Altered Plasmodium falciparum Sensitivity to the Antiretroviral Protease Inhibitor Lopinavir Associated with Polymorphisms in pfmdr1

ABSTRACT The HIV protease inhibitor lopinavir inhibits Plasmodium falciparum aspartic proteases (plasmepsins) and parasite development, and children receiving lopinavir-ritonavir experienced fewer episodes of malaria than those receiving other antiretroviral regimens. Resistance to lopinavir was selected in vitro over ∼9 months, with ∼4-fold decreased sensitivity. Whole-genome sequencing of resistant parasites showed a mutation and increased copy number in pfmdr1 and a mutation in a protein of unknown function, but no polymorphisms in plasmepsin genes.

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Whole-Genome Sequencing for Detecting Antimicrobial Resistance in Nontyphoidal Salmonella

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.01030-16 ◽

2016 ◽

Vol 60 (9) ◽

pp. 5515-5520 ◽

Cited By ~ 148

Author(s):

Patrick F. McDermott ◽

Gregory H. Tyson ◽

Claudine Kabera ◽

Yuansha Chen ◽

Cong Li ◽

...

Keyword(s):

Antimicrobial Resistance ◽

Whole Genome Sequencing ◽

Genome Sequencing ◽

Resistance Genes ◽

The United States ◽

Whole Genome ◽

Content Type ◽

Structural Resistance ◽

Retail Meat

ABSTRACTLaboratory-basedin vitroantimicrobial susceptibility testing is the foundation for guiding anti-infective therapy and monitoring antimicrobial resistance trends. We used whole-genome sequencing (WGS) technology to identify known antimicrobial resistance determinants among strains of nontyphoidalSalmonellaand correlated these with susceptibility phenotypes to evaluate the utility of WGS for antimicrobial resistance surveillance. Six hundred fortySalmonellaof 43 different serotypes were selected from among retail meat and human clinical isolates that were tested for susceptibility to 14 antimicrobials using broth microdilution. The MIC for each drug was used to categorize isolates as susceptible or resistant based on Clinical and Laboratory Standards Institute clinical breakpoints or National Antimicrobial Resistance Monitoring System (NARMS) consensus interpretive criteria. Each isolate was subjected to whole-genome shotgun sequencing, and resistance genes were identified from assembled sequences. A total of 65 unique resistance genes, plus mutations in two structural resistance loci, were identified. There were more unique resistance genes (n =59) in the 104 human isolates than in the 536 retail meat isolates (n =36). Overall, resistance genotypes and phenotypes correlated in 99.0% of cases. Correlations approached 100% for most classes of antibiotics but were lower for aminoglycosides and beta-lactams. We report the first finding of extended-spectrum β-lactamases (ESBLs) (blaCTX-M1andblaSHV2a) in retail meat isolates ofSalmonellain the United States. Whole-genome sequencing is an effective tool for predicting antibiotic resistance in nontyphoidalSalmonella, although the use of more appropriate surveillance breakpoints and increased knowledge of new resistance alleles will further improve correlations.

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pfmdr1Amplification Is Related to Increased Plasmodium falciparum In Vitro Sensitivity to the Bisquinoline Piperaquine

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.06350-11 ◽

2012 ◽

Vol 56 (7) ◽

pp. 3615-3619 ◽

Cited By ~ 21

Author(s):

M. I. Veiga ◽

P. E. Ferreira ◽

M. Malmberg ◽

L. Jörnhagen ◽

A. Björkman ◽

...

Keyword(s):

Plasmodium Falciparum ◽

Copy Number ◽

Statistical Significance ◽

Resistance Locus ◽

Content Type ◽

Enhanced Sensitivity ◽

In Vitro Sensitivity ◽

Artemisinin Combination Therapies

ABSTRACTThe 4-aminoquinoline bisquinoline piperaquine is an important partner drug in one of the presently recommended artemisinin combination therapies. Recent clinical trials have confirmed its high efficacy in combination with dihydroartemisinin. Resistance to piperaquine alone has, however, been documented. Amplification in copy number of thePlasmodium falciparummultidrug resistance locus on chromosome 5, containing thepfmdr1gene, has been shown to confer resistance to structurally unrelated antimalarials. Through the determination of the 50% inhibitory concentrations (IC50s) and IC90s for piperaquine and chloroquine in a set of 46 adaptedP. falciparumcultures originating from the Thai-Burmese border, we have characterized the regions around thepfmdr1gene and identified a significant association between the presence ofpfmdr1duplications and enhanced sensitivity to piperaquine (P= 0.005 for IC50andP= 0.002 for IC90) and chloroquine, reaching statistical significance at IC90s (P= 0.026). These results substantiate the potential importance ofpfmdr1copy number amplifications in the efficacy of the combination therapy piperaquine-dihydroartemisinin. It supports the rational use of 4-aminoquinolines and artemisinin-based compounds, as they independently select for mutually incompatible combinations of mutations.

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Plasmodium falciparum Founder Populations in Western Cambodia Have Reduced Artemisinin SensitivityIn Vitro

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.03055-14 ◽

2014 ◽

Vol 58 (8) ◽

pp. 4935-4937 ◽

Cited By ~ 33

Author(s):

Chanaki Amaratunga ◽

Benoit Witkowski ◽

Dalin Dek ◽

Vorleak Try ◽

Nimol Khim ◽

...

Keyword(s):

Plasmodium Falciparum ◽

Parasite Clearance ◽

Parasite Development ◽

Ring Stage ◽

Content Type ◽

Short Course ◽

Founder Populations ◽

Admixed Population ◽

Survival Assay

ABSTRACTReducedPlasmodium falciparumsensitivity to short-course artemisinin (ART) monotherapy manifests as a long parasite clearance half-life. We recently defined three parasite founder populations with long half-lives in Pursat, western Cambodia, where reduced ART sensitivity is prevalent. Using the ring-stage survival assay, we show that these founder populations have reduced ART sensitivityin vitroat the early ring stage of parasite development and that a genetically admixed population contains subsets of parasites with normal or reduced ART sensitivity.

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Conjugal Transfer, Whole-Genome Sequencing, and Plasmid Analysis of Four mcr-1-Bearing Isolates from U.S. Patients

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.02417-18 ◽

2019 ◽

Vol 63 (4) ◽

Cited By ~ 1

Author(s):

Wenming Zhu ◽

Adrian Lawsin ◽

Rebecca L. Lindsey ◽

Dhwani Batra ◽

Kristen Knipe ◽

...

Keyword(s):

Escherichia Coli ◽

Antimicrobial Resistance ◽

Genome Sequencing ◽

Resistance Genes ◽

Whole Genome ◽

Colistin Resistance ◽

Content Type ◽

Plasmid Analysis ◽

Antimicrobial Resistance Genes

ABSTRACT Four Enterobacteriaceae clinical isolates bearing mcr-1 gene-harboring plasmids were characterized. All isolates demonstrated the ability to transfer colistin resistance to Escherichia coli; plasmids were stable in conjugants after multiple passages on nonselective media. mcr-1 was located on an IncX4 (n = 3) or IncN (n = 1) plasmid. The IncN plasmid harbored 13 additional antimicrobial resistance genes. Results indicate that the mcr-1-bearing plasmids in this study were highly transferable in vitro and stable in the recipients.

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Whole-Genome Sequencing of a Brucella melitensis Strain (BMWS93) Isolated from a Bank Clerk and Exhibiting Complete Resistance to Rifampin

Microbiology Resource Announcements ◽

10.1128/mra.01645-18 ◽

2019 ◽

Vol 8 (33) ◽

Author(s):

Zhi-guo Liu ◽

Xiao-an Cao ◽

Miao Wang ◽

Dong-ri Piao ◽

Hong-yan Zhao ◽

...

Keyword(s):

Whole Genome Sequencing ◽

Genome Sequencing ◽

Inner Mongolia ◽

Brucella Melitensis ◽

Public Health Problem ◽

Human Brucellosis ◽

Whole Genome ◽

Zone Of Inhibition ◽

Content Type

Human brucellosis has become the most severe public health problem in the Ulanqab region of Inner Mongolia, China. Brucella melitensis BMWS93 was obtained from a blood sample taken from a bank clerk in the Ulanqab region of Inner Mongolia, China, and antimicrobial susceptibility testing in vitro showed no zone of inhibition, which confirmed resistance to rifampin. Therefore, whole-genome sequencing of this isolate was performed to better understand the mechanism of this resistance.

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Decreasingpfmdr1Copy Number Suggests that Plasmodium falciparum in Western Cambodia Is RegainingIn VitroSusceptibility to Mefloquine

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.05163-14 ◽

2015 ◽

Vol 59 (5) ◽

pp. 2934-2937 ◽

Cited By ~ 25

Author(s):

Pharath Lim ◽

Dalin Dek ◽

Vorleak Try ◽

Sokunthea Sreng ◽

Seila Suon ◽

...

Keyword(s):

Plasmodium Falciparum ◽

Combination Therapy ◽

Molecular Marker ◽

Copy Number ◽

Artemisinin Combination Therapy ◽

Clinical Isolates ◽

Content Type

ABSTRACTDihydroartemisinin-piperaquine is the current frontline artemisinin combination therapy (ACT) forPlasmodium falciparummalaria in Cambodia but is now failing in several western provinces. To investigate artesunate plus mefloquine (AS+MQ) as a replacement ACT, we measured the prevalence of multiplepfmdr1copies—a molecular marker for MQ resistance—in 844P. falciparumclinical isolates collected in 2008 to 2013. Thepfmdr1copy number is decreasing in Western Cambodia, suggesting thatP. falciparumis regainingin vitrosusceptibility to MQ.

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Analysis of the Conformation and Function of the Plasmodium falciparum Merozoite Proteins MTRAP and PTRAMP

Eukaryotic Cell ◽

10.1128/ec.00039-12 ◽

2012 ◽

Vol 11 (5) ◽

pp. 615-625 ◽

Cited By ~ 21

Author(s):

Onyinyechukwu Uchime ◽

Raul Herrera ◽

Karine Reiter ◽

Svetlana Kotova ◽

Richard L. Shimp ◽

...

Keyword(s):

Plasmodium Falciparum ◽

Binding Assay ◽

Actin Binding ◽

Parasite Development ◽

Content Type ◽

Content Development ◽

Falciparum Merozoite ◽

Erythrocyte Binding ◽

And Function

ABSTRACT Thrombospondin repeat (TSR)-like domains are structures involved with cell adhesion. Plasmodium falciparum proteins containing TSR domains play crucial roles in parasite development. In particular, the preerythrocytic P. falciparum circumsporozoite protein is involved in hepatocyte invasion. The importance of these domains in two other malaria proteins, the merozoite-specific thrombospondin-related anonymous protein (MTRAP) and the thrombospondin-related apical membrane protein (PTRAMP), were assessed using near-full-length recombinant proteins composed of the extracellular domains produced in Escherichia coli . MTRAP is thought to be released from invasive organelles identified as micronemes during merozoite invasion to mediate motility and host cell invasion through an interaction with aldolase, an actin binding protein involved in the moving junction. PTRAMP function remains unknown. In this study, the conformation of recombinant MTRAP (rMTRAP) appeared to be a highly extended protein (2 nm by 33 nm, width by length, respectively), whereas rPTRAMP had a less extended structure. Using an erythrocyte binding assay, rMTRAP but not rPTRAMP bound human erythrocytes; rMTRAP binding was mediated through the TSR domain. MTRAP- and in general PTRAMP-specific antibodies failed to inhibit P. falciparum development in vitro . Altogether, MTRAP is a highly extended bifunctional protein that binds to an erythrocyte receptor and the merozoite motor.

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Early Detection of Emergent Extensively Drug-Resistant Tuberculosis by Flow Cytometry-Based Phenotyping and Whole-Genome Sequencing

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.01834-18 ◽

2019 ◽

Vol 63 (4) ◽

Cited By ~ 2

Author(s):

Max R. O’Donnell ◽

Michelle H. Larsen ◽

Tyler S. Brown ◽

Paras Jain ◽

Vanisha Munsamy ◽

...

Keyword(s):

Drug Resistance ◽

Genome Sequencing ◽

Low Frequency ◽

Whole Genome ◽

Drug Resistant ◽

Drug Resistant Tuberculosis ◽

Content Type ◽

Resistant Tuberculosis ◽

Extensively Drug Resistant

ABSTRACTA critical gap in tuberculosis (TB) treatment is detection of emergent drug resistance. We hypothesized that advanced phenotyping with whole-genome sequencing (WGS) will detect low-frequencyMycobacterium tuberculosisdrug resistance. We assessed a reporter mycobacteriophage (Φ2GFP10)in vitroto detect drug-resistant subpopulations and predictM. tuberculosisbactericidal activity in this pilot study. Subsequently, we prospectively studied 20 TB patients with serial Φ2GFP10, Xpert MTB/RIF, andM. tuberculosisculture through end of treatment. WGS was performed, and single nucleotide polymorphisms (SNPs) were examined to detect mixed infection in selectedM. tuberculosisisolates. ResistantM. tuberculosisisolates were detected at 1:100,000, and changes in cytometry-gated events were predictive ofin vitroM. tuberculosisbactericidal activity using the Φ2GFP10 assay. Emergent drug resistance was detected in one patient by Φ2GFP10 at 3 weeks but not by conventional testing (M. tuberculosisculture and GeneXpert). WGS revealed a phylogeographically distinct extensively drug-resistant tuberculosis (XDR-TB) genome, identical to an XDR-TB isolate from the patient’s spouse. Variant lineage-specific SNPs were present early, suggesting mixed infection as the etiology of emergent resistance with temporal trends providing evidence for selection during treatment. Φ2GFP10 can detect low-frequency drug-resistantM. tuberculosisand with WGS characterize emergentM. tuberculosisresistance. In areas of high TB transmission and drug resistance, rapid screening for heteroresistance should be considered.

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ReducedIn VitroDoxycycline Susceptibility in Plasmodium falciparum Field Isolates from Kenya Is Associated with PfTetQ KYNNNN Sequence Polymorphism

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.02788-13 ◽

2014 ◽

Vol 58 (10) ◽

pp. 5894-5899 ◽

Cited By ~ 10

Author(s):

Angela O. Achieng ◽

Luiser A. Ingasia ◽

Dennis W. Juma ◽

Agnes C. Cheruiyot ◽

Charles A. Okudo ◽

...

Keyword(s):

Plasmodium Falciparum ◽

Amino Acid ◽

Genetic Polymorphisms ◽

Copy Number ◽

Sequence Polymorphism ◽

Limited Information ◽

Content Type ◽

Field Isolates ◽

Potential Marker

ABSTRACTDoxycycline is widely used for malaria prophylaxis by international travelers. However, there is limited information on doxycycline efficacy in Kenya, and genetic polymorphisms associated with reduced efficacy are not well defined.In vitrodoxycycline susceptibility profiles for 96Plasmodium falciparumfield isolates from Kenya were determined. Genetic polymorphisms were assessed inP. falciparummetabolite drug transporter (Pfmdt) andP. falciparumGTPasetetQ(PftetQ) genes. Copy number variation of the gene and the number of KYNNNN amino acid motif repeats within the protein encoded by PftetQwere determined. Reducedin vitrosusceptibility to doxycycline was defined by 50% inhibitory concentrations (IC50s) of ≥35,000 nM. The odds ratio (OR) of having 2 PfTetQ KYNNNN amino acid repeats in isolates with IC50s of >35,000 nM relative to those with IC50s of <35,000 nM is 15 (95% confidence interval [CI], 3.0 to 74.3;Pvalue of <0.0002). Isolates with 1 copy of the Pfmdtgene had a median IC50of 6,971 nM, whereas those with a Pfmdtcopy number of >1 had a median IC50of 9,912 nM (P= 0.0245). Isolates with 1 copy of PftetQhad a median IC50of 6,370 nM, whereas isolates with a PftetQcopy number of >1 had a median IC50of 3,422 nM (P< 0.0007). Isolates with 2 PfTetQ KYNNNN motif repeats had a median IC50of 26,165 nM, whereas isolates with 3 PfTetQ KYNNNN repeats had a median IC50of 3,352 nM (P= 0.0023). PfTetQ sequence polymorphism is associated with a reduced doxycycline susceptibility phenotype in Kenyan isolates and is a potential marker for susceptibility testing.

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Application of Whole-Genome Sequencing Data for O-Specific Antigen Analysis andIn SilicoSerotyping of Pseudomonas aeruginosa Isolates

Journal of Clinical Microbiology ◽

10.1128/jcm.00349-16 ◽

2016 ◽

Vol 54 (7) ◽

pp. 1782-1788 ◽

Cited By ~ 17

Author(s):

Sandra Wingaard Thrane ◽

Véronique L. Taylor ◽

Ole Lund ◽

Joseph S. Lam ◽

Lars Jelsbak

Keyword(s):

Pseudomonas Aeruginosa ◽

Whole Genome Sequencing ◽

Genome Sequencing ◽

Specific Antigen ◽

Whole Genome Sequencing Data ◽

Whole Genome ◽

Sequencing Data ◽

Chronic Infections ◽

Content Type

Accurate typing methods are required for efficient infection control. The emergence of whole-genome sequencing (WGS) technologies has enabled the development of genome-based methods applicable for routine typing and surveillance of bacterial pathogens. In this study, we developed thePseudomonas aeruginosaserotyper (PAst) program, which enabledin silicoserotyping ofP. aeruginosaisolates using WGS data. PAst has been made publically available as a web service and aptly facilitates high-throughput serotyping analysis. The program overcomes critical issues such as the loss ofin vitrotypeability often associated withP. aeruginosaisolates from chronic infections and quickly determines the serogroup of an isolate based on the sequence of the O-specific antigen (OSA) gene cluster. Here, PAst analysis of 1,649 genomes resulted in successful serogroup assignments in 99.27% of the cases. This frequency is rarely achievable by conventional serotyping methods. The limited number of nontypeable isolates found using PAst was the result of either a complete absence of OSA genes in the genomes or the artifact of genomic misassembly. With PAst,P. aeruginosaserotype data can be obtained from WGS information alone. PAst is a highly efficient alternative to conventional serotyping methods in relation to outbreak surveillance of serotype O12 and other high-risk clones, while maintaining backward compatibility to historical serotype data.

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