Volume 7, Number 2—April 2001
4th Decennial International Conference on Nosocomial and Healthcare-Associated Infections
State of the Art
Molecular Approaches to Diagnosing and Managing Infectious Diseases: Practicality and Costs
|Organism(s)||Antimicrobial agent(s)||Gene||Detection method|
|Staphylococci||Methicillin Oxacillin||mec Ab||Standard DNA probe Branched chain DNA probe PCR|
|Enterococci||Vancomycin||van A, B, C, Dc||Standard DNA probe PCR|
|Enterobacteriaceae Haemophilus influenzae Neisseria gonorrhoeae||Beta-lactams||blaTEM and blaSHVd||Standard probe PCR and RFLP PCR and sequencing|
|Enterobacteriaceae and gram-positive cocci||Quinolones||Point mutations in gyr A, gyr B, par C and par E||PCR and sequencing|
|Mycobacterium tuberuclosise||Rifampin Isoniazid Ethambutol Streptomycin||Point mutations in rpo B Point mutations in kat G, inh A, and ahp C Point mutations in emb B Point mutations in rps L and rrs||PCR and SSCP PCR and sequencing PCR and SSCP PCR and sequencing PCR and RFLP|
|Herpes virusesf||Acyclovir and related drugs Foscarnet||Mutations or deletions in the TK gene Point mutations in DNA polymerase gene||PCR and sequencing PCR and sequencing|
|HIVg||Nucleoside reverse transcriptase inhibitors Protease inhibitors||Point mutations in RT gene Point mutations in PROT gene||PCR and sequencing PCR and LIPA PCR and sequencing|
aAdapted from Pfaller (2). bmecA encodes for the altered penicillin binding protein PBP2a'; phenotypic methods may require 48 hours incubation or more to detect resistance and are less than 100% sensitive. Detection of mecA has potential for clinical application in specific circumstances.
cVancomycin resistance in enterococci may be related to one of four distinct resistance genotypes of which vanA and vanB are most important. Genotypic detection of resistance is useful in validation of phenotypic methods.
dThe genetic basis of resistance to beta-lactam antibiotics is extremely complex. The blaTEM and blaSHV genes are the two most common sets of plasmid encoded beta-lactamases. The presence of either a blaTEM or blaSHV gene implies ampicillin resistance. Variants of the blaTEM and blaSHV genes (extended spectrum beta-lactamases) may also encode for resistance to a range of third-generation cephalosporins and to monobactams.
eM. tuberculosis is very slow growing. Four weeks or more may be required to obtain phenotypic susceptibility test results. Detection of resistance genes in M. tuberculosis has potential for clinical application in the short term.
fThere are no phenotypic methods sufficiently practical for routine clinical detection of resistance to antiviral agents. Genotypic methods represent a practical method for routine detection of antiviral resistance.
gAbbreviations not defined in text: RFLP, restriction fragment length polymorphism; SSCP, single-stranded conformational polymorphism; LIPA, line probe assay; TK, thymidine kinase; RT, reverse transcriptase; PROT, protease.
- Cormican MG, Pfaller MA. Molecular pathology of infectious diseases. In: Henry JB, editor. Clinical diagnosis and management by laboratory methods. 19th ed. Philadelphia: W.B. Saunders Company; 1996:1390-9.
- Pfaller MA. Diagnosis and management of infectious diseases: Molecular methods for the new millennium. Clinical Laboratory News. 2000;26:10–3.
- Kant JA. Molecular diagnostics: Reimbursement and other selected financial issues. Diagn Mol Pathol. 1995;4:79–81.
- Fredricks DN, Relman DA. Sequence-based identification of microbial pathogens: A reconsideration of Koch's postulates. Clin Microbiol Rev. 1996;9:18–33.
- Fredricks DN, Relman DA. Application of polymerase chain reaction to the diagnosis of infectious disease. Clin Infect Dis. 1999;29:475–88.
- Tang YW, Persing DH. Molecular detection and identification of microorganisms. In: Murray PR, Baron EJ, Pfaller MA, Tenover FC, Yolken RH, editors. Manual of clinical microbiology. 7th ed. Washington: American Society for Microbiology; 1999:215-44.
- Woods GL. Molecular techniques in mycobacterial detection. Arch Pathol Lab Med. 2001;125:122–6.
- Bergeron MG, Ouellette M. Preventing antibiotic resistance using rapid DNA-based diagnostic tests. Infect Control Hosp Epidemiol. 1998;19:560–4.
- Cockerill FR III. Genetic methods for assessing antimicrobial resistance. Antimicrob Agents Chemother. 1999;43:199–212.
- Hacek DM, Suriano T, Noskin GA, Kruszynski J, Reisberg B, Peterson LR. Medical and economic benefit of a comprehensive infection control program that includes routine determination of microbial clonality. Am J Clin Pathol. 1999;111:647–54.
- Ross JS. Financial determinants of outcomes in molecular testing. Arch Pathol Lab Med. 1999;123:1071–5.
- Pfaller MA. Molecular epidemiology in the care of patients. Arch Pathol Lab Med. 1999;123:1007–10.
- Nolte FS. Impact of viral load testing on patient care. Arch Pathol Lab Med. 1999;123:1011–4.
- Anthony RM, Brown TJ, French GL. Rapid diagnosis of bacteremia by universal amplification of 23S ribosomal DNA followed by hybridization to an oligonucleotide array. J Clin Microbiol. 2000;38:781–8.
- Marshall SA, Wilke WW, Pfaller MA, Jones RN. Staphylococcus aureus and coagulase-negative staphylococci from blood stream infections: Frequency of occurrence, antimicrobial susceptibility, and molecular (mec A) characterization of oxacillin resistance in the SCOPE Program. Diagn Microbiol Infect Dis. 1998;30:205–14.
- Hussain Z, Stoakes L, Massey V, Diagre D, Fitzgerald V, El Sayed S, Correlation of oxacillin MIC with mec A gene carriage in coagulase-negative staphylococci. J Clin Microbiol. 2000;38:752–4.
- Hecht FM, Grant RM, Petropoulos CJ, Dillon B, Chesney MA, Tian H, Sexual transmission of an HIV-1 variant resistant to multiple reverse-transcriptase and protease inhibitors. N Engl J Med. 1998;339:307–11.
- Stuyver L, Van Geyt C, de Gendt S, Van Reybroeck G, Zoulin F, Leroux-Rods G, Line probe assay for monitoring drug resistance in hepatitis B virus-infected patients during antiviral therapy. J Clin Microbiol. 2000;38:702–7.
- Courvalin P. Genotypic approach to the study of bacterial resistance to antibiotics. Antimicrob Agents Chemother. 1991;35:1019–23.
- Arbeit RD. Laboratory procedures for epidemiologic analysis of microorganisms. In: Murray PR, Baron EJ, Pfaller MA, Tenover FC, Yolken RH, editors. Manual of clinical microbiology. 7th ed. Washington: American Society for Microbiology; 1999:116-37.
- Pfaller MA, Herwaldt LA. The clinical microbiology laboratory and infection control: Emerging pathogens, antimicrobial resistance, and new technology. Clin Infect Dis. 1997;25:858–70.
- Ferreira-Gonzalez A, Garrett CG. Pitfalls in establishing a molecular diagnostic laboratory. Hum Pathol. 1996;27:437–40.
- Back NA, Linnemann CC, Pfaller MA, Staneck JL, Morthland V. Recurrent epidemics caused by a single strain of erythromycin-resistant Staphylococcus aureus: The importance of molecular epidemiology. JAMA. 1993;270:1329–33.