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Novel rapidly diversifiable antimicrobial RNA polymerase switch region inhibitors with confirmed mode of action in Haemophilus influenzae.

ABSTRACT: A series of inhibitors with a squaramide core was synthesized following its discovery in a high-throughput screen for novel inhibitors of a transcription-coupled translation assay using Escherichia coli S30 extracts. The inhibitors were inactive when the plasmid substrate was replaced with mRNA, suggesting they interfered with transcription. This was confirmed by their inhibition of purified E. coli RNA polymerase. The series had antimicrobial activity against efflux-negative strains of E. coli and Haemophilus influenzae. Like rifampin, the squaramides preferentially inhibited synthesis of RNA and protein over fatty acids, peptidoglycan, and DNA. However, squaramide-resistant mutants were not cross-resistant to rifampin. Nine different mutations were found in parts of rpoB or rpoC that together encode the so-called switch region of RNA polymerase. This is the binding site of the natural antibiotics myxopyronin, corallopyronin, and ripostatin and the drug fidaxomicin. Computational modeling using the X-ray crystal structure of the myxopyronin-bound RNA polymerase of Thermus thermophilus suggests a binding mode of these inhibitors that is consistent with the resistance mutations. The squaramides are the first reported non-natural-product-related, rapidly diversifiable antibacterial inhibitors acting via the switch region of RNA polymerase.

SUBMITTER: Buurman ET

PROVIDER: S-EPMC3458661 | biostudies-literature | 2012 Oct

REPOSITORIES: biostudies-literature

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Novel rapidly diversifiable antimicrobial RNA polymerase switch region inhibitors with confirmed mode of action in Haemophilus influenzae.

Buurman Ed T ET Foulk Melinda A MA Gao Ning N Laganas Valerie A VA McKinney David C DC Moustakas Demetri T DT Rose Jonathan A JA Shapiro Adam B AB Fleming Paul R PR

Journal of bacteriology 20120727 20

A series of inhibitors with a squaramide core was synthesized following its discovery in a high-throughput screen for novel inhibitors of a transcription-coupled translation assay using Escherichia coli S30 extracts. The inhibitors were inactive when the plasmid substrate was replaced with mRNA, suggesting they interfered with transcription. This was confirmed by their inhibition of purified E. coli RNA polymerase. The series had antimicrobial activity against efflux-negative strains of E. coli ...[more]

PMID: 22843845

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Project description:Antibiotics are important for the treatment and prevention of invasive Haemophilus influenzae disease. Reduced susceptibility to clinically relevant drugs, except ampicillin, has been uncommon in the United States. Susceptibility of 700 invasive H. influenzae isolates, collected through population-based surveillance during 2016, was assessed for 15 antibiotics using broth microdilution, according to the CLSI guidelines; a subset of 104 isolates were also assessed for rifampin susceptibility using Etest. Genomes were sequenced to identify genes and mutations known to be associated with reduced susceptibility to clinically relevant drugs. A total of 508 (72.6%) had reduced susceptibility to at least one antibiotic and more than half of the isolates exhibited reduced susceptibility to only one (33.6%) or two (21.6%) antibiotic classes. All tested isolates were susceptible to rifampin, a chemoprophylaxis agent, and <1% (n = 3) of isolates had reduced susceptibility to third generation cephalosporins, which are recommended for invasive disease treatment. In contrast, ampicillin resistance was more common (28.1%) and predominantly associated with the detection of a β-lactamase gene; 26.2% of isolates in the collection contained either a TEM-1 or ROB-1 β-lactamase gene, including 88.8% of ampicillin-resistant isolates. β-lactamase negative ampicillin-resistant (BLNAR) isolates were less common and associated with ftsI mutations; resistance to amoxicillin-clavulanate was detected in <2% (n = 13) of isolates. The proportion of reduced susceptibility observed was higher among nontypeable H. influenzae and serotype e than other serotypes. US invasive H. influenzae isolates remain predominantly susceptible to clinically relevant antibiotics except ampicillin, and BLNAR isolates remain uncommon. IMPORTANCE Antibiotics play an important role for the treatment and prevention of invasive Haemophilus influenzae disease. Antimicrobial resistance survey of invasive H. influenzae isolates collected in 2016 showed that the US H. influenzae population remained susceptible to clinically relevant antibiotics, except for ampicillin. Detection of approximately a quarter ampicillin-resistant and β-lactamase containing strains demonstrates that resistance mechanisms can be acquired and sustained within the H. influenzae population, highlighting the continued importance of antimicrobial resistance surveillance for H. influenzae to monitor susceptibility trends and mechanisms of resistance.

Project description:BackgroundThe widespread use of antimicrobials and Haemophilus influenzae type b (Hib) vaccine worldwide has altered the epidemiological patterns of invasive H. influenzae. Nonetheless, little is currently known on the epidemiological characteristics of H. influenzae in Guiyang, Guizhou, China.ObjectiveTo determine the serotype distribution, antimicrobial resistance and Multilocus Sequence Typing (MLST) of H. influenzae in hospitalized patients in Guiyang City.MethodsA total of 196 clinical isolates from hospitalized patients were collected. Serotypes were determined according to the specific capsule gene, bexA, amplified by PCR. According to the guidelines of Clinical and Laboratory Standards Institute (CLSI) 2020 drug susceptibility tested, and the results determined. The chromogenic cephalosporin nitrocefin method was used to detect β-lactamase production, β-lactamase negative, ampicillin-resistant (BLNAR) strains were detected by PCR amplification and sequencing of the penicillin-binding protein 3 (PBP3) locus of ftsI. Multilocus Sequence Typing was performed for molecular typing.ResultsAll isolates studied were non-typeable H. influenzae (NTHi). Most patients originated from the pediatrics department (78.6%, 154/196), and suffered from lung with respiratory tract infection (pneumonia and bronchitis, 68.4%, 134/196). The resistance rates of ampicillin, cefaclor and azithromycin were 71.4% (140/196), 36.7% (72/196) and 34.2% (67/196), respectively. 40.3% (79/196) of strains were β-lactamase positive ampicillin-resistant (BLPAR). All BLPAR carried the TEM-1 gene. 9.2% (18/196) were β-lactamase negative ampicillin-resistant strains (BLNAR). The PBP3 mutation was detected in the ampicillin-resistant strains (n = 113), of which 18 belonged to group IIa. A total of 49 sequence types (ST) and 23 clonal complexes (CC) were detected, among which CC107 (ST107, n = 27; ST1002, n = 5; ST1218, n = 5) was the most frequent clonal complexes. BLPAR isolates mostly belonged to ST107 (20/79), while BLNAR was predominantly distributed in ST12 (5/18).ConclusionH. influenzae infections are predominately caused by genetically diverse NTHi among hospitalized patients in Guiyang. The prevalence of β-lactamase production and PBP3 mutation may contribute to the high local ampicillin resistance rate.

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Novel rapidly diversifiable antimicrobial RNA polymerase switch region inhibitors with confirmed mode of action in Haemophilus influenzae.

Publications

Novel rapidly diversifiable antimicrobial RNA polymerase switch region inhibitors with confirmed mode of action in Haemophilus influenzae.

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