Identification of distinct genotypes in circulating RSV A strains based on variants in the virus replication-associated genes.

Respiratory syncytial virus (RSV) is a common cause of respiratory infection that often leads to hospitalization of infected younger children and older adults. RSV is classified into two strains, A and B, each with several subgroups or genotypes. One issue with the definition of these subgroups is the lack of a unified method of identification or genotyping.

Identification of distinct genotypes in circulating RSV A strains based on variants on the virus replication-associated genes.

Respiratory syncytial virus is a common cause of respiratory infection that often leads to hospitalization of infected younger children and older adults. RSV is classified into two strains, A and B, each with several subgroups or genotypes. One issue with the definition of these subgroups is the lack of a unified method of identification or genotyping.

Streptothricin F is a bactericidal antibiotic effective against highly drug-resistant gram-negative bacteria that interacts with the 30S subunit of the 70S ribosome.

The streptothricin natural product mixture (also known as nourseothricin) was discovered in the early 1940s, generating intense initial interest because of excellent gram-negative activity. Here, we establish the activity spectrum of nourseothricin and its main components, streptothricin F (S-F, 1 lysine) and streptothricin D (S-D, 3 lysines), purified to homogeneity, against highly drug-resistant, carbapenem-resistant Enterobacterales (CRE) and Acinetobacter baumannii. For CRE, the MIC50 and MIC90 for S-F and S-D were 2 and 4 μM, and 0.

Genotypic investigation of a rotavirus cluster at a quaternary-care pediatric hospital.

Rotavirus (RV) was a common healthcare-associated infection prior to the introduction of the RV vaccine. Following widespread RV vaccination, healthcare-associated rotavirus cases are rare. We describe an investigation of a cluster of rotavirus infections in a pediatric hospital in which an uncommon genotype not typically circulating in the United States was detected.

Novel Assays for Molecular Detection of Severe Acute Respiratory Syndrome Coronavirus 2.

From the onset of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)/COVID-19 pandemic, there has been a major emphasis on molecular laboratory tests for the virus. Shortages in various testing supplies, the desire to increase testing capacity, and a push to make point-of-care or home-based testing available have fostered considerable innovation for SARS-CoV-2 molecular diagnostics, advancements likely to be applicable to other diagnostic uses. The authors attempt to cover some of the most compelling novel types of molecular assays or novel approaches in adapting established molecular methodologies for SARS-CoV-2 detection or characterization.

The convergent total synthesis and antibacterial profile of the natural product streptothricin F.

A convergent, diversity-enabling total synthesis of the natural product streptothricin F has been achieved. Herein, we describe the potent antimicrobial activity of streptothricin F and highlight the importance of a total synthesis that allows for the installation of practical divergent steps for medicinal chemistry exploits. Key features of our synthesis include a Burgess reagent-mediated 1,2--diamine installation, diastereoselective azidation of a lactam enolate, and a mercury(ii) chloride-mediated desulfurization-guanidination.

Pyrazole-Thiazole Core-Containing Analogs Exhibit Adjunctive Activity with Meropenem against Carbapenem-Resistant Enterobacteriaceae (CRE).

Pyrazole-thiazole core-containing compound KP-40 and 20 novel derivatives were designed and synthesized through traditional SAR analysis. These molecules displayed adjunctive activity with meropenem against Gram-negative bacteria evidenced by a range of fractional inhibitory concentration (FIC=0.5-0.

The Limit of Detection Matters: The Case for Benchmarking Severe Acute Respiratory Syndrome Coronavirus 2 Testing.

Background: Resolving the coronavirus disease 2019 (COVID-19) pandemic requires diagnostic testing to determine which individuals are infected with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). The current gold standard is to perform reverse-transcription polymerase chain reaction (PCR) on nasopharyngeal samples. Best-in-class assays demonstrate a limit of detection (LoD) of approximately 100 copies of viral RNA per milliliter of transport media.

SARS-CoV2 Testing: The Limit of Detection Matters.

Resolving the COVID-19 pandemic requires diagnostic testing to determine which individuals are infected and which are not. The current gold standard is to perform RT-PCR on nasopharyngeal samples. Best-in-class assays demonstrate a limit of detection (LoD) of ~100 copies of viral RNA per milliliter of transport media.

Large-scale, in-house production of viral transport media to support SARS-CoV-2 PCR testing in a multi-hospital healthcare network during the COVID-19 pandemic.

The COVID-19 pandemic has severely disrupted worldwide supplies of viral transport media (VTM) due to widespread demand for SARS-CoV-2 RT-PCR testing. In response to this ongoing shortage, we began production of VTM in-house in support of diagnostic testing in our hospital network. As our diagnostic laboratory was not equipped for reagent production, we took advantage of space and personnel that became available due to closure of the research division of our medical center.

Open Development and Clinical Validation Of Multiple 3D-Printed Sample-Collection Swabs: Rapid Resolution of a Critical COVID-19 Testing Bottleneck.

The SARS-CoV-2 pandemic has caused a severe international shortage of the nasopharyngeal swabs that are required for collection of optimal specimens, creating a critical bottleneck in the way of high-sensitivity virological testing for COVID-19. To address this crisis, we designed and executed an innovative, radically cooperative, rapid-response translational-research program that brought together healthcare workers, manufacturers, and scientists to emergently develop and clinically validate new swabs for immediate mass production by 3D printing. We performed a rigorous multi-step preclinical evaluation on 160 swab designs and 48 materials from 24 companies, laboratories, and individuals, and shared results and other feedback via a public data repository (http://github.

Large-Scale, In-House Production of Viral Transport Media To Support SARS-CoV-2 PCR Testing in a Multihospital Health Care Network during the COVID-19 Pandemic.

The COVID-19 pandemic has severely disrupted worldwide supplies of viral transport media (VTM) due to widespread demand for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) reverse transcription-PCR (RT-PCR) testing. In response to this ongoing shortage, we began production of VTM in-house in support of diagnostic testing in our hospital network. As our diagnostic laboratory was not equipped for reagent production, we took advantage of space and personnel that became available due to closure of the research division of our medical center.

Open Development and Clinical Validation of Multiple 3D-Printed Nasopharyngeal Collection Swabs: Rapid Resolution of a Critical COVID-19 Testing Bottleneck.

The pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused a severe international shortage of the nasopharyngeal swabs that are required for collection of optimal specimens, creating a critical bottleneck blocking clinical laboratories' ability to perform high-sensitivity virological testing for SARS-CoV-2. To address this crisis, we designed and executed an innovative, cooperative, rapid-response translational-research program that brought together health care workers, manufacturers, and scientists to emergently develop and clinically validate new swabs for immediate mass production by 3D printing. We performed a multistep preclinical evaluation of 160 swab designs and 48 materials from 24 companies, laboratories, and individuals, and we shared results and other feedback via a public data repository (http://github.

Bringing Antimicrobial Susceptibility Testing for New Drugs into the Clinical Laboratory: Removing Obstacles in Our Fight against Multidrug-Resistant Pathogens.

There are now several new antibiotics available to treat multidrug-resistant pathogens, and susceptibility testing methods for these drugs are increasingly available at the time of drug approval. However, lack of clarity regarding verification requirements remains a formidable barrier to introducing such testing in clinical laboratories, making these drugs practically unavailable for patient use. We propose a change in the framework for bringing in testing for new antibiotics, focusing on quality control rather than underpowered verification studies.

Rapid Susceptibility Testing Methods.

With emerging antimicrobial resistance, rapid antimicrobial susceptibility testing (AST) is needed to provide early definitive therapeutic guidance to optimize patient outcome. Genotypic methods are fast, but can identify only a subset of known resistance elements. Phenotypic methods determine clinically predictive minimal inhibitory concentrations and include very sensitive optical and biophysical methods to detect changes in replication or physiology of pathogens in response to antibiotics.

A Whole-Cell Screen for Adjunctive and Direct Antimicrobials Active against Carbapenem-Resistant Enterobacteriaceae.

Carbapenem-resistant Enterobacteriaceae (CRE) are an emerging antimicrobial resistance threat for which few if any therapeutic options remain. Identification of new agents that either inhibit CRE or restore activity of existing antimicrobials is highly desirable. Therefore, a high-throughput screen of 182,427 commercially available compounds was used to identify small molecules which either enhanced activity of meropenem against a carbapenem-resistant ST258 screening strain and/or directly inhibited its growth.

Evaluation of apramycin against spectinomycin-resistant and -susceptible strains of Neisseria gonorrhoeae.

Background: The emergence of Neisseria gonorrhoeae resistant to all currently available antimicrobial therapies poses a dire public health threat. New antimicrobial agents with activity against N. gonorrhoeae are urgently needed.

Evaluation of apramycin activity against methicillin-resistant, methicillin-sensitive, and vancomycin-intermediate Staphylococcus aureus clinical isolates.

We evaluated the in vitro activity of apramycin against clinical strains of vancomycin-intermediate and methicillin-resistant and -susceptible Staphylococcus aureus. Apramycin demonstrated an MIC/MIC of 8/16 μg/mL. No strains had an MIC above the epidemiological cutoff value of 32 μg/mL, suggesting apramycin resistance mechanisms are rare in this strain population.

The Inoculum Effect in the Era of Multidrug Resistance: Minor Differences in Inoculum Have Dramatic Effect on MIC Determination.

The observed MIC may depend on the number of bacteria initially inoculated into the assay. This phenomenon is termed the inoculum effect (IE) and is often most pronounced for β-lactams in strains expressing β-lactamase enzymes. The Clinical and Laboratory Standards Institute (CLSI)-recommended inoculum is 5 × 10 CFU ml with an acceptable range of 2 × 10 to 8 × 10 CFU ml IE testing is typically performed using an inoculum 100-fold greater than the CLSI-recommended inoculum.

Efficacy of Apramycin against Multidrug-Resistant Acinetobacter baumannii in the Murine Neutropenic Thigh Model.

Apramycin, an aminocyclitol aminoglycoside, was rapidly bactericidal against In a neutropenic murine thigh infection model, treatment-associated CFU reductions of >4 log per thigh were observed for all exposures for which area under the curve (AUC)/MIC ratio was >50 and maximum concentration of drug in serum ()/MIC was ≈10 or higher. Based on these findings, we suggest that apramycin deserves further preclinical exploration as a repurposed therapeutic for multidrug-resistant Gram-negative pathogens, including .

Automated Interpretation of Blood Culture Gram Stains by Use of a Deep Convolutional Neural Network.

Microscopic interpretation of stained smears is one of the most operator-dependent and time-intensive activities in the clinical microbiology laboratory. Here, we investigated application of an automated image acquisition and convolutional neural network (CNN)-based approach for automated Gram stain classification. Using an automated microscopy platform, uncoverslipped slides were scanned with a 40× dry objective, generating images of sufficient resolution for interpretation.

Screening for synergistic activity of antimicrobial combinations against carbapenem-resistant Enterobacteriaceae using inkjet printer-based technology.

Background: Synergistic combination antimicrobial therapy may provide new options for treatment of MDR infections. However, comprehensive in vitro synergy data are limited and facile methods to perform synergy testing in a clinically actionable time frame are unavailable.

Objectives: To systematically investigate a broad range of antibiotic combinations for evidence of synergistic activity against a collection of carbapenem-resistant Enterobacteriaceae (CRE) isolates.

Development of MAST: A Microscopy-Based Antimicrobial Susceptibility Testing Platform.

Antibiotic resistance is compromising our ability to treat bacterial infections. Clinical microbiology laboratories guide appropriate treatment through antimicrobial susceptibility testing (AST) of patient bacterial isolates. However, increasingly, pathogens are developing resistance to a broad range of antimicrobials, requiring AST of alternative agents for which no commercially available testing methods are available.

Activity of minocycline against Klebsiella pneumoniae carbapenemase (KPC)-producing Enterobacteriaceae clinical isolates, with comparison to doxycycline and tigecycline.

We examined the in vitro activity of minocycline against 103 Klebsiella pneumoniae carbapenemase (KPC)-producing Enterobacteriaceae isolates and found approximately half had susceptible (26%) or intermediate (26%) MICs. For a subset of 35 isolates, susceptibility was highest to tigecycline (71% FDA vs. 20% EUCAST) followed by minocycline (14%) and then doxycycline (6%).