Bacterial ResistanceProblem Space

Ehren BucholtzSaint Mary-of-the-Woods College

Mark GalloNiagara University

Background

• Microbial drug resistance (DR) is a growing concern among health professionals and others around the world.

• DR Issues that can be studied in a problem space environment include– Types of Drug classes– Types of Bacteria– Mechanism of Action– Mechanisms of DR

Microbial drug resistance

Biological PrinciplesNext slide

ToolsBiology Workbench

NCBI

Data SetsCDCFDA

Drug DigestRxList

Drug Resistance

Drug Resistance

Bacteriatypes

DrugClasses

Mechanism of Action

ResistanceMechanisms

Cellularlevel

Mutations

Antibacterials

Antibiotics

Phylogeny

Geneticexchange

Hostinteraction

Bacteriostatic

Bacteriocidal

Coevolution

EcologicalFactors

Resources

• Centers for Disease Control– Case studies– Outbreak data

• Genbank

• Biology Workbench

Multidrug Resistance Region of Salmonella

enterica Serovar Typhimurium DT104

• Outbreaks of MDR Typhimurium DT104 have also been reported in poultry, beef, cheese, and swine in numerous countries

• Resistant to ampicillin, chloramphenicol, streptomycin, sulfonamides, and tetracycline

• however, isolates have been identified which are also resistant to fluoroquinolones, trimethoprim, and kanamycin

• antimicrobial resistance gene is clustered on a fragment of less than 46 kb

Task: Identify Resistance ORFs

• Students are given pieces of nucleotide sequence of SGi1 to identify homology– beta-lactamase – Tetracycline resistance– Integrase– Transposase

• beta_lactamase (partial sequence shown) NEGKLGDLRDTTTPKAIASTLNKFLFGSALSEMNQKKLESWMVNNQVTGNLLRSVLPAGW

Find results for

• 4377727_4377728Vibrio cholerae non O1, non O139 plasmid class A beta-lactamase

• 216842_216843Proteus mirabilis plasmid pCS229 blaP gene for bata-lactamase,

• 30314015_30314018Pasteurella multocida plasmid pJR2, complete sequence.

• 9719055_9719057Vibrio cholerae class 1 integron integrase (intI1) gene,partial

• 12719011_12719033Salmonella enterica subsp. enterica serovar Typhimurium genomic

• 151074_151076P.aeruginosa beta-lactamase (PSE-1) gene, complete cds

• 47647_47648S.typhimurium beta-lactamase gene.

BlastP and Clustal for beta lactamase unknown

• 30314015_3031401 NEGKLGDLRDTTTPKAIASTLNKFLFGSALSEMNQKKLESWMVNNQVTGNLLRSVLPAGW• 216842_216843 NEGKLGDLRDTTTPKAIASTLNKFLFGSALSEMNKKKLESWMVNNQVTGNLLRSVLPAGW • 47647_47648 NEGKLGDLRDTTTPKAIASTLNKFLFGSALSEMNQKKLESWMVNNQVTGNLLRSVLPAGW • 151074_151076 NEGKLGDLRDTTTPKAIASTLNKFLFGSALSEMNQKKLESWMVNNQVTGNLLRSVLPAGW • 12719011_12719033 NEGKLGDLRDTTTPKAIASTLNKFLFGSALSEMNQKKLESWMVNNQVTGNLLRSVLPAGW • 9719055_9719057 NEGKLGDLRDTTTPKAIASTLNKFLFGSALSEMNQKKLESWMVNNQVTGNLLRSVLPAGW• beta_lactamase NEGKLGDLRDTTTPKAIASTLNKFLFGSALSEMNQKKLESWMVNNQVTGNLLRSVLPAGW • 4377727_4377728 NEGKLGDLRDTTTPKAIASTLNQLLFGSTLSEASQKKLESWMVNNQVTGNLLRSVLPVKW • **********************::****:*** .:**********************. *

Rooted Tree from ClustalW

Other project ideas

• Use wealth of CDC case study data to fill out project space

CDC Case Study

• Variant Salmonella Genomic Island 1 Antibiotic Resistance Gene Cluster in Salmonella enterica Serovar Albany– Benoît Doublet et.al. Emerg Infect Dis 2003 May

Available from: URL: http://www.cdc.gov/ncidod/EID/vol9no5/02-0609.htm

Something’s fishy

• A variant SGI1, Salmonella genomic island 1, was identified as an antibiotic-resistance gene cluster in a multidrug-resistant strain of S. enterica serovar Albany isolated from food fish from Thailand and imported to France. 

• In this strain, the streptomycin resistance aadA2 gene cassette in one of the SGI1 integrons was replaced by a dfrA1 gene cassette, conferring resistance to trimethoprim and an open reading frame of unknown function.

Gene Function Treasure Hunt

• Clustering of Salmonella isolates