Gentamicin-resistant Bacteria Research Articles

Prevalence and Characterization of Gentamicin Resistance Genes in Escherichia coli Isolates from Beef Cattle Feces in Japan.

Gentamicin is an important antibiotic for the treatment of opportunistic infections in the clinical field. Gentamicin-resistant bacteria have been detected in livestock animals and can be transmitted to humans through the food supply or direct contact. We have previously revealed that gentamicin-resistant Escherichia coli are distributed at a comparatively high rate from beef cattle in Japan, but few studies have focused on the molecular epidemiology of gentamicin-resistant bacteria. To understand these bacteria, this study examined the prevalence of various gentamicin resistance genes in gentamicin-resistant E. coli isolates from beef cattle feces. Of the 239 gentamicin-resistant E. coli isolates, the presence of the aacC2, aadB, or aac(3)-VIa genes was confirmed in 147, 84, and 8 isolates, respectively. All aac(3)-VIa-harboring isolates had an MIC value of 64μg/mL for gentamicin and exhibited resistance to 11 antibiotic agents. An analysis of the representative aac(3)-VIa-harboring E. coli strain GC1-3-GR-4 revealed that the aac(3)-VIa gene was present on the IncA/C plasmid together with the aadA and blaCMY genes. Furthermore, the upstream region of the aac(3)-VIa gene contained the aadA gene and the class 1 integron-integrase gene (intI1). The aac(3)-VIa gene was detected for the first time in Japan and is expected to be able to transfer between bacteria via the IncA/C plasmid and integron. These results reveal the expansion of the distribution or diversity of gentamicin resistance genes in Japan.

Intensive care unit time and prolonged enucleation to processing interval are associated with donor cornea contamination.

To determine donor cornea contamination rate and to determine factors associated with cornea contamination. To assess the effect of hospitalization, intensive care unit (ICU) time, and antibiotic use on corneal contamination rate. To determine the spectrum of the contaminating microorganisms. The contamination rate of 212 corneas, obtained by enucleation from April 2014 to January 2015 in a single eye bank, was assessed retrospectively according to age, sex, cause of death, systemic antibiotic use, hospitalization time, ICU time, mechanical ventilation (MV), death to enucleation interval (DEI), enucleation to processing interval (EPI), and corneal epithelial exposure grading. The relative risk (RR) and adjusted RR with a 95% confidence interval were calculated using IBM-SPSS 20.0. The contamination rate was 35.6% (n = 75). On multivariate analysis, ICU stay of 4days or longer and enucleation to processing interval (EPI) greater than 7.4h (RR 1.58, CI 0.96-2.60, P= 0.06) were associated with donor cornea contamination. Corneal contamination risk was highest from 4 to 6days at the ICU (RR 3.40, CI 1.54-7.51, P< 0.01) and decreased after 7days (RR 2.22, CI 1.00-4.93, P= 0.05). Coagulase-negative Staphylococcus was the most common isolated bacteria (69.6%). The frequency of gentamicin-resistant bacteria was higher among patients who stayed 4days or longer at the ICU. Patients staying at the intensive care unit 4days or longer showed increased risk of corneal contamination. This is an important result to consider further indication for cornea donation.

An antibiotic loaded ceramic sternum to treat destroyed infected sternum: 4 cases.

BackgroundAfter its destruction during refractory deep sternal wound infection (DSWI), current sternum reconstructions mainly rely on muscle flaps technique, but such technique have pitfalls and limits. To tackle the limited possibilities to use device implantation because of the risk of infection, we developed a self-protected device allowing its implantation in an infected area.MethodsWe used gentamicin alone or in combination with vancomycin loaded in a porous ceramic sternum to replace sternums destroyed during DSWI. The aim was to mechanically replace the sternum and to secure the implantation by killing the remaining bacteria in the wound thanks to the loaded antibiotic.ResultsThis device was implanted in four infected patients during DWSI with sternal dehiscence. No complication occurred during surgeries, and wound healing was obtained quickly. Local antibiotic concentrations largely exceeded the ones needed for their efficacy while no antibiotic was found in the blood. All patients are well-being. However previously unknown gentamicin resistant bacteria, present in the surgical wound at the time of positioning, required sternal implant removal for one patient after 19 months. For all patients, pulmonary function tests (PFT) improved after implantation.ConclusionsThe ceramic sternum played its role consolidating the thoracic cage without stiffening. The antibiotic loaded in the sternum allowed a secure implantation, killing bacteria before the colonization of the implant even in this infected area. These four implantations are promising for patients with sternal destruction after DSWI.

Physico-chemical and bacteriological screening of Hussain Sagar lake: An urban wetland

Hussain Sagar is a man-made lake originally designed for drinking water purpose that receives domestic sewage and industrial effluents through drainage canals due to rapid residential and industrial growth. Also, every year thousands of idols are immersed into the lake during festivals. In this context, a comprehensive study was initiated for monitoring the lake water quality. Various physico-chemical parameters such as temperature, pH, EC, TDS, COD and chlorophyll a were analyzed using APHA standard methods Water was also monitored for heterotrophic bacteria, total coliforms, Escherichia coli and antibiotic resistant bacteria. The average values for heterotrophs were found to be 8.6 × 104 and 2.8 × 104 CFU/mL before and after idol immersion, respectively. While the average values for total coliforms and E. coli were 5 × 104 and 5 × 102; 1.2 × 104 and 7.2 × 101 CFU/mL, for the respective sampling periods. The mean values for ampicillin and gentamicin resistant bacteria were 5.9 × 103 and 6.9 × 102; and 2.2 × 103 and 5.4 × 102 CFU/mL, respectively. It was found that TDS, COD and chlorophyll a values were decreased after idol immersion due to extensive cleaning. The statistical results showed no correlation between faecal bacteria and physico-chemical parameters and one way-ANOVA indicated statistically significant differences between the mean values of different sampling locations, with respect to COD and E. coli at 95% confidence. However, enormous load of coliforms and E. coli indicated severe contamination of the lake with domestic sewage and human excreta. Thus, the water is not suitable for human consumption/drinking purpose. Notably, incidence of antibiotic resistant bacteria in lake water is a potential threat to both public health and the environment. Thus, regular monitoring and applying appropriate corrective actions are needed to improve the water quality.

2-stage revision of 120 deep infected hip and knee prostheses using gentamicin-PMMA beads

Background and purpose — A 2-stage revision is the most common treatment for late deep prosthesis-related infections and in all cases of septic loosening. However, there is no consensus about the optimal interval between the 2 stages.Patients and methods — We retrospectively studied 120 deep infections of total hip (n = 95) and knee (n = 25) prostheses that had occurred over a period of 25 years. The mean follow-up time was 5 (2–20) years. All infections had been treated with extraction, 1 or more debridements with systemic antibiotics, and implantation of gentamicin-PMMA beads. There had been different time intervals between extraction and reimplantation: median 14 (11–47) days for short-term treatment with uninterrupted hospital stay, and 7 (3–22) months for long-term treatment with temporary discharge. We analyzed the outcome regarding resolution of the infection and clinical results.Results — 88% (105/120) of the infections healed, with no difference in healing rate between short- and long-term treatment. 82 prostheses were reimplanted. In the most recent decade, we treated patients more often with a long-term treatment but reduced the length of time between the extraction and the reimplantation. More reimplantations were performed in long-term treatments than in short-term treatments, despite more having difficult-to-treat infections with worse soft-tissue condition.Interpretation — Patient, wound, and infection considerations resulted in an individualized treatment with different intervals between stages. The 2-stage revision treatment in combination with local gentamicin-PMMA beads gave good results even with difficult prosthesis infections and gentamicin-resistant bacteria.

Occurrence of airborne vancomycin- and gentamicin-resistant bacteria in various hospital wards in Isfahan, Iran.

Background:Airborne transmission of pathogenic resistant bacteria is well recognized as an important route for the acquisition of a wide range of nosocomial infections in hospitals. The aim of this study was to determine the prevalence of airborne vancomycin and gentamicin (VM and GM) resistant bacteria in different wards of four educational hospitals.Materials and Methods:A total of 64 air samples were collected from operating theater (OT), Intensive Care Unit (ICU), surgery ward, and internal medicine ward of four educational hospitals in Isfahan, Iran. Airborne culturable bacteria were collected using all glass impingers. Samples were analyzed for the detection of VM- and GM-resistant bacteria.Results:The average level of bacteria ranged from 99 to 1079 CFU/m3. The highest level of airborne bacteria was observed in hospital 4 (628 CFU/m3) and the highest average concentration of GM- and VM-resistant airborne bacteria were found in hospital 3 (22 CFU/m3). The mean concentration of airborne bacteria was the lowest in OT wards and GM- and VM-resistant airborne bacteria were not detected in this ward of hospitals. The highest prevalence of antibiotic-resistant airborne bacteria was observed in ICU ward. There was a statistically significant difference for the prevalence of VM-resistant bacteria between hospital wards (P = 0.012).Conclusion:Our finding showed that the relatively high prevalence of VM- and GM-resistant airborne bacteria in ICUs could be a great concern from the point of view of patients' health. These results confirm the necessity of application of effective control measures which significantly decrease the exposure of high-risk patients to potentially airborne nosocomial infections.

Risk Factors and Significance of Gentamicin Resistant Positive Urine Culture in Patients Undergoing Urological Surgery: A Short Report

We assessed patients who had pre-operative urine that grew gentamicin-resistant bacteria but were given gentamicin prophylaxis because urine result was not available. Our aim was to identify postoperative-sepsis rates, risk factors to acquire resistant-bacteria, and to optimize our prophylactic regime. Total 4,933 pre-operative urine-samples were reviewed and those positive for E.coli, Klebsiella or Proteus (n = 979) were analysed. Forty-four (4.4%) had gentamicin-resistant bacteria. Of those, 8 were immunosuppressed, 38 (86%) had a recent urological procedure and 29 (66%) had received recent antibiotics. Eighteen (41%) had a urinary catheter and 11 (25%) had double J stent. Three patients (7%) developed post-operative sepsis/febrile urinary tract infection. Although the majority of gentamicin-resistant samples represent colonization, the incidence of post-operative sepsis was significant. Amikacin may be a superior alternative. Our new protocol aims to pre-operatively identify patients at risk of prophylaxis failure with gentamicin and select amikacin as an alternative.

Practical survey on antibiotic-resistant bacterial communities in livestock manure and manure-amended soil

Through livestock manure fertilization, antibiotics, antibiotic-resistant bacteria and genes are transferred to agricultural soils, resulting in a high prevalence of antibiotic-resistant bacteria in the soil. It is not clear, however, whether a correlation exists between resistant bacterial populations in manure and manure-amended soil. In this work, we demonstrate that the prevalence of cephalexin-, amoxicillin-, kanamycin- and gentamicin-resistant bacteria as well as bacteria simultaneously resistant to all four antibiotics was much higher in manure-amended soils than in manure-free soil. 454-pyrosequencing indicated that the ARB and multiple antibiotic-resistant bacteria (MARB) in swine or chicken manure and manure-amended soil were mainly distributed among Sphingobacterium, Myroides, Enterococcus, Comamonas and unclassified Flavobacteriaceae. The genus Sphingobacterium was highly prevalent among ARB from swine manure and manure-amended soil, and was also the most dominant genus among MARB from chicken manure and manure-amended soil. Other dominant genera among ARB or MARB populations in manure samples, including Myroides, Enterococcus and Comamonas, could not be detected or were detected at very low relative abundance in manure-amended soil. The present study suggests the possibility of transfer of ARBs from livestock manures to soils and persistence of ARB in these environments.

‘To bead or not to bead?’ Treatment of osteomyelitis and prosthetic joint-associated infections with gentamicin bead chains

Gentamicin-containing polymethylmethacrylate (PMMA) beads are frequently used to prevent and treat orthopaedic infections. The beads are typically inserted to fill anatomical defects secondary to surgical debridement. Local gentamicin use results in low serum levels whilst achieving high concentrations at the site of infection. However, a systematic review of the available literature showed that, despite these theoretical advantages, no prospective study has thus far proven gentamicin-containing PMMA beads to be effective in treating orthopaedic infections. Available studies are based on small patient numbers and do not show significantly better results when local and parenteral antibiotics are combined compared with systemic therapy alone. These poor results may be explained partially by reduced aminoglycoside efficacy when biofilms or gentamicin-resistant bacteria are present. Moreover, little is known regarding the potential side effects of gentamicin-containing beads. In this paper, the pros and cons regarding the use of gentamicin-containing PMMA beads are discussed. It is concluded that more well-executed, prospective studies are needed to settle the discussion on the use of gentamicin-containing beads in the treatment of orthopaedic infections.

Spraying of oxytetracycline and gentamicin onto field-grown coriander did not affect the abundance of resistant bacteria, resistance genes, and broad host range plasmids detected in tropical soil bacteria

Horticultural supplements containing oxytetracycline and gentamicin, two clinically relevant biocides, are widely marketed to prevent or control infections by bacterial phytopathogens. Despite their regular consumption in the world’s less developed countries, it is unknown whether exposure of tropical farmlands to these drugs results in an enrichment of resistant bacteria, resistance genes, and/or mobile genetic elements in the soil. These concerns were investigated under field conditions by repeatedly spraying recommended amounts of a commercial product containing oxytetracycline-HCl, and gentamicin-\({\text{SO}}^{{ - 2}}_{4} \) onto two coriander plots. Subsequent to five applications within 16 months, composite soil samples from control and treated sections were compared with respect to the abundance of resistant bacteria and the prevalence of conserved nucleotide sequences from tetracycline efflux proteins, tetracycline ribosomal protection proteins, four different families of gentamicin-modifying enzymes, and broad host range plasmids of the IncP-1 and IncQ incompatibility groups. The isolation frequency of oxytetracycline- and gentamicin-resistant bacteria and the detection rate of the aforementioned genes and elements were unrelated to application of the supplement. Despite the omnipresence of sequences from IncP-1 plasmids, conjugative plasmids conferring resistance to oxytetracycline or gentamicin were not captured in biparental matings. The widespread occurrence of resistant bacteria and resistance genes at the beginning of the trial emerges as a reasonable explanation for the lack of anticipated responses. Moreover, we assume that the biocides applied were inactivated by biotic and abiotic factors under tropical conditions.

Lettuce for human consumption collected in Costa Rica contains complex communities of culturable oxytetracycline- and gentamicin-resistant bacteria.

The present widespread use of antimicrobials in crop farming is based upon their successful application in human medicine. However, recent evidence suggests that the massive anthropogenic release of antimicrobials into the biosphere has selected for resistant bacteria and facilitated the transfer of resistance genes among them. This work deals with the examination of iceberg lettuce collected at 10 farms from two regions in Costa Rica. Farmers from nine sampling sites regularly apply commercial formulations containing gentamicin, oxytetracycline, streptomycin, or a combination of them without being able to indicate how often and how much of these products have been sprayed onto the crops. One organic farm was also investigated for comparative purposes. Oxytetracycline- and gentamicin-resistant bacteria were abundantly detected using selective enrichment cultures. Furthermore, colony mixtures from selective plates were characterized by chemotaxonomical and molecular fingerprinting methods. Both types of resistant communities accounted for a significant fraction of all culturable bacteria and included several resistance genes as well as factors for their potential horizontal transfer. Given the fact that lettuce is eaten raw, it may contribute to the dissemination of antimicrobial-resistant bacteria and/or their resistance genes from the environment to the microbial biota of the human intestine.

Role of Microfilaments and Microtubules in the Invasion of EPC Cells by Aeromonas hydrophila

The influence of the cytoskeleton on the invasion of Aeromonas hydrophila strain AhJ-1, isolated from diseased fish, in the monolayer cell of epithelioma papillosum cells of carp (EPC) was evaluated by the recovery of gentamicin-resistant bacteria from Triton X-100 cell lysates. The depolymerization of microfilaments (MF) by cytochalasin B and D inhibited the uptake of A. hydrophila in a dose-dependent manner and that of microtubules (MT) by colchicines and nocodazole did not affect the invasion of A. hydrophila in EPC cells significantly. The invasion frequency decreased approximately 62% with the addition of 0.1 microg/ml cytochalasin D and nearly 86% by the addition of 5.0 microg/ml. Invasion decreased approximately 49% and 83% by addition of cytochalasin B in a concentration of 2.5 microg/ml and 10.0 microg/ml. Colchicine and nocodazole, inhibitors of MT formation appears to have little effect on the invasion of EPC cells by strain Ah J-1. Thus MF formation, but not MT formation seems to play an important role in the internalization of A. hydrophila J-1.

Making sense of antimicrobial use and resistance surveillance data: application of ARIMA and transfer function models

Making sense of antimicrobial use and resistance surveillance data: application of ARIMA and transfer function models

Aminoglycoside resistance. Enzymatic mechanisms in clinical bacterial strains in Slovakia during the last decade

We determined the importance and dissemination of enzymatic mechanisms of aminoglycoside resistance in 239 gentamicin-resistant strains of Gram-negative bacilli isolated in Slovakia during the past decade. Over the past 5 years, the resistance to tobramycin has risen by 1.1%, to netilmicin by 27.4%, to amikacin by 78.3%, and resistance to isepamicin was high too (81.9%). Sixteen different aminoglycoside-modifying enzymes were detected and their significance for resistance to aminoglycosides was confirmed. It is evident that within the past 5 years gentamicin-resistant bacteria have obtained genetic information on resistance to the other aminoglycosides tested and the percentage of strains with different enzyme combinations has risen. These observations confirm the continuously increasing complexity of antibiotic resistance in clinical bacterial isolates under study in Slovakia during the last decade.

Efficacy and Safety of Gentamicin and Streptomycin in Optisol-GS, a Preservation Medium for Donor Corneas

Increasing reports of gentamicin-resistant bacteria contaminating donor corneas and causing endophthalmitis have indicated that preservation of corneal storage media with 100 micrograms/ml of gentamicin alone needs reevaluation. We investigated the stability and possible cytotoxicity of streptomycin as a supplement to gentamicin in Optisol corneal storage medium. The combination of gentamicin and streptomycin in Optisol solution was stable at room temperature for at least 4 weeks and inhibited the growth of Staphylococcus aureus, S. epidermidis, alpha hemolytic streptococci, Streptococcus Group D, Propionibacterium acnes, Escherichia coli, and diphtheroids, but not Pseudomonas aeruginosa. The addition of vancomycin did not significantly improve the antibacterial effectiveness of the gentamicin and streptomycin combination when stored at 4 degrees C. The growth of 15 of 20 clinical ocular isolates of Ps. aeruginosa was suppressed by the gentamicin-streptomycin combination. Streptomycin in concentrations of up to 1,000 micrograms/ml did not decrease the mitotic activity of corneal endothelial cells as evaluated by the in vitro incorporation of tritiated thymidine or cause cytotoxicity. The addition of 200 micrograms/ml of streptomycin to Optisol corneal storage medium containing 100 micrograms/ml of gentamicin may significantly improve activity against gentamicin-sensitive and gentamicin-resistant contaminants.

Escherichia coli expressing a Neisseria gonorrhoeae opacity-associated outer membrane protein invade human cervical and endometrial epithelial cell lines.

Members of the opacity-associated (Opa) outer membrane protein family of Neisseria gonorrhoeae have been proposed to mediate adherence to and invasion of cultured human epithelial cells. We transformed Escherichia coli with a plasmid containing a gonococcal opa gene fused in-frame to the leader sequence of the beta-lactamase gene as described by Palmer et al. [Palmer, L., Brooks, G. F. & Falkow, S. (1989) Mol. Microbiol. 3, 663-671]. These transformed E. coli [E. coli (opa)] expressed the heat-modifiable opa gene product (the Opa protein) in their outer membrane and adhered to and invaded ME-180 human endocervical epithelial cells. In a 2-h adherence assay, an average of 26.7 E. coli (opa) adhered per ME-180 cell, whereas the control E. coli carrying only the expression vector (pKT279) did not adhere at all (less than 0.15 bacterium per cell). We investigated the ability of the adherent E. coli (opa) to invade ME-180 epithelial cells by using a gentamicin selection assay. We recovered up to 1 x 10(6) gentamicin-resistant bacteria per monolayer when ME-180 cells were infected with E. coli (opa) compared to less than 10 bacteria when the epithelial cells were infected with the same number of control E. coli (pKT279). The kinetics and level of invasion by E. coli (opa) were similar to invasion by Opa+ N. gonorrhoeae. Maximum invasion occurred 4 h after infection with 4 x 10(7) bacteria. Transmission electron microscopy studies confirmed that E. coli (opa) invaded ME-180 cells. In comparative studies, the number of E. coli (opa) that invaded HEC-1-B human endometrial epithelial cells was about an order of magnitude less than the number that invaded ME-180 cells, and E. coli (opa) did not invade Chang human conjunctival epithelial cells at all. The observations that early (less than 4 h) invasion by E. coli (opa) was dramatically inhibited, in a dose-responsive manner, by the actin-disrupting reagent cytochalasin D but later invasion (8-24 h) was not suggest that invasion mediated by Opa proteins may occur by two mechanisms, only one of which is dependent upon microfilament function. Transmission electron microscopy also revealed that infected epithelial cells had a dramatically increased amount of cytoplasmic fibrillar material surrounding the nucleus. The function and genesis of this material remain unclear. These studies indicate that at least one gonococcal Opa protein is an invasin.

Donor Cornea Bacterial Contamination

The incidence of culture-positive cornea rims from 446 consecutive donor corneas cultured at the Doheny Eye Institute between 1986 and 1988 was determined. Both the identity and antibiotic sensitivities of the contaminating organisms were reviewed. Sixty-three (14.1%) of 446 cornea rims were culture-positive, but none of the 63 patients who received these contaminated donor corneas developed endophthalmitis. Streptococcus (26 of 63), Propionibacterium (15 of 63), Stphylococcus (14 of 63) species and diptheroids (8 of 63) were the most common cornea rim contaminants, and in most cases were resistant to gentamicin (i.e., 21 of 26 or 81%, 9 of 15 or 60%, 10 of 14 or 71%, 4 of 8 or 50%, respectively). Virtually all of the gentamicin-resistant bacteria isolated from cornea rims were found to be sensitive to vancomycin. Eye banks should consider the addition of other antibiotics to storage media to reduce donor cornea contamination. Surgeons performing corneal transplantation should also consider these results when selecting antibiotics for use at the time of surgery and in the postoperative period.

Penetration and replication of Edwardsiella spp. in HEp-2 cells

The ability of 22 Edwardsiella strains to penetrate and replicate in cultured epithelial cells was initially evaluated by light microscopy methods and by the recovery of gentamicin-resistant (Gmr) bacteria from the Triton X-100 cell lysates of HEp-2-infected monolayers. Giemsa-stained HEp-2 cells revealed the presence of numerous internalized bacteria 3 h postinfection, often appearing as parallel rows of replicated bacteria within the cytosol and sometimes obliterating the cytoplasm because of the large numbers of bacilli present. Invasive bacteria were also sometimes found within cytoplasmic vacuoles in infected cells; thin-section electron micrographs of HEp-2-infected cells supported these conclusions. Results of light microscopy studies and cell lysate assays indicated that most Edwardsiella tarda (92%) and some Edwardsiella hoshinae strains were invasion positive on one or more occasions, while Edwardsiella ictaluri isolates were uniformly negative. HEp-2 invasion by E. tarda was a microfilament-dependent (cytochalasin B- and D-sensitive) process, with maximum numbers of Gmr CFU recorded between 3 and 6 h postinfection. The small percentage (0.01 to 1.0%) of the challenge inoculum recoverable as Gmr progeny 3 to 6 h postinfection was attributed to a strong cell-associated (not filterable) hemolysin that was produced by a majority (85%) of the E. tarda strains but not by E. ictaluri and only minimally by E. hoshinae. This cytolysin/hemolysin was responsible for the toxic effects observed in HEp-2 cells during the infection-replication process of edwardsiellae and appears to play a role in the release of internalized and replicated bacteria from infected cells. The results suggest an invasion strategy with some similarities to and differences from those of other recognized enteroinvasive pathogens.

The elimination of gentamicin-resistant gram-negative bacteria in a newborn intensive care unit.

The aminoglycosides play a central role in the treatment of infectious diseases caused by gram-negative bacteria. During the period of January to June 1984, 45 clinical specimens collected in our neonatal intensive care unit grew Enterobacter cloacae; 41 of them were gentamicin resistant. One neonate developed septicemia. The routine antibiotic protocol was then changed from gentamicin-ampicillin to amikacin-ampicillin for a period of six months. During this period the resistance to gentamicin declined to a minimum. Only eight of 122 specimens proved to harbor gram-negative organisms resistant to gentamicin. The gentamicin-resistant E. cloacae vanished. No isolate was resistant to amikacin. The gentamicin-ampicillin regimen was then reintroduced.

Antibiotic Resistance Patterns During Aminoglycoside Restriction

When amikacin first became available its use was restricted to prevent the emergence of resistant strains of gram-negative bacilli to this new agent. Gentamicin was the aminoglycoside most widely used at this time, and the incidence of gentamicin-resistant bacteria was 14%, while only 2.4% were resistant to amikacin. For a period of 15 months gentamicin use was restricted, and amikacin was used almost exclusively. Amikacin use was associated with a fall in the incidence of gentamicin-resistant bacteria to 9.2% (p less than .00005), while amikacin resistance remained unchanged at 2.2% (NS). During a period of 21 months after all aminoglycoside restrictions were lifted, gentamicin use again increased, and was accompanied by a return of gentamicin resistance to the baseline level of 15.3%. During this period, amikacin resistance also increased to 4.0% (p less than .0000001) but was due primarily to an increase in resistant Pseudomonas aeruginosa. Escherichia coli was the most frequently isolated gram-negative bacillus during all three periods, and it remained sensitive to both antibiotics regardless of the drug in use. In contrast, P. aeruginosa showed a high level of resistance to gentamicin, which fell when this antibiotic was restricted, only to return to a high level with reinstitution of gentamicin. While there was also an increase in amikacin resistant strains of P. aeruginosa with unrestricted aminoglycoside use, there was no apparent shift in the pattern of aminoglycoside modifying enzymes among a small random selection of amikacin-resistant bacteria. Impaired uptake of antibiotic was the predominant mechanism responsible for P. aeruginosa resistance among strains that did not produce aminoglycoside acetyltransferase (AAC)(6').