Rafael Gómez-Lus
Department of Microbiology, Hospital Clínico Received 25 June 1998Accepted 25 July 1998 Summary Bacterial resistance to antibiotics is often plasmid-mediated and the associated
genes encoded by transposable elements. These elements play a central role in evolution
by providing mechanisms for the generation of diversity and, in conjunction with DNA
transfer systems, for the dissemination of resistances to other bacteria. At the University
Hospital of Zaragoza, extensive efforts have been made to define both the dissemination
and evolution of antibiotic resistance by studying the transferable R plasmids and
transposable elements. Here we describe the research on bacterial resistance to antibiotics
in which many authors listed in the references have participated. The aspects of bacterial
resistance dealt with are: (i) transferable resistance mediated by R plasmids in Gram-
negative bacteria, (ii) R plasmid-mediated resistance to apramycin and hygromycin
in clinical strains, (iii) the transposon Tn1696 and the integron In4, (iv) expression of Department of Microbiology. Hospital Clínico Universitario. Escherichia coli resistance genes in Haemophilus influenzae, (v) aminoglycoside- modifying-enzymes in the genus Mycobacterium with no relation to resistance, and (vi) macrolide-resistance and new mechanisms developed by Gram-positive bacteria Tel.: +34-976761759. Fax: +34-976761693. E-mail: [email protected] characterized the plasmid pUZ1 (initially R1033) from a strain of Pseudomonas aeruginosa coming from the Regional Centerfor Traumatology, near the University Hospital of Zaragoza. The Bacterial resistance to antibiotics in Gram-negative bacilli is plasmid had a size of 68 kb. Its incompatibility group was P (IncP) most commonly mediated by R plasmids and by genes carried and it was transferable to Escherichia coli J62 by conjugation.
by transposons (Tn) and integrons (In). In Gram-positive cocci The plasmid coded for resistance to ampicillin, tetracycline, the conjugative chromosomal transposons are fundamental for gentamicin, kanamycin, streptomycin, chloramphenicol, antibiotic resistance. They are characterized by their related sulfamides and mercuric chloride [19]. The donor as well as features to R plasmids, bacteriophages and the classic the recipient strain coded for the enzymes TEM-1, AAC(3)-I, transposons (for instance, Tn5 and Tn10). All these genetic APH(3’)-I, and ANT(3’’) [9]. Plasmid pUZ1 contained two elements play a central role in bacterial evolution, providing transposons, the already known Tn3, which carries the blaTEM-1 mechanisms to generate diversity and having DNA transfer gene, and a new transposon, Tn1696 [17], which bears the genes systems able to disseminate sequences among other bacteria.
aacC1, aadA1, sul1 and mer, which will be discussed later.
Good examples of those phenomena are the participation of Four years later, we isolated different strains of transposons in the rapid spread of antibiotic resistance among enterobacteria and of P. aeruginosa (Table 1), which contained bacterial populations—above all in those causing nosocomial also plasmids of the IncP group, of the same size, with identical infections—and the role of integrons in the maintenance and restriction patterns and enzymes as pUZ1 [6, 15]. These findings spread of the sul1 gene in enterobacteria.
supported the hypothesis that R plasmids spread to proximalareas, with the advantage of being able to propagate betweenthe Enterobacteriaceae and Pseudomonadaceae, which is In 1976 we detected a 73 kb plasmid which belonged to the group IncM and, thus, only transferable to enterobacteria. The At the University Hospital of Zaragoza and other hospital centres plasmid conferred resistance to ampicillin, tetracycline, gentamicin, in Zaragoza, research has been carried out regarding the evolution tobramycin and dibekacin and produced the enzymes TEM-1 and and spread of antibiotic resistance over the last 30 years. We have AAC(3)-V [14]. Following this, numerous strains of enterobacteria studied conjugative R plasmids (tra+), transposable elements and were isolated which also carried plasmids from the IncM group, the biochemical mechanisms of resistance. In 1974 we with identical properties, genotypes and phenotypes (Table 2).
In one of the strains of Proteus vulgaris (strain 18182) two unit initiated by the blaTEM-1 gene promoter, which is conjugative plasmids were detected. One was from the IncM responsible for the mobilization of one of the IS140.
group (pUZ3a) and the other from the IncP group (pUZ3b). The The capacity for transposition between replicons pattern of resistance transferred corresponds to that described in containing the bla genes is noteworthy, and it is reflected in Tables 2 and 1, respectively. In addition, as a result of the the numerous plasmid incompatibility groups, in which the epidemiological monitoring of the strains carrying R plasmids, bla genes have been detected: 18 for β-lactamase TEM-1, 7 we found that certain plasmids which were initially transferable for TEM-2, 3 for OXA-1, 4 for OXA-2, and 2 for OXA-3.
(tra+) later lost this capacity (tra–). We also found plasmid loss or We have already referred to part of our experiments involving recombination of plasmid genes with the chromosome, which the blaTEM-1 genes isolated from plasmids in the groups IncP and IncM. In France, the gene blaTEM-3 has also been A case to point out was a transconjugant of E. coli characterized in R plasmids from the IncM group. This gene (strain 3644) which had lost the plasmid it was carrying but codes for plasmid β-lactamases of extended spectrum.
had conserved its pattern of resistance. The alternative strategy Previously, in the same country, an IncC plasmid had been was to use the plasmid pUZ8, lacking transposable elements, described which carried a blaTEM-4 gene coding for an and introduce it into the aforementioned transconjugant of E. extended β-lactamase spectrum, named TEM-4.
coli J62 (F– his lac nalR pro trp). Through conjugation withE. coli J53 (F– met pro rifR), a plasmid (pUZ3644) was obtainedwhich contained a 27 kb fragment, carrying the bla aacC5 genes, flanked by two copies of an insertion sequence, IS140. In this segment, the composite transposon Tn2922 was characterized. Tn2922 is a co-integrate capable of transposingas a unit and formed by the fusion of the genes tnpR, blaTEM- In scientific literature there are examples of antibiotic resistance 1 and aacC5 [10]. These two genes make up a transcription genes that have been transported to man by bacteria of animal Table 1 Incompatibility group P plasmids isolated in the Hospital Clínico Universitario, Zaragoza, from 1974 until 1978. Resistance pattern transferred to
Escherichia coli K-12: Ap, Tc, Gm, Km, Cm, Sm, Su*
* Ap, Tc, Gm, Km, Cm, Sm, Su: (ampicillin, tetracycline, gentamicin, kanamycin, chloramphenicol, streptomycin, sulfonamide).
‡ Month/year. ¶ Size of all plasmids: ca. 68 kb. ** Cerebrospinal fluid.
Table 2 Incompability group M plasmids isolated in the Hospital Clínico Universitario, Zaragoza, from 1976 until 1983. Resistance pattern transferred to
Escherichia coli K-12: Ap, Tc, Gm, Tm, Dkb, Nt*
* Ap, Tc, Gm, Tm, Dkb, Nt: (ampicillin, tetracycline, gentamicin, tobramycin, dibekacin, netilmicin).
‡ Month/year. ¶ Size of all plasmids: ca. 73 kb. ** Intensive care units.
origin. It is well known that the correct use of antibiotics in Other genes have used this system, among them the most veterinary medicine is of great concern, as it was reported in prominent are: aadA2, aadB, aacA4, dhfrI, dhfrIIb, dhfrIIc, the Swann Report (1969) (Report of Joint Comittee on the Use dhfrV, oxa1, oxa2, pse2, etc. The majority of dhfr genes for of Antibiotics in Animal Husbandry and Veterinary Medicine, resistance to trimethoprim are found near integrons that also Her Majesty’s Stationery Office, London [1969]). Such a carry the gene sul1, reflecting the joint clinical use of transfer happened with the use of the aminoglycosides trimethoprim and sulfamethoxazol. In Tn1696, the close union apramycin and hygromycin in cattle and pigs respectively. In of the aadA and aacC1 genes stands out, which implies cross- fact, through their application, resistant strains emerged, which resistance on the one hand to streptomycin and spectinomycin were isolated from animals which had undergone treatment.
and on the other to gentamicin, sisomicin and fortimicin. The The resistance was plasmid mediated, and genes were detected stable position of the resistance genes in an integron, present for apramycin-acetyltransferase [AAC(3)-IV] and hygromycin- in an R plasmid, assures their dissemination and the capture of phosphotransferase [APH-(4)-I] enzymes, which inactivated The mechanism of integron mobility is not well known, but In 1989 we were able to confirm the isolation of two clinical there exist at least three possibilities: (i) self mobility, which is strains (Escherichia coli and Klebsiella pneumoniae) resistant deduced from the different genetic locations; (ii) by means of to the abovementioned aminoglycosides, with R plasmids of transposons of the Tn21 family, in which case they are integrated; 110 kb which carried the aacC4 and aph4 genes. Subsequently, and (iii), horizontal transfer of integrons by means of R plasmids four strains of enterobacteria with similar properties were such as the IncP (pUZ1) and IncW (pSa and R388).
isolated. The analysis of these gene organizations showed thatthey were adjacent and grouped in the same orientation as thosewhich had been isolated from animal origin. The two genes for resistance to apramycin and hygromycin form an operon and are associated to IS140 sequences, which implies a transposable structure [18]. Given that the enzyme AAC(3)-IV inactivatesgentamicin and tobramycin, in addition to apramycin, it The efficiency of genetic mechanism of transference is reflected constitutes a potential risk for selecting resistances in clinically in the spread of multiple resistance to sulfonamides, β-lactam used aminoglycosides. Furthermore, the six strains isolated, antibiotics, chloramphenicol, aminoglycosides, etc. The carrying the plasmids pUZ6734, pUZ6743, pUZ6776, pUZ7852 selection can occur simply through the use of one of the and pUZ7874, also showed transferable resistance to ampicillin antimicrobial agents which affects all the genetic determinants.
(β-lactamase TEM-1) and streptomycin [phosphotransferase An interesting example is the resistance of Haemophilus influenzae to chloramphenicol in our geographical region(Aragón) (12%), despite limited clinical use of the drug. Incontrast, 35% were resistant to ampicillin (ApR). And of the chloramphenicol resistant strains, all except one were also resistant to ampicillin. This resistance is mediated by β-lactamases TEM-1. We have not detected any β-lactamase While trying to characterize the mycobacterial version of the of type ROB-1, supporting the previously suggested hypothesis aacC3 gene, the sul3 gene was found, as well as the of a different ROB-1 reservoir for β-lactam resistance such determinants int and tnpA. These discoveries allowed us to re- examine the organization of Tn1696. This 16 kb transposon chloramphenicol is due to the synthesis of an acetyltransferase carries the aacC1 gene and the determinants of resistance to (CAT) which shares biochemical properties with E. coli CAT sulfamides (sul1), chloramphenicol (cmlA), streptomycin- II and III. Some of these strains hybridized with a probe for spectinomycin (aadA1), and mercuric chloride (mer). These cmlA (a gene present in the integron In4 which is located in determinants are located in the integron In4, which has a site- Tn1696), demonstrating permeability changes equivalent to specific recombination system that permits the capture of various cassettes of resistance genes [3]. Figure 1 shows the P. aeruginosa caused by the protein CmlA. In our laboratory, structure of several integrons, among them In4, including the we found in H. influenzae the genes which code for the HI1254 cassettes of genes inserted in the variable region [9]. Near protein, related to the polypeptide CmlA mentioned above, and the recombination site (res), the int gene was found. It belongs for the HI1716 protein, which could participate in altering the to the family of integrases, enzymes essential for recombination.
permeability (Vergara Y, 1996, Ph.D. Thesis, University of In addition, the cmlA gene, the determinant for non-enzymatic Zaragoza). Both of these genes have been characterized in the resistance to chloramphenicol, had a different feature: it has genome of H. influenzae Rd. On the other hand, there is a low its own promoter. This does not occur in other integrons of level of homology between the cmlA gene and the HI1254 protein, which is reflected in weak hybridization with the cmlA Fig. 1 Structural features of various
naturally occurring integrons. Modified
from reference [9]. intI: integrase gene
situated in the 5’ conserved segment;
sul1: sulfonamide resistance gene situated
in the 3’ conserved segment; gene
cassettes: aadB, aadA1, etc. with circles
(O), which represent recombinaton
sites of 59 bp
probe. 37.5% of the H. influenzae strains ApR and CmR were with enterobacteria, and it seems probable that the species also resistant to kanamycin, neomycin and lividomycin through H. parainfluenzae acted as an intermediate.
the production of a phosphotransferase APH(3’)-I. Transfer ofresistance was achieved by electroporation of a competent strain (E. coli DH5α) with the total DNA of H. influenzae, followed by selection with ampicillin and kanamycin. Using electroporation, 180 colonies of E. coli resistant to ampicillinand kanamycin were obtained. DNA was extracted from 9 of The discovery of strains of Mycobacterium fortuitum producing the colonies and in each a 7 kb plasmid was found. Restriction aminoglycoside modifying enzymes [AAC(6’), AAC(3) and enzimes (EcoRI and PstI) analysis demonstrated that the APH(3’)] and in particular acetylases subclass AAC(3)-III, has plasmids were identical. Crude enzyme extracts from the strains not been related to resistance nor to the presence of plasmids.
showed β-lactamase TEM-1 (isoelectric point, pI = 5.4) activity A new enzymatic activity that phosphorylated only streptomycin and 3’-O-phosphotransferase-[APH(3’)-I] activity, which (APH(6)) was demonstrated in strains of M. fortuitum and M. modified kanamycin, neomycin and lividomycin [8].
chelonae. Hybridizations with a probe from Streptomyces griseus The purified phosphotransferase had a molecular weight (aph-6 gene) were positive not only in the new strains that of 26,000 Da and a pI = 5.3, with an aminoacid sequence at produced 6-O-phosphotransferase activity but also in 20 strains the aminoterminal extreme similar to other phosphotransferases that did not possess such phosphorylating activity. This suggested in enterobacteria. Hybridizations with the intragenic probe that the aph-6 gene was localized on the chromosome, that could aph(3’)-Ia, demonstrated that the gene in H. influenzae is not be phenotypically expressed or not, and that did not have any located in the transposon Tn903, which is a feature that relation to streptomycin resistance. The investigation was extended differentiates it from the situation in enterobacteria. In spite of to other fast growing species (M. smegmatis and M. phlei) and to this, it was obvious that the determinants of resistance to slow growing species (M. avium, M. scrofulaceum, M. xenopi ampicillin, chloramphenicol and kanamycin came mostly from and M. terrae), verifying in all a positive hybridization, although E. coli, with peculiarities in genetic reorganization and to a varying degree. Besides, studying a strain of M. aurum, phenotypic expression that reflect the change in bacterial host.
the synthesis of an acetylase AAC(2’) with a characteristic profile In the enterobacteria, the catI, catII and catIII genes are found was detected. The substrates were gentamicin C1 and C1a, mainly in IncM plasmids, whereas cmlA and related genes are netilmicin, 6’-ethyl-netilmicin, tobramycin and kanamycin B, in IncP plasmids. The coexistance of both types of genes in the without modifying the 2’-deamino-kanamycins—kanamycin A genus Haemophilus, situated in plasmids and/or in the and amikacin—nor the 2’-ethyl-netilmicin (Sánchez-Yangüela chromosome, shows the heterogeneity of genetic exchange E, 1990, M.D. Thesis, University of Zaragoza). However, the genes encoding acetylase AAC(3) subclass III or the related activation of aac(2’)-Ib in M. smegmatis mc2 155 allowed the enzyme were not identified, although the enzyme was present in presence of the gene to be correlated with the resistance to all the strains of M. fortuitum that were studied and in the majority lysozyme. Considering this, it is remarkable that the enzyme AAC(2’)-Ia has been shown to contribute to the acetylation of In order to detect and characterize the candidate gene aac3, peptidoglycan (PG) in P. stuartii. Presumably, aminoglycosides probes derived from genes and transposons coding for antibiotic are not the true substrate of AAC(2’)-Ia but are acetylated due resistance were applied to samples of M. fortuitum chromosomal to their structural similarity with PG [13]. As far as we know, DNA. A positive hybridization was obtained with a probe the genes coding for aminoglycoside-modifying enzymes are containing the aacC1 gene, localized in the previously not strictly speaking resistance genes in mycobacteria.
mentioned transposon Tn1696. On determining the nucleotide Paradoxically, if these enzymes are implicated in the sequence of the hybridizing region, similarities were detected biosynthesis of the mycobacterial cell wall, they could act as at the 5’ region of the aacC1 gene. This region comprises the int gene, which encodes an integrase, a determinant for sitespecific recombination. The mycobacterial sequence alsocontained the sul3 gene, a homologue to the sul1 gene present in Tn1696 and in other members of the Tn21 family. The region occupied by the sul3 gene was defined after verifying that itwas flanked by two 880 bp sequences, which were called The resistance of Streptococcus pneumoniae to penicillin is a IS6100, and that it encoded a transposase. These mycobacterial serious problem in Spain. It has become more complicated insertion sequences contained inverted repetitions of 14 bp, since the 1980s with an increase in erythromycin resistance, which differed by only one nucleotide from IS15. The sequence which rose from 0.9% in 1979 to 10% in 1989. Although was similar to IS6, and it is closely related to elements isolated Streptococcus pyogenes remains sensitive to penicillin, in the from Gram-positive and Gram-negative bacteria. The int and last few years more and more strains resistant to macrolides sul3 genes, marked by two IS600 sequences, make up the have been identified. For this reason, epidemiological transposon Tn610 [11]. The codons of the IS6100 transposase gene proved to be similar to those of a gene which encoded the Of the mechanisms that these streptococci have developed main antigenic protein of M. tuberculosis. This suggests that for macrolide resistance, the two fundamental ones are the this 65,000 Da protein originated in mycobacteria.
production of both methyltransferases, encoded by the erm The presence of the sul3 gene seemed obvious, since (A, B, C, TR) genes [2], and the efflux systems, which are the sulfonamides have been used since 1935 and were applied in phenotypic expressions of the mefA (S. pyogenes) and mefB genes the treatment of tuberculosis in the 1950’s. Therefore, finding (S. pneumoniae), and of the msrA (Staphylococcus epidermidis the gene in the M. fortuitum FC1 strain suggested a broad and Staphylococcus aureus) and msrB genes (Staphylococcus diffusion between this species and others from the genus xylosus). The methylases encoded by the erm genes cause Mycobacterium. To test whether this hypothesis was valid, ribosomal modification at the level of 23S RNA subunit 50S, hybridizations were carried out with a probe from the int + sul giving rise to the MLSB phenotype (macrolides, lincosamides, genes (1.4 kb, plasmid pIPC1::Tn610). All the hybridizations streptogramin B), either inducible or constitutive. In our medium, done on 30 strains of M. fortuitum, 21 strains of M. chelonae the genes detected in Gram-positive cocci belonged to different and 125 strains of M. tuberculosis were negative. It seemed classes depending on the genus: Staphylococcus (ermA, ermC, obvious that we had been lucky in choosing M. fortuitum FC1.
ermA + ermC), Enterococcus (ermB) and Streptococcus (ermB), In fact, initially this strain was selected for its intense acetylating the expression of which can be inducible or constitutive, activity AAC(3), and it turned out to house the int, sul3, aph- irrespective of the class of erm gene [7].
6 and blaF genes (Timm J, 1994, Ph.D. Thesis, University of The first description of an efflux mechanism against Paris VII), and also, as we will describe, the aac-2’-Ib gene.
erythromycin was made in 1989, proving the existence of the MS A gene encoding an AAC(2’) was cloned from phenotype (14 and 15 membered macrolides, and streptogramine M. fortuitum. DNA sequencing results identified an B) as the expression of the msrA gene in S. epidermidis. One year ORF(aac(2’)-Ib) coding for a putative protein with a predicted later the gene was characterized and sequenced, demonstrating molecular mass of 28,400 Da [1]. The deduced AAC(2’)-Ib that it formed a complex with two other determinants, smp and protein showed homology to the AAC(2’)-Ia from Providencia stp [16]. In 1992 the msrB gene was cloned and sequenced from stuartii. The presence of the aac(2’)-Ib in all 34 M. fortuitum a plasmid from S. xylosus, which expressed the same phenotype strains was not correlated with any aminoglycoside-resistance [12]. Using initiators M1 and M2 in one strain of group A phenotype. This suggested that the product of this gene streptococcus, a 0.4 kb amplification product was obtained, a size could be implicated in functions other than antibiotic- compatible with the msrA and msrB genes. A gene which showed modification, such as the synthesis of intermediate metabolites, 98% homology to the genes characterized in Staphylococcus was ribosomal proteins, and cell wall components. The insertional cloned and sequenced from the DNA band (Adrián FJ, 1997, Ph.D. Thesis, University of Zaragoza).
integron of Tn1696: similarity of the product to transmembrane transport A new phenotype M has been described in Streptococcus 4. Clancy J, Petitpas JW, Dib-Hajj F, Bergeron J, Yuan W, Cronan M, Kamath pneumoniae and S. pyogenes, which produces resistance AV, Retsema JA (1996) Molecular cloning and functional analysis of a exclusively to 14 and 15 membered macrolides. Investigating novel macrolide-resistant determinant, mefA, from Streptococcus pyogenes.
this mechanism, a new class of genes was characterized called mef (for macrolide efflux): mefA in S. pyogenes and mefE in 5. Clancy J, Dib-Hajj F, Petitpas JW, Yuan W (1997) Cloning and S. pneumoniae [4, 20]. In 1997 a new efflux gene mreA (for characterization of a novel macrolide efflux gene, mreA, fromStreptococcus agalactiae. Antimicrob Agents Chemother 41:2719–2723 macrolide resistance efflux) was described in Streptococcus 6. Gómez-Lus R, Larrad L, Rubio-Calvo MC, Navarro M, Lasierra MP (1980) AAC(3) and AAC(6’) enzymes coded by R plasmid isolated in a In our laboratory, using primers specific for the mefA and general hospital. In: Mitsuhashi S, Rosival L, Krcmery V (eds) Antibiotic mefE genes we have confirmed the efflux mechanism in the resistance. Heidelberg: Springer-Verlag, pp 295–303 majority of M phenotype strains of the species S. pyogenes, 7. Gómez-Lus S, Gómez-Lus R (1993) Detection of new macrolide- lincosamide-streptogramin resistance phenotypes in gram-positive cocci S. pneumoniae and different species of the viridans group. The and its genetic basis. In: Einhorn J, Nord CE, Ragnar Norrby S (eds) differentiating feature is that the ermB gene is much more Recent Advances in Chemother. Washington, DC: American Society for prevalent in the S. pneumoniae population with MLSB phenotype. Whereas in S. pyogenes the M phenotype expression 8. Gómez-Lus R, Vergara Y (1995) Aminoglycoside resistance in of the mefA gene is dominant. The M phenotype is infrequent Haemophilus influenzae. J Chemother 40:97–99 9. Hall RM, Collis CM (1995) Mobile cassettes and integrons: capture and in pneumococci in our geographical region, in contrast to what spread of genes by site-specific recombination. Molec Microbiol 15:593–600 has been described in other geographical regions. In almost all Martín C, Gómez-Lus R, Ortiz JM, García-Lobo JM (1987) Structure strains the gene mefE was detected.
and mobilization of an ampicillin and gentamicin resistance determinant.
Other resistance mechanisms such as enzymatic Antimicrob Agents Chemother 31:1266–1270 modification (acetylation, phosphorylation, glycosylation and 11. Martín C, Timm J, Rauzier J, Gómez-Lus R, Davies J, Gicquel B (1990) Transposition of an antibiotic resistance element in mycobacteria. Nature nucleotidation) and hydrolysis (esterases), have minor practical significance in Gram-positive bacteria.
12. Milton ID, Hewitt CL, Harwood CR (1992) Cloning and sequencing of The transfer of macrolide resistance genes in S. pneumoniae a plasmid-mediated erythromycin resistance determinant from is mediated by conjugative chromosomal transposons with a broad Staphylococcus xylosus. FEMS Microbiol Lett 97:141–148 host spectrum: the genera Streptococcus, Staphylococcus and 13. Payie KG, Rather PN, Clarke AJ (1995) Contribution of gentamicin 2’-N-acetyltransferase to the O acetylation of peptidoglycan in Providencia Enterococcus. Given that resistance to chloramphenicol, stuartii. J Bacteriol 177:4303–4310 tetracyclines and aminoglycosides (high level) is also transmitted 14. Rivera MJ, Vitoria MA, Navarro M, Robledano L, Chocarro P, Gómez- by conjugative transposons, these antibiotics can indirectly co- Lus R (1984) Hospital dissemination among Gram-negative bacillus select erythromycin, as is the case with Tn1545, which carries the strains of an IncM plasmid encoding an AAC(3) and a TEM-1 erm, tet, and aph-3’ genes. In contrast, the mutation of the pbp β-lactamase. Drugs Exptl Clin Res 11:789–795 15. Rivera MJ, Martín C, Gómez-Lus ML, Robledano L, Otal I, Gómez-Lus genes and transformation by pbp genes from other species, R (1988) Characterization of a hospital disseminated plasmid encoding constitutes the well-documented mechanism of penicillin resistance resistance to gentamicin and other antimicrobial agents. Microbiología in pneumococci. S. pyogenes exchanges resistance genes by means of conjugative transposons, for R plasmids and for bacteriophages 16. Ross JI, Eady EA, Cove JH, Cunliffe WJ, Baumberg S, Wooton JC (1990) (transduction). Nevertheless, studies on resistance gene transfer Inducible erythromycin resistance in staphylococci is encoded by amember of the ATP-binding transport super-gene family. Molec Microbiol made in hospitals tend to be retrospective, hence we must try to examine the process while it is happening, and track the in vivo 17. Rubens CE, McNeill WF, Farrar WE (1979) Transposable plasmid traffic of R plasmids, transposons and integrons.
deoxyribonucleic acid sequence in Pseudomonas aeruginosa whichmediates resistance to gentamicin and four other antimicrobial agents. JBacteriol 139:877–888 18. Salauze D, Otal I, Gómez-Lus R, Davies J (1990) Aminoglycoside acetyltransferase 3-IV (aaC4) and hygromycin B 4-I phosphotransferase(hphB) in bacteria isolated from human and animal sources. Antimicrob Aínsa JA, Martín C, Gicquel B, Gómez-Lus R (1996) Characterization of the chromosomal aminoglycoside 2’-N-acetyltransferase gene from 19. Smith DI, Gómez-Lus R, Rubio-Calvo MC, Datta N, Jacob A, Hedges Mycobacterium fortuitum. Antimicrob Agents Chemother 40:2350–2355 RW (1975) Third type of plasmid conferring gentamicin resistance in Arthur M, Molinas C, Mabilat C, Courvalin P (1990) Detection of Pseudomonas aeruginosa. Antimicrob Agents Chemother 8:227–230 erythromycin resistance by the polymerase chain reaction using primers 20. Sutcliffe J, Tait-Kamradt A, Wondrack L (1996) Streptococcus in conserved regions of erm rRNA methylase genes. Antimicrob Agents pneumoniae and Streptococcus pyogenes resistant to macrolides but sensitive to clindamycin: a common resistance pattern mediated by an Bissonnette L, Champetier C, Buisson JP, Roy PH (1991) Characterization efflux system. Antimicrob Agents Chemother 40:1817–1824 of the non-enzymatic chloramphenicol resistance (cmlA) gene of the In4


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My lecture outline o Describe a brief history of the development of hospice care o Describe the hospice philosophy of care o Describe the referral process o Describe hospice services o Describe reimbursement in hospice care o Recognize hospice myths • Course outline based on Objectives: o Describe a brief history of the development of hospice care, especially ƒ Dame Cice

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