽

Cited By ~ 2

Author(s):

Masoumeh Douraghi ◽

Johanna J. Kenyon ◽

Parisa Aris ◽

Mahla Asadian ◽

Sedighe Ghourchian ◽

...

Keyword(s):

Middle East ◽

Acinetobacter Baumannii ◽

Resistance Genes ◽

Carbapenem Resistance ◽

Antibiotic Resistant ◽

Content Type ◽

Carbapenem Resistant ◽

East Region ◽

Lineage 2 ◽

Middle East Region

ABSTRACT The worldwide distribution of carbapenem-resistant Acinetobacter baumannii (CRAB) has become a global concern, particularly in countries where antibiotic prescription is not tightly regulated. However, knowledge of the genomic aspects of CRAB from many parts of the world is still limited. Here, 50 carbapenem-resistant A. baumannii isolates recovered at a single hospital in Tehran, Iran, during several outbreaks in 2012 and 2013 were found to be resistant to multiple antibiotics. They were examined using PCR mapping and multilocus sequence typing (MLST). All Iranian strains belonged to sequence type 328 in the Institut Pasteur MLST scheme (ST328IP), a single-locus variant of ST81IP, and all Iranian strains contained two carbapenem resistance genes, oxa23 and oxa24. The oxa23 gene is in the transposon Tn2006 in AbaR4, which interrupts the chromosomal comM gene. Phylogenetic analysis using whole-genome sequence (WGS) data for 9 isolates showed that they belonged to the same clade, designated the ST81/ST328 clade, within lineage 2 of global clone 1 (GC1). However, there were two groups that included either KL13 or KL18 at the K locus (KL) for capsular polysaccharide synthesis and either a tet39 or an aadB resistance gene, respectively. The genetic context of the resistance genes was determined, and the oxa24 (OXA-72 variant) and tet39 (tetracycline resistance) genes were each in a pdif module in different plasmids. The aadB gene cassette (which encodes gentamicin, kanamycin, and tobramycin resistance) was harbored by pRAY*, and the aphA6 gene (which encodes amikacin resistance) and sul2 gene (which encodes sulfamethoxazole resistance) were each harbored by a different plasmid. The sequences obtained here will underpin future studies of GC1 CRAB strains from the Middle East region. IMPORTANCE Carbapenem-resistant Acinetobacter baumannii strains are among the most critical antibiotic-resistant bacteria causing hospital-acquired infections and treatment failures. The global spread of two clones has been responsible for the bulk of the resistance, in particular, carbapenem resistance. However, there is a substantial gap in our knowledge of which clones and which specific lineages within each clone are circulating in many parts of the world, including Africa and the Middle East region. This is the first genomic analysis of carbapenem-resistant A. baumannii strains from Iran. All the isolates, from a single hospital, belonged to lineage 2 of global clone 1 (GC1) but fell into two groups distinguished by genes in the locus for capsule biosynthesis. The analysis suggests a potential origin of multiply antibiotic-resistant lineage 2 in the Middle East region and highlights the ongoing evolution of carbapenem-resistant GC1 A. baumannii strains. It will enhance future studies on the local and global GC1 population structure.

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Whole genome sequence analysis of antimicrobial resistance genes, multilocus sequence types and plasmid sequences in ESBL/AmpC Escherichia coli isolated from broiler caecum and meat

International Journal of Food Microbiology ◽

10.1016/j.ijfoodmicro.2019.108361 ◽

2020 ◽

Vol 315 ◽

pp. 108361 ◽

Cited By ~ 1

Author(s):

M. Päivärinta ◽

S. Latvio ◽

M. Fredriksson-Ahomaa ◽

A. Heikinheimo

Keyword(s):

Escherichia Coli ◽

Sequence Analysis ◽

Antimicrobial Resistance ◽

Genome Sequence ◽

Resistance Genes ◽

Whole Genome Sequence ◽

Whole Genome ◽

Genome Sequence Analysis ◽

Antimicrobial Resistance Genes ◽

Sequence Types

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Molecular and Phenotypic Characteristics ofEscherichia coliIsolates from Farmed Minks in Zhucheng, China

BioMed Research International ◽

10.1155/2019/3917841 ◽

2019 ◽

Vol 2019 ◽

pp. 1-12 ◽

Cited By ~ 3

Author(s):

Jianhua Qiu ◽

Zhiyu Jiang ◽

Zijing Ju ◽

Xiaonan Zhao ◽

Jie Yang ◽

...

Keyword(s):

Antibiotic Resistance ◽

Resistance Genes ◽

Large Scale ◽

Clonal Complex ◽

Sensitivity Testing ◽

Class 1 Integron ◽

Pcr Screening ◽

E Coli ◽

Antimicrobial Resistance Genes ◽

Class 1

In this study, the prevalence, phenotypes, and clonal relationships ofEscherichia coli(E. coli) strains isolated from minks were investigated. In July 2017, a total of 62 fresh faecal swab samples were randomly collected from one large-scale mink farm in Zhucheng, Shandong Province, China. In all the samples, 50E. colistrains were isolated and then assigned to serotyping, antimicrobial susceptibility test, detection of antimicrobial resistance genes and the Class 1 integrons, and multilocus sequence typing (MLST). Four pathogenic serotypes were identified among all the isolates, while the most common serotype was enterohemorrhagicE. coliO104:H4 (6.0 %). Antimicrobial sensitivity testing revealed that most isolates were susceptible to cefoxitin (96.0 %) and amikacin (82.0 %), while most isolates were resistant to ampicillin (92.0 %) and tetracycline (90.0 %). An analysis of the nucleotide sequences revealed that 7 isolates (14.0%) carried 4 types of Class 1 integron cassette, includingdfrA27+aadA2+qnrA(57.1%),dfrA17+aadA5(14.3%),dfrA12+aadA2(14.3%), anddfrA1+aadA1(14.3%). PCR screening showed that 14 antibiotic resistance genes were presented in 50 isolates, while the most prevalent resistance gene wasqnrS, which was detected in 60.0 % of isolates, followed bysul2(40.0%) andoqxA(38.0%). MLST analysis showed that 32 sequence types (STs) were identified, while ST46 was the predominant genotype among all isolates. Clonal complex 3 (CC3) was dominant. Compared with 340 humanE. coliSTs reported in China, the ST10 clonal complex, known as the largest human clonal complex, was also found in the 50 minkE. coliisolates. Meanwhile, mink-derived strain ST206 formed a new clonal complex, CC206, which was different from human ST strains. Our results showed that farmed minks could be reservoirs of antimicrobial-resistantE. coliwith Class 1 integron cassettes and resistance genes, which were likely to pose a threat to public health. Therefore, continuous inspections and monitoring ofE. coliin minks are essential for detecting and controlling emergingE. coliwith different serovars as well as antibiotic resistance.

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Research and Technological Advances Regarding the Study of the Spread of Antimicrobial Resistance Genes and Antimicrobial-Resistant Bacteria Related to Animal Husbandry

International Journal of Environmental Research and Public Health ◽

10.3390/ijerph16244896 ◽

2019 ◽

Vol 16 (24) ◽

pp. 4896

Author(s):

Na Li ◽

Chong Liu ◽

Zhiguo Zhang ◽

Hongna Li ◽

Tingting Song ◽

...

Keyword(s):

Antimicrobial Resistance ◽

Resistance Genes ◽

High Throughput Sequencing ◽

Animal Husbandry ◽

Control Measures ◽

Research Progress ◽

Future Research ◽

Resistant Bacteria ◽

Technological Advances ◽

Antimicrobial Resistance Genes

The extensive use of antimicrobials in animal farms poses serious safety hazards to both the environment and public health, and this trend is likely to continue. Antimicrobial resistance genes (ARGs) are a class of emerging pollutants that are difficult to remove once introduced. Understanding the environmental transfer of antimicrobial-resistant bacteria (ARB) and ARGs is pivotal for creating control measures. In this review, we summarize the research progress on the spread and detection of ARB and ARG pollution related to animal husbandry. Molecular methods such as high-throughput sequencing have greatly enriched the information about ARB communities. However, it remains challenging to delineate mechanisms regarding ARG induction, transmission, and tempo-spatial changes in the whole process, from animal husbandry to multiple ecosystems. As a result, future research should be more focused on the mechanisms of ARG induction, transmission, and control. We also expect that future research will rely more heavily on metagenomic -analysis, metatranscriptomic sequencing, and multi-omics technologies

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Assessing the Genetic Diversity of Austrian Corynebacterium diphtheriae Clinical Isolates, 2011-2019

Journal of Clinical Microbiology ◽

10.1128/jcm.02529-20 ◽

2020 ◽

Author(s):

Justine Schaeffer ◽

Steliana Huhulescu ◽

Anna Stoeger ◽

Franz Allerberger ◽

Werner Ruppitsch

Keyword(s):

Genetic Diversity ◽

Whole Genome Sequencing ◽

Genome Sequencing ◽

Resistance Genes ◽

Diphtheria Toxin ◽

Corynebacterium Diphtheriae ◽

Toxin Gene ◽

Whole Genome ◽

Skin Infections ◽

Antimicrobial Resistance Genes

Background: Diphtheria is a vaccine preventable disease with a high potential for re-emergence. One of its causative agent is Corynebacterium diphtheriae, some strains producing the diphtheria toxin. From 2011 to 2019, 57 clinical C. diphtheriae strains were isolated in Austria, either from the respiratory track or from skin infections. Objectives: The aim of the study was to investigate the genetic diversity of these C. diphtheriae isolates using whole genome sequencing. Methods: Isolates were characterized by genome wide comparison using single nucleotide polymorphism or core genome multilocus sequence typing, and by searching sequence data for antimicrobial resistance genes and genes involved in diphtheria toxin production. Results: Genetic diversity between the isolates was high, with no clear distribution over time or place. C. belfantii isolates were separated from other strains, and were strongly associated with respiratory infections (OR = 57). Two clusters, limited in time and space, were identified. Almost 40% of strains carried resistance genes against tetracycline or sulfonamides, mostly from skin infections. Microbiological tests showed that 55% of isolates were resistant to penicillin, but did not carry genes conferring β-lactam resistance. Diphtheria toxin gene with no non-synonymous mutation was found in three isolates only. Conclusion: This study showed that sequencing can provide valuable information complementing routine microbiological and epidemiological investigations. It allowed us to identify unknown clusters, evaluate antimicrobial resistances more broadly and support toxigenicity results obtained by PCR. For these reasons, C. diphtheriae surveillance could strongly benefit from the routine implementation of whole genome sequencing.

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