Development of a quadruple qRT-PCR assay for simultaneous identification of hypervirulent and carbapenem-resistant Klebsiella pneumoniae.

IMPORTANCE: Globally, the increasing number of hypervirulent Klebsiella pneumoniae (hvKp) and carbapenem-resistant Kp (CR-Kp) infections poses a huge public health challenge with high morbidity and mortality. Worrisomely, due to the mobility of elements carrying virulence and drug-resistance genes, the increasing prevalence of CR-hvKp has also been found with an overwhelming mortality rate in recent years. However, the current detection methods for hvKp and CR-Kp have many disadvantages, such as long turnaround time, complex operation, low sensitivity, and specificity. Herein, a more sensitive, rapid, single-reaction, and multiplex quantitative real-time PCR was developed and validated to differentiate the circulating lineages of Kp with excellent performance in sensitivity and specificity, providing a useful tool for the differential diagnosis and the surveillance of the circulating Kp.

High-Throughput Experimentation and Machine Learning-Assisted Optimization of Iridium-Catalyzed Cross-Dimerization of Sulfoxonium Ylides.

A novel and convenient approach that combines high-throughput experimentation (HTE) with machine learning (ML) technologies to achieve the first selective cross-dimerization of sulfoxonium ylides via iridium catalysis is presented. A variety of valuable amide-, ketone-, ester-, and N-heterocycle-substituted unsymmetrical E-alkenes are synthesized in good yields with high stereoselectivities. This mild method avoids the use of diazo compounds and is characterized by simple operation, high step-economy, and excellent chemoselectivity and functional group compatibility. The combined experimental and computational studies identify an amide-sulfoxonium ylide as a carbene precursor. Furthermore, a comprehensive exploration of the reaction space is also performed (600 reactions) and a machine learning model for reaction yield prediction has been constructed.

A deep learning framework for accurate reaction prediction and its application on high-throughput experimentation data.

In recent years, it has been seen that artificial intelligence (AI) starts to bring revolutionary changes to chemical synthesis. However, the lack of suitable ways of representing chemical reactions and the scarceness of reaction data has limited the wider application of AI to reaction prediction. Here, we introduce a novel reaction representation, GraphRXN, for reaction prediction. It utilizes a universal graph-based neural network framework to encode chemical reactions by directly taking two-dimension reaction structures as inputs. The GraphRXN model was evaluated by three publically available chemical reaction datasets and gave on-par or superior results compared with other baseline models. To further evaluate the effectiveness of GraphRXN, wet-lab experiments were carried out for the purpose of generating reaction data. GraphRXN model was then built on high-throughput experimentation data and a decent accuracy (R2 of 0.712) was obtained on our in-house data. This highlights that the GraphRXN model can be deployed in an integrated workflow which combines robotics and AI technologies for forward reaction prediction.

Assessment of antibody responses against SARS-CoV-2 in unvaccinated individuals and vaccinees from Omicron-BA.2 infection in Zhaoqing, Guangdong Province, China.

Currently, the majority of the global population has been vaccinated with the COVID-19 vaccine, and characterization studies of antibodies in vivo from Omicron breakthrough infection and naive infection populations are urgently needed to provide pivotal clues about accurate diagnosis, treatment, and next-generation vaccine design against SARS-CoV-2 infection. We showed that after infection with Omicron-BA.2, the antibody levels of specific IgM against the Wuhan strain and specific IgG against Omicron were not significantly elevated within 27 days of onset. Interestingly, in this study, the levels of humoral immunity against Omicron-specific IgM were significantly increased after breakthrough infection, suggesting that the detection of Omicron-specific IgM antibodies can be used as a test criterion of Omicron breakthrough infection. In addition, we observed that serums from unvaccinated individuals and the majority of vaccinated infections possessed only low or no neutralizing activity against Omicron at the onset of Omicron breakthrough infections, and at the later stage of Omicron-BA.2 breakthrough infection, levels of neutralization antibody against the Wuhan and Omicron strains were elevated in infected individuals. The findings of this study provide important clues for the diagnosis of Omicron breakthrough infections, antibody characterization studies and vaccine design against COVID-19.

Copper-Catalyzed Nitrogen Atom Transfer to Isoquinolines via C-N Triple Bond Cleavage and Three-Component Cyclization.

A copper(I)-catalyzed tandem reaction of 2-bromoaryl ketones, terminal alkynes, and CH3CN is developed, which combines N atom transfer and three-component [3 + 2 + 1] cyclization, and efficiently produces densely functionalized isoquinolines in a facile, highly selective, and general manner. In the reaction, the formation of aromatic C-N bonds along with the complete C-N triple bond cleavage is first realized; Cu(III)-acetylide species might serve as the intermediates, which allow highly selective 6-endo-dig cyclization.

Copper-Catalyzed 6-endo-dig Cyclization-Coupling of 2-Bromoaryl Ketones and Terminal Alkynes toward Naphthyl Aryl Ethers in Water.

The cyclization-coupling reaction of 2-bromoaryl ketones and terminal alkynes is first realized by copper catalysis, which produces polyfunctional naphthyl aryl ethers in moderate to excellent yields with broad substrate scope and good functional group tolerance. This reaction proceeds via 6-endo-dig cyclization and C(sp2)-O coupling using green H2O as the unique solvent and 5-bromopyrimidin-2-amine as the critical additive. Mechanistically, a unique Cu(III)-acetylide probably is the key intermediate, which allows exclusive 6-endo-dig selectivity.

Molecular Epidemiological Analysis of ST11-K64 Extensively Drug-Resistant Klebsiella pneumoniae Infections Outbreak in Intensive Care and Neurosurgery Units Based on Whole-Genome Sequencing.

Klebsiella pneumoniae (Kp) is the primary causative bacteria for nosocomial infections and hospital outbreaks. In particular, extensively drug-resistant K. pneumoniae (XDRKp) causes severe clinical infections in hospitalized patients. Here, we used pulsed-field gel electrophoresis (PFGE), drug susceptibility tests, and the whole-genome sequencing (WGS) technology to examine genetic relatedness and phenotypic traits of the strains isolated during an outbreak period. Based on PFGE, a distinct clones cluster comprised of eight XDRKp was observed. These strains were confirmed as ST11-K64 via multiple-locus sequence typing database of Kp. The strains also had genes related to the regulation of biofilm biosynthesis (type 1 & 3 fimbriae, type IV pili biosynthesis, RcsAB, and type VI secretion system) and multiple drug resistance (ß-lactamase and aminoglycoside antibiotic resistance). WGS data based on core-single nucleotide polymorphisms and epidemiological investigation showed that the neurosurgery unit was likely the source of the outbreak, the strain was likely to have been transmitted to the ICU through patients. In addition, the two highly probable transmission routes were in the ICU (exposure through shared hospital beds) and the neurosurgery units (all cases were treated by the same rehabilitation physician and were most likely infected during the physical therapy). Notably, the bed mattress had played a crucial transmission role of this outbreak, served as a pathogen reservoir.

Cu(i) catalysis for selective condensation/bicycloaromatization of two different arylalkynes: direct and general construction of functionalized C-N axial biaryl compounds.

Selective condensation/bicycloaromatization of two different arylalkynes is firstly developed under ligand-free copper(i)-catalysis, which allows the direct synthesis of C-N axial biaryl compounds in high yields with excellent selectivity and functional group tolerance. Due to the critical effects of Cu(i) catalyst and HFIP, many easily occurring undesired reactions are suppressed, and the coupled five-six aromatic rings are constructed via the selective formation of two C(sp2)-N(sp2) bonds and four C(sp2)-C(sp2) bonds. The achievement of moderate enantioselectivity verifies its potential for the simplest asymmetric synthesis of atropoisomeric biaryls. Western blotting demonstrated that the newly developed compounds are promising targets in biology and pharmaceuticals. This unique reaction can construct structurally diverse C-N axial biaryl compounds that have never been reported by other methods, and might be extended to various applications in materials, chemistry, biology, and pharmaceuticals.

Cu-Catalyzed Oxidative Thioesterification of Aroylhydrazides with Disulfides.

An alternative thioesterification reaction via copper-catalyzed oxidative coupling of readily available aroylhydrazides with disulfides is developed, in which oxidative expulsion of N2 overcomes the activation barrier between the carboxylic acid derivatives and the products. The reaction produces various thioesters in good to excellent yields with good functional group tolerance. In the reaction, stable and easily available aroylhydrazides are used as acyl sources and the relatively odorless disulfides are used as S sources. Mechanistic investigations demonstrate that the reaction of copper salt and oxidant (NH4)2S2O8 allows for achievement of tandem processes, including deprotonation, free-radical-mediated denitrogenation, and C-S bond formation.

Selective Oxidative Cleavage of 3-Methylindoles with Primary Amines Affording Quinazolinones.

A selective functionalization of C-C═C bonds toward N-C═O bonds is realized by an n-Bu4NI-catalyzed reaction of 3-methylindoles with primary amines using TBHP as the unique oxidant. The systematic process involves oxygenation, nitrogenation, ring-opening, and recyclization, affording a broad range of quinazolinones in good to excellent yields.

General Oxidative Aryl C-P Bond Formation through Palladium-Catalyzed Decarbonylative Coupling of Aroylhydrazides with P(O)H Compounds.

Oxidative C-C/P-H cross-coupling is achieved via Pd-catalyzed decarbonylative cross-coupling of aroylhydrazides with P(O)H compounds. The unique cooperative reaction system, especially the Brønsted acid and bidentate phosphine ligand, allows the selective activation of the inert C-C bond and the suppression of the undesired oxidation and coordination of >P(O)-H compounds, leading to a general oxidative synthesis of aryl phosphorus compounds from easily available substrates.

Cu(I)-Catalyzed 6- endo-dig Cyclization of Terminal Alkynes, 2-Bromoaryl Ketones, and Amides toward 1-Naphthylamines: Applications and Photophysical Properties.

Functional group substituted 1-naphthylamines, especially N-methylated ones, play important roles in numerous chemical and biological processes. However, these compounds' general and step-economic syntheses are highly limited, which seriously restricts efforts to improve the properties and develop new functions for this kind of compound. In this report, we describe the development of an efficient, convenient, and general method for the synthesis of valuable functionalized 1-naphthylamines directly from readily available terminal alkynes, 2-bromoaryl ketones, and amides via Cu(I)-catalyzed benzannulation in a green solvent (i.e., water) under Pd- and ligand-free conditions. A total of 82 functionalized 1-naphthylamines, especially synthetically and biologically useful N-methylated compounds, are synthesized in isolated yields up to 95%. Some unique features of the reaction are as follows: (1) exclusive 6 -endo-dig selectivity, (2) ready incorporation of a broad range of functional groups directly from easily available substrates, and (3) amides that can be used as aminating agents and that are excellent alternatives to toxic and/or odorous amines. Due to facile tuning of functional groups for the reaction, the products possess good electronic donor-acceptor structures and exhibit intriguing photophysical properties, such as tunable and polarity-sensitive fluorescence emission and large Stokes shifts (up to 258 nm). Utilizing the products' unique polarity-sensitive fluorescence response, we successfully applied the 1-naphthylamine derivatives, such as compound 91, to image lipid droplets (LDs) and monitor cellular LDs growth. The previously mentioned advantages of this methodology, along with the mild conditions, simple operation, and scalable synthesis, may allow this novel reaction to be extended to varied applications in chemistry, biology, and materials science.

Palladium-Catalyzed Regio- and Stereoselective Coupling-Addition of Propiolates with Arylsulfonyl Hydrazides: A Pattern for Difunctionalization of Alkynes.

A new pattern for difunctionalization of alkynes via a palladium-catalyzed regio- and stereoselective coupling-addition of propiolates with arylsulfonyl hydrazides is disclosed. The approach enables the synthesis of various highly functionalized ( E)-vinylsulfones in satisfactory yields. Arylsulfonyl hydrazides act as both aryl and sulfonyl sources via selective cleavage of Ar(C)-S and S-N bonds, which are simultaneously incorporated onto the terminal carbon atom of an alkyne molecule.

Cyclization of Ketones with Nitriles under Base: A General and Economical Synthesis of Pyrimidines.

A facile, general, and economical synthesis of diversely functionalized pyrimidines has been realized under basic conditions via the copper-catalyzed cyclization of ketones with nitriles. The reaction proceeds via a novel pathway involving the nitriles acting as electrophiles and consecutive C-C bond and two C-N bond formations and shows broad substrate scope and good tolerance of many important functional groups. This strategy represents a new platform for constructing pyrimidine structures.

Copper Catalysis for Selective Heterocoupling of Terminal Alkynes.

A Cu-catalyzed selective aerobic heterocoupling of terminal alkynes is disclosed, which enables the synthesis of a broad range of unsymmetrical 1,3-diynes in good to excellent yields. The results disprove the long-held belief that homocouplings are exclusively favored in the Glaser-Hay reaction.