Review on Laboratory Testing Methods for Staphylococcus Aureus - Book Report/Review Example

Name of university Name of college/department Name of student Student number Course title Course number REVIEW OF LABORATORY TESTING METHODS FOR STAPHYLOCOCCUS AUREUS TO BETA LACTAM SUSCEPTIBILITY ANTIBIOTICS DECLARATION: This is my original work and has never been submitted before for any award or published in any medical journal or magazine Signature of student Date of submission Definition of terms BHI: Brain Heart Infusion CA-MRSA: Community Associated Multiple/Methicillin Resistant Staphylococcus Aureus GISA: Glycopeptides Intermediate Staphylococcus Aureus HA-MRSA: Healthcare- Associated Multiple/Methicillin Resistant Staphylococcus Aureus MIC: Minimum Inhibitory Concentration MRSA: Methicillin-Resistant Staphylococcus Aureus MRSA: Multiple-Resistant Staphylococcus Aureus NCCLS: National Committee for Clinical Laboratory Standards ORSA: Oxacilin Resistant Staphylococcus Aureus VISA: Vancomycin Intermediate Staphylococcus Aureus VRE: Vancomycin Resistant Enterococcus VRSA: Vancomycin Resistant Staphylococcus Aureus VSSA: Vancomycin Sensitive/Susceptible Staphylococcus Aureus Table of contents Definition of terms 2 Review of laboratory testing methods for Staphylococcus Aureus susceptibility to Beta-Lactams antibiotics 3 Introduction 3 Method of laboratory testing 5 Effectiveness of the methods 6 Epidemiology, profile testing and population analysis 8 Table 1: phenotypic characteristics 12 Bibliography 12 Review of laboratory testing methods for Staphylococcus Aureus susceptibility to Beta-Lactams antibiotics Introduction Noble et al (1992:195-198), Hiramatsu et al (1997:135-136), CDC (1997:624-626) and Collignon et al (1998:145) argue that Staphylococcus Aureus is a gram positive1 bacterium that causes skin infections2 like cellulitis, boils, impetigo. NCCLS (2000) indicates S. Aureus causes soft tissue infections like abscesses and sinusitis and bone, joint and endovascular infections like endocarditis and vascular graft infections. NCCLS (2000) indicates S. Aureus severe infections are bacteremia, endocarditis, metastatic infections, sepsis and staphylococcal toxic shock syndrome (waldvogel 1995:1754-1777). According to NCCLS (2000), resistance to vancomycin or teicoplanin is measured by Minimum Inhibitory Concentration (MIC) that relies on breakpoints that are laid down by National Committee for Clinical Laboratory Standards (NCCLS). According to NCCLS (2000) and Johnson (1998:189-291), staphylococci that needs vancomycin concentrations of less than or equal to 4µg/mL for inhibition is classified as susceptible/sensitive hence termed as Vancomycin Susceptible/Sensitive Staphylococcus Aureus (VSSA) and Staphylococcus Aureus that needs vancomycin concentration ranging from 8µg/mL to 16µg/mL is classified as intermediate hence termed as Vancomycin Intermediate Staphylococcus Aureus (VISA). If glycopeptide is used it is termed as Glycopeptide Intermediate Staphylococcus Aureus (GISA3). While Staphylococcus Aureus that requires vancomycin concentrations equal to or above 32µg/mL is termed as vancomycin resistant Staphylococcus Aureus (VRSA). Himaratsu et al (1997:135-136), CDC (1997:765-766) and Tenover (2000:49-53) show that VRSA4 is also termed as MRSA that denotes Methicillin/Multiple Resistant Staphylococcus Aureus or in terms of teicoplanin or oxacillin, Oxacillin Resistant Staphylococcus Aureus (ORSA). VRSA is also used to refer to staphylococcus aureus that is able to grow on Brain Heart Infusion (BHI) screening agar plate that has vancomycin of 4µg/mL concentration within 24 hours provided that the vancomycin broth dilution MIC concentration is at least 8µg/mL. Method of laboratory testing Ryan and Ray (1994:221-250) argue that susceptibility testing can be done by using Kirby-Bauer technique. Levinson and Jawetz (1994:54-59) and NCCLS (2000) agree that epsilometer or e-test technique is used to investigate inhibitory concentration of an antibiotic. Kaplan and Mason (1998:2866-2873) indicate agar dilution and broth technique are effective in determining susceptibility tests of staphylococcus aureus. Cockeril (1998:1007-1021) indicates genetic laboratory tests offer prompt susceptibility testing and are based on genetic markers of antibiotic resistance. Mandell et al (2000:184-221) indicates use of automated techniques like microscan (dade microscan, west sacramento, CA) rapid panels. According to NCCLS (2000) and Working party of the British society for antimicrobial chemotherapy (1998:701-710), the best method for susceptibility testing of staphylococcus aureus is MIC that may be carried out through broth dilution, agar dilution, agar gradient diffusion. Black, Thomson and Pitout (2004:2203-2206) and Bauer et al (1966:493-496) indicate disk diffusion method. Hindler and Inderlied (1985:205-210) suggested that agglutination method is a quick technique for testing S. aureus. Effectiveness of the methods Kondo et al (2004), Liu et al (2000:361-366), Sato et al (1997:401-404) and Shibata et al (2003:1478-1483) all agree that measures should be put in place to ensure contamination is reduced because this is very critical for antibiotic susceptibility testing. Shimizu et al (2001:3198-3201), Suzuki et al (1997:327-331) and Yamase et al (1996:459-465) indicate that contaminants occur from specimens that originate from sterile conditions and are passed via colonized area during collection. Isenberg (1992), Komatsuzawa et al (1997:2355-2361), Shimizu et al (2001:3198-3201), Suzuki et al (1997:327-331) and Yamase et al (1996:459-465) provide that needle aspiration, surgical drainage of infected material and sufficient antiseptic preparations assists in preventing contamination. Thornsberry and McDougal (1983:1084-1091) argue that agar microdilution technique provides critical information on concentration of antibiotic that is needed to kill the Staphylococcus Aureus. Thornsberry and McDougal (1983) suggest standard inoculum of the S. Aureus is placed into a series of test tubes that are equipped with appropriate S. Aureus growth medium and an increasing concentration of the antibiotic is added into each test tube and the S .Aureus is given time to incubate. Sato et al (2004:1357-1360) and Thornsberry and McDougal (1983) indicate that broth micro-dilution technique is concerned more with determination of turbidity that confirms growth of S. Aureus. The test tube that has lowest concentration of the antibiotic and has no turbidity is used to determine MIC. Sato et al (2004) and Sato et al (2004b: 28-35) indicate that a sample is drawn from every test tube and inoculated on fresh agar plates to determine concentration of the antibiotic that killed the bacteria. Thornsberry and McDougal (1983) argue that if the antibiotic killed the S. Aureus, there will be no evidence of S. Aureus growth on the agar plates. NCCLS (2000) suggest that the minimum concentration of antibiotic that kills 99.9% of the S. Aureus is termed as Minimal Bactericidal Concentration (MBC). NCCLS (2000) and Hansen and Freedy (1984:494-499) agree that genetic laboratory tests provide prompt susceptibility testing. This, according to Hansen and Freedy (1984), is based on genetic markers of antibiotic resistance. Hansen and Freedy (1984) indicates that genotype susceptibility testing with polymerase chain reaction restriction fragment result into information on length polymorphisms and branched DNA technology determines genetic source of resistance. This method is better compared with other techniques that rely on S. Aureus growth. NCCLS (2000) argues that Epsilometer or E-test technique is a valuable tool when determining concentration of antibiotic. This process is carried out by laying antibiotic strip on agar plate inoculated with S. Aureus. The strip should have a gradient of single antibiotic and be marked at determined regular intervals to show concentration at every level of the strip. If S. Aureus is sensitive or susceptible, a ring of no growth will be formed at the end of the strip that contains highest concentration of the antibiotic. Zhao et al (2001:1737-1742) indicates that the perimeter of the ring of no growth will intersect the strip at a point where concentration of the antibiotic is high enough to inhibit S. Aureus growth showing that it attains minimal inhibitory concentration (MIC) or the lowest concentration of an antibiotic that can inhibit S. Aureus multiplication. Peacock et al (1981:575-582) and Shiota et al (1999:1388-1390) indicated Kirby-Bauer technique as a S. Aureus susceptibility test that is carried out by placing several antibiotic disks where every disk has a different antibiotic on a agar plate that is inoculated with S. Aureus. According to Shiota et al (2000:3198-3201), varying rings of no growth of different sizes form around the disks and this is depended on sensitivity or susceptibility of the S. Aureus to the antibiotic. Shiota et al (1999; 2000) indicates that large rings of ‘no growth’ is an indicator of high sensitivity and suggest that if the S. aureus is resistant to the antibiotic, then, there is no ring that is formed. Kirby-Bauer technique shows that a tested antibiotic inhibits growth of S. Aureus. Epidemiology, profile testing and population analysis Utsul and Tokota (1985:397-403) and Chambers and Sachdenim (1990:1170-1176) argue that MRSA provokes Penicillin Binding Protein 2’ (PBP2’). Chambers and Sachdenim (1990:1170-1176) and Asada et al (1995:517-524) indicate that PBP2’ is hard to inhibit by using beta Lactams antibiotics because of remarkable low binding affinities to beta-lactam antibiotics like ampicillin and penicillin G that has better affinities to PBP2’ (IC50=14-20mg/l. these beta-lactams are not sufficiently strong to treat MRSA illnesses and its opportunistic infections as monotherapeutic agents because of limited tissue concentration. Chamber and Sachdenim (1990) suggests that genetic analysis of chromosomal DNA of MRSA elucidates 5 clones. Hiramatsu, Kondo and Ito (1996:117-129) and Tanaka et al (1995:40) classified the clonotypes as clonotype I-A that was witnessed in UK in 1960s and was represented as NCTC10442, clonotype III-A observed in Britain, Clonotype II-A represented as N315 that was evident in Japan and USA , Clonotype II-A that was identified in Japan in 1990. Tanaka et al (1995) indicates that it is from MRSA clonotype II-A which gave rise into first isolate of VRSA. Hiramatsu et al (1997:135-136) indicate that strains of VISA with MIC equivalent to 8µg/ml were reported in Japan, united States of America (Smith et al 1999:493-501), France (Ploy et al (1998:691-696), UK (Howe et al (1998:602) and Germany (Sieradzki et al 1999:517-523). Smith et al (1999:493-501) indicate opportunistic infections caused by VISA have vancomycin MICs of 8mg/l and are refractory to vancomycin therapy. CDC has shown that there are other infections caused by S. Aureus where MICs are 4 mg/l showing that the patients studied had little improvement on vancomycin therapy. According to Climo et al (1999:1747-1753) vancomycin monotherapy is not sufficient for VISA or GISA strains. Climo et al (1999) suggest a combination of Oxacillin and Vancomycin is synergestic both in vitro and in vivo in endocarditis models. Sieradzki (1999:517-523) and Bierbaum et al (1999:691-696) have observed similar outcomes on synergy of beta-lactams and vancomycin for VISA and GISA. Sieradzki et al (1999:517-523) points out that accumulated work on human beings is not adequate to make a firm conclusion on loss of effectiveness especially opportunitstic infections that are caused by S. Aureus that are heteroresistant to glycopeptides. Hiramatsu, kondo and Ito (1996) indicate that the first MRSA strain Mu50 had a vancomycin MIC of 8mg/L and was isolated from a four month old male baby. Other strains have been identified like Mu3 and Mu45. Both Mu45 and Mu50 differ through vancomycin resistance and genetically. Tenover, Lancaster and Bill et al (1998:1020-1027) indicate that Mu50 is termed as VISA (Vancomycin intermediate Staphylococcus Aureus or GISA (glycopeptides intermediate Staphylococcus Aureus) in the USA and this is based on NCCLS breakpoints for vancomycin namely lessthan 8mg/l (susceptible), 8-16mg/l (intermediate and more than 16mg/l (resistant). Hiramatsu, Aritaka, Hanaki et al (1997:1668-1671) and Tanaka et al (1998:552-553) indicate that population analysis of Mu50 (VISA/GISA) show a PFGE banding pattern and has mutation at gyrA and grlA genes. Tanaka et al (1998:552-553) therefore concludes that Hetero-VRSA is a precursor of VRSA and is responsible for causing therapeutic failures of vancomycin by generating VRSA at high frequency that has appearance rate of 1 into a million cells. NCCLS (2000) indicates that glycopepetide antibiotics namely vancomycin and teicoplanin act on Staphylococcus aureus and other bacteria by binding to carbonyl-terminal D-Alanyl-D-Alanine reactive sites of peptidoglycan named murein monomer. This is depended on the electron withdrawing and electron donating effect of side groups attached to the reactive sites at β and α carbon atoms. NCCLS (2000) shows that the peptidoglycan bound murein monomer is not able to be added (addition reaction) to the nascent peptidoglycan hence peptidoglycan biosynthesis is halted in the presence of glycopetide antibiotic to carry out its antimicrobial activity. Hiramatsu (1998:1020-1027) indicates Vancomycin must therefore penetrate layers of cell wall in order to get at outer surface of cytoplasmic membrane where transglycosylation takes place. Hiramatsu (1998:1020-1027) indicates that cell wall of S. aureus has numerous sites for binding with vancomycin. The sites are D-Alanyl-D-Alanine that fail action of PBP which cross bridges nascent peptidoglycan by cleaving D-Alanyl-D-Alanine residues to the penta-glycine. This means molecules of vancomycin are trapped in the cell wall of S. aureus before reaching their recommended sites. In Mu50, there is overpopulation of PBP2. PBP2’ overproduction does not contribute to vancomycin resistance because loss of mecA gene does not influence vancomycin resistance. It is PBP2 that augments vancomycin and teicoplanin resistance. Introduction of PBP2 gene on plasmid increases teicoplanin MIC from 1 to 8mg/l and vancomycin from 1 to 2 mg/l. Therefore resistance buildup is based on affinity trapping mechanism. Resistance to beta-lactams occurs secondary to either enzyme production or alterations of penicillin binding proteins (PBP). Reistance occurs through development of three manin phenotypes with respect to vancomycin. These are illustrated in table 1 below. Table 1: phenotypic characteristics Phenotype Characteristics vanA MIC (vancomycin) 64 to greater than 1000mcg/ml High level of vancomycin resistance Resistant to teicoplanin Commonly evident with E. faecium Transferable between organisms Inducible by antibiotics VanB MIC (vancomycin) 4 to 1000 mcg/ml Moderately resistant to vancomycin Susceptible to teicoplanin Transferable between organisms vanC MIC (vancomycin) 2 to 32 mcg/ml Low level of vancomycin resistance Susceptible to teicoplanin Not evident in E. Faecalis or E. Faecium Not transferable Bibliography Asada, K.; Inaba, Y.; Tateda-Suzuki,E.; Kuwahara-Arai, K.; Ito,T. and Hiramatsu,K. (1995): Evolution and resistance expression of MRSA: evaluation of beta-lactam antibiotics against a set strains with different types of phenotypic expression. Acta Bio chimica polonica; Volume 42; pp. 517-524. Bauer, A.W.; Kirby, W.M.; Sherris, J.C.; Turck, M. (1966 April): Antibiotic susceptibility testing by a standardized single disk method. Am J Clin Pathol., Volume 45, issue number 4; pp.493–496. Bierbaum G, Fuchs K, Lenz W, Szekat C, Sahl H-G. (1999): Presence of Staphylococcus aureus with reduced susceptibility to vancomycin in Germany. Eur J Clin Microbiol Infect Dis, Volume 18; pp.691-696 Black, J. A., Thomson, K. S., and Pitout, J. D. D. (2004): Use of beta-lactamase inhibitors in disk tests to detect plasmid-mediated AmpC beta-lactamases. J Clin Microbiol. Volume 42, pp.2203-2206. Centers for Disease Control and Prevention (1997): Staphylococcus aureus with reduced susceptibility to vancomycin-United States, 1997. MMWR Morb Mortal Wkly Rep; Volume 46, pp.765-766. Chambers, H.F. and Sachdenim, M. (1990): Binding of beta-lactam antibiotics to penincillin binding protein in methicillin resistant staphylococcus aureus. J infect Dis; Volume 42, pp.1170-1176 Climo MW, Patron RL, Archer GL. (1999): Combinations of vancomycin and beta-lactams are synergistic against staphylococci with reduced susceptibility to vancomycin. Antimicrob Agents Chemother; Volume 43, pp. 1747-53 Cockeril, F.R. (1998): Conventional and Genetic Laboratory Tests Used to Guide Antimicrobial Therapy. Mayo Clin Proc.;L73: pp.1007 –1021 Gill, V.J.; Fedorko,D.P. and Witebsky,F.G. In: Mandell, G.L., Bennett. J.E. and Dolin. R.D., eds. (2000): The Clinician and the Microbiology Lab. Principles and Practice of Infectious Disease. 5th ed. Philadelphia, Pa: Churchill Livingstone; pp.184 –221 Hansen, S.L. and Freedy, P.K. (1984 September): Variation in the abilities of automated, commercial, and reference methods to detect methicillin-resistant (heteroresistant) Staphylococcus aureus. J Clin Microbiol. Volume 20, issue no.3; pp.494–499 Hindler, J.A. and Inderlied, C.B. (1985:February): Effect of the source of Mueller-Hinton agar and resistance frequency on the detection of methicillin-resistant Staphylococcus aureus. J Clin Microbiol. Volume 21,issue number 2; pp.205–210. Hiramatsu K, Hanaki H, Ino T, Yabuta K, Oguri T, Tenover FC. (1997): Methicillin-resistant Staphylococcus aureus clinical strain with reduced vancomycin susceptibility. J Antimicrob Chemother ; Volume 40: pp.135-136 Hiramatsu, K, Aritaka, N., Hanaki, H. et al (1997): Dissemination in Japanese Hospitals of starins of staphylococcus aureus hetero-geneously resistant to vancomycin. Lancet. Vol. 350;pp.1668-1671. Hiramatsu, K. (1998): Vancomycin resistance in staphylococci. Drug resist updates. Volume 1; pp. 135-150. Hiramatsu, K., Hanaki, H., Ino, T., Yabuta, K., Oguri, T. and Tenover, F.C. (1997): Methicillin-resistant Staphylococcus aureus clinical strain with reduced vancomycin susceptibility. J Antimicrob Chemother, Volume 40; pp.135-136. Hiramatsu, K; Kondo, N. and Ito, T. (1996): genetic basis for molecular epidemiology of MRSA. J infect chemother. Volume 2; pp. 117-129. Howe RA, Bowker KE, Walsh TR, Feest TG, MacGowan AP. (1998): Vancomycin-resistant Staphylococcus aureus. Lancet; pp.351:602. Isenberg, H. D. (1992): Clinical microbiology procedures handbook, vol. 1. American Society for Microbiology, Washington, D.C Johnson, A.P. (1998): Intermediate vancomycin resistance in Staphylococcus aureus: a major threat or a minor inconvenience? J. Antimicrob Chemother; Volume 42, pp.289-91. Kaplan, S.L. and Mason, E.O. In: Feigin RD, Cherry JD, eds. (1998): Use of Bacteriology, Mycology, and Parasitology Laboratories. Textbook of Pediatric Infectious Diseases. 4th ed. Philadelphia, Pa: WB Saunders Co; pp.2866 –2873 Komatsuzawa, H.; Sugai, M.; Ohta, K.; Fujiwara, T.; Nakashima, S.; Suzuki, J.; Lee, C. Y. and Suginaka, H. (1997): Cloning and characterization of the fmt gene which affects the methicillin resistance level and autolysis in the presence of triton X-100 in methicillin-resistant Staphylococcus aureus. Antimicrob. Agents Chemother. Volume 41; pp.2355-2361 Kondo, K.; Takaishi, Y. ; Shibata, H. and Higuti, T. (2004): ILSMRs (intensifier of ß-lactam-susceptibility in methicillin-resistant Staphylococcus aureus) from tara [Caesalpinia spinosa (Molina) Kuntze] Levinson, W.E. and Jawetz, E., eds. (1994): Antimicrobial Drugs: Resistance. In: Medical Microbiology & Immunology. 3rd ed. Norwalk, Conn: Appleton and Lange; pp.54 –59 Liu, I. X., Durham, D. G. and Richards, R. M. (2000): Baicalin synergy with ß-lactam antibiotics against methicillin-resistant Staphylococcus aureus and other ß-lactam-resistant strains of S. aureus. J. Pharm. Pharmacol. Volume 52; pp.361-366 McDougal, L.K.; and Thornsberry, C. (1984 April): New recommendations for disk diffusion antimicrobial susceptibility tests for methicillin-resistant (heteroresistant) staphylococci. J Clin Microbiol. Volume 19,issue no. 4; pp.482–488 National Committee for Clinical Laboratory Standards (2000): Methods for dilution antimicrobial susceptibility tests for bacteria that grow aerobically. 5th ed. Approved standard M7-A5. Wayne (PA): The Committee National Committee for Clinical Laboratory Standards (2000): Methods for dilution antimicrobial susceptibility tests for bacteria that grow aerobically. 5th ed. Approved standard M7-A5. Wayne (PA): The Committee National Committee for Clinical Laboratory Standards.( 2000): Methods for dilution antimicrobial susceptibility tests for bacteria that grow aerobically. 5th ed. Approved standard M7-A5. Wayne (PA): The Committee National Committee for Clinical Laboratory Standards.( 2000): Performance standards for antimicrobial disk susceptibility tests. 7th ed. Approved standard M2-A7. Wayne (PA): The Committee. Noble, W.C., Virani, Z. and Cree, R.G. (1992): Co-transfer of vancomycin and other resistance genes from Enterococcus faecalis NCTC 12201 to Staphylococcus aureus. FEMS Microbiol Lett; Vol. 72, pgs. 195-198. Hiramatsu, K., Hanaki, H., Ino, T., Yabuta, K., Oguri, T. and Tenover, F.C. (1997): Methicillin-resistant Staphylococcus aureus clinical strain with reduced vancomycin susceptibility. J Antimicrob Chemother ; vol. 40, pp.135-136. Centers for Disease Control and Prevention (CDC) (1997): Reduced susceptibility of Staphylococcus aureus to vancomycin - Japan, 1996. MMWR; vol. 46, pp. 624-626. Collignon P et al. (1998): Community-acquired methicillin-resistant Staphylococcus aureus in Australia. Lancet, Vol. 352, pp.145 Peacock, J.E. Jr; Moorman, D.R.; Wenzel, R.P.; Mandell, G.L. (1981 December): Methicillin-resistant Staphylococcus aureus: microbiologic characteristics, antimicrobial susceptibilities, and assessment of virulence of an epidemic strain. J Infect Dis. Volume 144,issue number 6; pp.575–582 Ploy MC, Grélaud C, Martin C, de Lumley L, Denis F. (1998): First clinical isolate of vancomycin-intermediate Staphylococcus aureus in a French hospital. Lancet pp.351:1212 Ryan, K.J. and Ray, C.G. In: Ryan, K.J., ed. (1994): Principles of Laboratory Diagnosis of Infectious Disease. Sherris Medical Microbiology.3rd Ed. Norwalk, Conn: Appleton and Lange; pp.221-250 Sato, Y.; Oketani, H.; Singyouchi, K.; Ohtsubo, T.; Kihara, M.; Shibata, H. and Higuti, T. (1997): Extraction and purification of effective antimicrobial constituents of Terminalia chebula RETS against methicillin-resistant Staphylococcus aureus. Biol. Pharm. Bull. Volume 20; pp.401-404 Sato, Y.; Shibata, H.; Arakaki, N. and Higuti, T. (2004): 6,7-Dihydroxyflavone dramatically intensifies the susceptibility to ß-lactam antibiotics in methicillin-resistant and -sensitive Staphylococcus aureus. Antimicrob. Agents Chemother; Volume 48; pp.1357-1360 Sato, Y.;Shibata, H.; Arai, T.; Yamamoto, A.; Okimura, Y.; Arakaki, N. and Higuti,T. (2004b): Variation in synergistic activity by flavone and its related compounds on the increased susceptibility of various strains of methicillin-resistant Staphylococcus aureus to ß-lactam antibiotics. Int. J. Antimicrob. Agents Volume 24; pp.28-35 Shibata, H., Shirakata, C.; Kawasaki, H.; Sato, Y.; Kuwahara, T.; Ohnishi, Y.; Arakaki, N. and Higuti, T. (2003): Flavone markedly affects phenotypic expression of ß-lactam resistance in methicillin-resistant Staphylococcus aureus strains isolated clinically. Biol. Pharm. Bull. T. volume 26; pp. 1478-1483 Shimizu, M.; Shiota, S.; Mizushima, T.; Ito, H.; Hatano, T.; Yoshida, T. and Tsuchiya, T.(2001): Marked potentiation of activity of ß-lactams against methicillin-resistant Staphylococcus aureus by corilagin. Antimicrob. Agents Chemother. Volume 45; pp.3198-3201 Shiota, S.; Shimizu, M.; Mizushima, T.; Ito, H.; Hatano, T.; Yoshida, T. and Tsuchiya, T. (1999): Marked reduction in the minimum inhibitory concentration (MIC) of ß-lactams in methicillin-resistant Staphylococcus aureus produced by epicatechin gallate, an ingredient of green tea (Camellia sinensis). Biol. Pharm. Bull. Volume 22; pp.1388-1390 Shiota, S.; Shimizu, M.; Mizusima, T.; Ito, H.; Hatano, T.;Yoshida, T. and Tsuchiya, T. (2000): Restoration of effectiveness of ß-lactams on methicillin-resistant Staphylococcus aureus by tellimagrandin I from rose red. FEMS Microbiol. Lett. Volume 185; pp.135-138 Sieradzki K, Roberts RB, Haber SW, Tomasz A. (1999): The development of vancomycin resistance in a patient with methicillin-resistant Staphylococcus aureus infection. N Engl J Med; Volume 340; pp.517-523 Smith T, Pearson ML, Wilcox KR, Cruz C, Lancaster ML, Robinson-Dunn B, et al. (1999): Emergence of vancomycin resistance in Staphylococcus aureus: epidemiology and clinical significance. N Engl J Med Volume 340: pp.493-501 Suzuki, J.; Komatsuzawa, H.; Sugai, M.; Ohta, K.; Kozai, K.; Nagasaka, N. and Suginaka, H. (1997): Effects of various types of Triton X on the susceptibilities of methicillin-resistant staphylococci to oxacillin. FEMS Microbiol. Lett. Volume 153; pp.327-331. Tanaka, M.; Wada, N.; Mori-Kurosaka, S.; Chiba, M.Sato, k. and Hiramatsu, K. (1998): In vitro Activity of DU-6859a against methicillin resistant staphylococcus aureus isolates with reduced susceptibility to vancomycin. J antimicrob chemother. Volume 42; pp. 552-553. Tanaka, T.; Okuzum, K.; Iwamoto, T and Hiramatsu, K. (1995): A retrospective study on methicillin resistant staphylococcus aureus clinical strains. Tokyo university hospital. J infect chemother. Vol. 1 pp. 40. Tenover, F.C. (2000): VRSA, VISA, and GISA: the dilemma behind the name game. Clinical Microbiology Newsletter; Volume 22, pp.49-53. Thornsberry, C. and McDougal, L.K. (1983 November): Successful use of broth microdilution in susceptibility tests for methicillin-resistant (heteroresistant) staphylococci. J Clin Microbiol. Volume 18,issue no.5; pp.1084–1091 Utsul, Y. and Tokota,T. (1985): Role of an altered penicillin binding protein in methicillin and Cephem-resistant staphylococcus aureus. Antimicrob agents chemother, Volume 28; pp.397-403. Waldvogel, F.A. (1995): Staphylococcus aureus (including toxic shock syndrome) In: Mandell GL, Bennett JE, Dolin R. Principles and Practice of Infectious Diseases: Fourth Edition; Volume2, pp.1754-777. Working Party of the British Society for Antimicrobial Chemotherapy. Breakpoints in in-vitro antibiotic susceptibility testing. J Antimicrob Chemother 1988;Volume 21;pp.701-10. Yamase, T.; Fukuda, N. and Tajima, Y. (1996): Synergistic effect of polyoxotungstates in combination with ß-lactam antibiotics on antibacterial activity against methicillin-resistant Staphylococcus aureus. Biol. Pharm. Bull. Volume 19; pp.459-465 Zhao, W. H.; Hu, Z. Q.; Okubo, S.; Hara, Y. and Shimamura, T. (2001): Mechanism of synergy between epigallocatechin gallate and ß-lactams against methicillin-resistant Staphylococcus aureus. Antimicrob. Agents Chemother. Volume 45,pp.1737-1742 Read More