The value of respect in human research ethics: a conceptual analysis and a practical guide.

In order to continue to maintain public trust and confidence in human research, participants must be treated with respect. Researchers and Human Research Ethics Committee members need to be aware that modern considerations of this value include: the need for a valid consenting process, the protection of participants who have their capacity for consent compromised; the promotion of dignity for participants; and the effects that human research may have on cultures and communities. This paper explains the prominence of respect as a value when considering the ethics of human research and provides practical advice for both researchers and Human Research Ethics Committee members in developing respectful research practices.

The -521 C/T variant in the dopamine-4-receptor gene (DRD4) is associated with skiing and snowboarding behavior.

Sensation seeking is the tendency to seek out new and thrilling experiences and to take risks for the sake of such experiences. A single-nucleotide polymorphism, -521 C/T (rs1800955) in the promoter region of the dopamine-4-receptor gene (DRD4), is associated with approach-related traits including novelty seeking and extraversion, in some, but not all studies. To our knowledge, no studies have been conducted on the genetics of risk-taking behavior in sports. Using a joint-analysis approach, we measured sensation seeking in two cohorts of experienced male and female skiers and snowboarders (n = 503) using a sports-specific tool developed for this study, the Contextual Sensation Seeking Questionnaire for Skiing and Snowboarding (CSSQ-S), and a more general trait measure, the Zuckerman-Kuhlman Personality Questionnaire impulsive sensation-seeking subscale. We detected, and then replicated a significant association between the DRD4 -521CC genotype and sports-specific sensation seeking as measured using the CSSQ-S (P < 0.001). These data suggest that the DRD4 -521 C/T polymorphism contributes to a "risk-taking phenotype" in skiers and snowboarders, but the variant was not associated with impulsive sensation seeking (P = 0.9).

TRC-1: emergence of a clavulanic acid-resistant TEM beta-lactamase in a clinical strain.

A novel TEM-derived plasmid-encoded beta-lactamase, resistant to inhibition by clavulanic acid, has been identified in a clinical strain of Escherichia coli found in Scotland. The beta-lactamase gene was carried on an 81-kb plasmid that conferred no other resistances. The novel enzyme conferred resistance to the amoxycillin/clavulanic acid combination on the host bacterium. The beta-lactamase has a pI of 5.25 and lies between the PSE-4 and SAR-1 beta-lactamases on an isoelectric focusing gel. This beta-lactamase has a Mr value of 25,000, similar to the TEM-1 enzyme and a comparable substrate profile. Its most significant difference is that it is inhibited by clavulanic acid 100-fold less efficiently than the TEM-1 enzyme. The enzyme was confirmed to be derived from the TEM enzymes by probing the plasmid DNA with an intragenic gene probe for TEM-1. This is the first report of a clinical bacterium carrying a TEM-enzyme that confers resistance to clavulanic acid combinations and we have designated the beta-lactamase as TRC-1.

Back mutations to the TEM-1 beta-lactamase from TRC-1 lead to restored sensitivity to clavulanic acid.

Back mutations from the TRC-1 beta-lactamase to the TEM-1 enzyme were selected in vitro. The revertant beta-lactamase was obtained from Escherichia coli strain J62.2 carrying plasmid pUK901 which encodes the TRC-1 beta-lactamase. The revertant was obtained after repeated subculture of E. coli J62.2 (pUK901) in amoxycillin 512 mg/L for 5 days. The revertant beta-lactamase had the same pI as TEM-1 (5.4) and had restored inhibition by clavulanic acid (ID50 reduced from 4.2 microM to 0.15 microM). The prevalence of these beta-lactamases in the clinical population may be the result of a two-way flux, with mutations in both forward and backward directions.

N-terminal amino-acid sequence and subunit structure of the type IV trimethoprim-resistant plasmid-encoded dihydrofolate reductase.

The type IV plasmid-mediated dihydrofolate reductase (DHFR), from a clinical strain of Escherichia coli isolated in South India, was prepared from a transconjugant containing the original clinical plasmid, E. coli J62-2 (pUK1123), and from E. coli C600 (pUK1150) containing a 2.6-kb HindIII fragment of pUK1123 cloned into plasmid pBR322. Both preparations were purified by methotrexate affinity chromatography. Automatic amino-acid sequencing of the N-terminal of the purified type IV enzyme from both sources gave an identical sequence which was clearly distinct from other plasmid-mediated trimethoprim-resistant DHFRs. The type IV DHFR showed most homology with the endogenous, chromosomally-encoded E. coli enzyme. Amino-acid sequence analysis also showed that the type IV enzyme preparation from E. coli J62-2 harbouring the original clinical plasmid, pUK1123, also contained the E. coli DNA-binding protein NS1. Analysis by polyacrylamide gel electrophoresis suggested that the type IV enzyme, in its native form, consists of a DHFR of Mr 33,000 coupled to a DNA-binding protein.

The role of thymine starvation in the expression of type IV plasmid-encoded trimethoprim-resistant dihydrofolate reductase.

Hyperproduction of the type IV plasmid-encoded dihydrofolate reductase was studied in Escherichia coli J62-2 (pUK1123). Hyperproduction of the enzyme was shown to occur not simply as a response to a given concentration of trimethoprim but also to the presence of thymidine in the medium. Before hyperproduction occurred the bacteria began to elongate and die, thus showing the symptoms of thymine starvation. Hyperproduction also required the presence of L-methionine, adenine and glycine, suggesting that the elevated production of the enzyme was a response to the ability of trimethoprim to starve the cell of thymine metabolites.

The induction of trimethoprim resistance encoded by the type IV dihydrofolate reductase gene.

The effect of plasmid pUK1123, which confers low level resistance to trimethoprim when tested on solid minimal medium, but also no resistance when tested on IsoSensitest agar, was investigated in liquid media. The growth of Escherichia coli J62-2, harbouring pUK1123, was unaffected in liquid minimal medium containing trimethoprim 10 mg/L. However, in IsoSensitest broth, exposure to this drug concentration resulted in bacteriostasis. After an initial delay, resistance to trimethoprim was induced in IsoSensitest broth containing trimethoprim 10 mg/L, by the imposition of thymine starvation. This response was immediately reversible when trimethoprim was removed, confirming that resistance resulted from induction rather than selection of resistant mutants.

Identification and cloning of the type IIIa plasmid-encoded dihydrofolate reductase gene from trimethoprim-resistant gram-negative bacteria isolated in Britain.

A clinical strain of Escherichia coli isolated in Nottinghamshire in 1980 was shown to harbour the type IIIa trimethoprim-resistant dihydrofolate reductase gene, previously identified on only one occasion, in New Zealand in 1979. The gene was identified by hybridisation with an 855-bp type III gene probe and its classification as a type IIIa dihydrofolate reductase was confirmed by detailed biochemical analysis of the enzyme product. The dihydrofolate reductase was identical in size and isoelectric point with the original type IIIa enzyme and shared similar inhibitory and kinetic profiles. The trimethoprim resistance gene was subsequently cloned and the type IIIa dihydrofolate reductase gene was localised to a 700-bp EcoRI-PstI fragment. This smaller fragment may prove to be a more specific DNA probe for the future identification of type IIIa dihydrofolate reductase genes.

A piperacillin-tazobactam resistant Escherichia coli strain isolated from a faecal sample of a healthy volunteer.

As part of a surveillance programme of the prevalence of antibiotic resistance, the faecal bacteria of healthy people (n = 1348) were examined, and the antibiotic resistance of the Escherichia coli strains determined. One strain out of 142 amoxycillin-resistant isolates, E. coli strain 1662, was also resistant to piperacillin-tazobactam but susceptible to amoxycillin-clavulanic acid. The piperacillin-tazobactam resistance determinant was transferable to standard E. coli strains by conjugation. However, the strain produced a beta-lactamase with several characteristics very similar to those of the TEM-1 beta-lactamase, i.e. pI of 5.4, an M(r) value of 22,000 and a comparable substrate profile. The enzyme was as efficiently inhibited by clavulanic acid and tazobactam as the TEM-1 and TEM-2 beta-lactamases but more than the amoxycillin-clavulanic acid-resistant TRC-1 enzyme. The transferable resistance to piperacillin-tazobactam appears to be mediated by a novel resistance mechanism that has previously not been described.

beta-Lactum resistance in aerobic commensal faecal flora.

Faecal specimens from 100 healthy volunteers living in Edinburgh were examined for the presence of antibiotic-resistant bacteria. A high incidence of ampicillin resistance was found as 42% of specimens containing normally sensitive bacteria were resistant to the drug; however, only 12% of the specimens contained trimethoprim-resistant bacteria. There was no detectable resistance to the third-generation cephalosporin, ceftazidime or the 4-quinolone, ciprofloxacin. Identification of the beta-lactamases produced by the ampicillin-resistant isolates demonstrated that the TEM-1 beta-lactamase predominated particularly in E. coli where it was identified in 86% of isolates. Thirty-three percent of the ampicillin-resistant isolates were able to transfer their resistance to E. coli K12 strain J62-2 and analysis of these transconjugants by iso-electric focusing revealed that the TEM-1 beta-lactamase was present in 100% of the transconjugants. Restriction endonuclease fingerprinting of the TEM-1 containing plasmids suggested the presence of an epidemic plasmid in the community.

The incidences of resistance to cephalosporins and fluorinated 4-quinolones in similar strains isolated in Glasgow and Edinburgh.

Bacterial sensitivity to cefuroxime, ceftazidime, cefotaxime, ciprofloxacin and ofloxacin was determined for 1386 urinary and bacteraemia isolates from Glasgow and Edinburgh to detemine the impact of these antibacterials on the development of resistance. The MIC(50) and MIC(90) values were determined for each species or genus. Cefuroxine was the least effective antibacterial drug and cefotaxime was the most potent cephalosporin, but it rarely matched the efficacy of the 4-quinolones. There was little difference in the sensitivities of Gram-negative bacteria from Edinburgh of Glasgow but Gram-positive bacteria isolated in Glasgow were usually more resistant. There has been no significant emergence of resistant Gram-negative bacteria even amongst the Pseudomonas spp.; however, the proportion of Gram-positive bacteria resistant to these drugs is higher in Scotland than elsewhere.

Antibiotic resistance in urinary bacteria isolated in central Scotland.

The levels of antibacterial amongst 991 strains responsible for significant bacteriuria, isolated in central Scotland at the end of 1990, have been determined by breakpoint sensitivity testing. Overall resistance to the commonly used antibacterials for UTI, trimethoprim and ampicillin was 23% and 36%, revealing that resistance to these agents in central Scotland had not significantly changed over the last ten years. High levels of ampicillin resistance have led to the widespread use of amoxicillin in combination with the beta-lactamase inhibitor clavulanic acid. The effectiveness of this approach was demonstrated by the fact that resistance among these urinary isolates to amoxicillin/clavulanic acid was only 6%. More detailed examination of Escherichia coli isolates, which were ampicillin-resistant, revealed that the addition of clavulanic acid restored sensitivity in 97.5% of the strains.

Antibiotic resistance in nosocomial isolates in Scotland.

In this first multi-centre study in Scotland, 1028 consecutive Gram-negative and staphylococci strains were obtained from four major teaching hospitals. E. coli was the most common organism among both intensive care units (ICUs) (39%) and non-ICU strains (46.6%). The prevalence of antibiotic resistance among E. coli was always higher in isolates from ICUs than non-ICUs: ceftazidime (14.1%, 7.2%), ceftriaxone (12.7%, 6.1%), cefotaxime (15.5%, 8.7%), cefuroxime (28.8%, 20.8%), amoxicillin plus clavulanic acid (52.2%, 38.8%) and gentamicin (7.0%, 2.8%). The highest incidences of resistance were identified among Enterobacter/Citrobacter spp. from ICUs; 43.8%, 41.7%, 45.8%, 54.2%, 87.5% and 10.4% of these organisms were resistant to ceftazidime, ceftriaxone, cefotaxime, cefuroxime, amoxicillin plus clavulanic acid and gentamicin, respectively.

Dissemination of the TEM-I beta-lactamase gene into disparate plasmids in faecal strains of Escherichia coli isolated in Vellore, south India.

The genetic basis for ampicillin resistance in commensal strains of Escherichia coli isolated in Vellore, south India has been examined. Of the 58 strains tested, 41% could transfer ampicillin resistance to a standard E. coli host strain. With the exception of one isolate, transferable ampicillin resistance was shown to result from the presence of the TEM-I beta-lactamase which was found on a wide variety of plasmid types.

Molecular Approaches for the Detection and Identification of ß-Lactamases.

ß-lactamases confer resistance to ß-lactam antibiotics, which are the most widely used family of antibiotics. It is, therefore, essential that one can identify the production of ß-lactamases by clinical isolates and have effective ways of distinguishing the different enzymes. This is necessary for epidemiologic surveys, predicting future resistance trends, and to ensure that patients receive the appropriate ß-lactam or alternative therapy.

Quinolone susceptibility of Vibrio cholerae O1 & O139 isolates from Vellore.

BACKGROUND & OBJECTIVES: Vellore is an endemic area for cholera. The relative prevalence of clinical cases of Vibrio cholerae O1 and O139 has been fluctuating. Few studies have examined the susceptibility of local isolates to quinolones. The objective of the present study was to look at quinolone susceptibility and determine MIC of ciprofloxacin to representative clinical isolates of V. cholerae O1 and O139 in Vellore, obtained between 1997 and 1999. METHODS: Antimicrobial susceptibility testing of V. cholerae strains was performed by disc diffusion technique and MIC determination by E test. RESULTS: Five of 30 O1 and all the O139 serogroup isolates were susceptible to nalidixic acid. All isolates of both serogroups were sensitive to norfloxacin. All isolates of both serogroups gave MIC results in the susceptible range to ciprofloxacin; the MICs being lower for V. cholerae O139 (MIC50 = 0.004 microgram/ml and MIC90 = 0.047 microgram/ml) than for O1 serogroup (MIC50 = 0.38 microgram/ml and MIC90 = 0.5 microgram/ml). INTERPRETATION & CONCLUSION: V. cholerae O1 and O139 show differences in quinolone susceptibility, the reason for this is not clear. This could be because of longer exposure of the O1 serogroup to quinolone antimicrobials as compared to the O139 serogroup.

Trimethoprim and brodimoprim resistance of gram-positive and gram-negative bacteria.

Trimethoprim and brodimoprim act by selectively inhibiting bacterial dihydrofolate reductase. There are a number of mechanisms by which bacteria can develop resistance to these agents. These include thymineless mutation, impermeability, alteration in chromosomal dihydrofolate reductase and the plasmid-encoded production of an additional dihydrofolate reductase which is insensitive to inhibition by antifolate agents. Clinically the most important of these is the plasmid-encoded production of additional dihydrofolate reductases and such resistance is found in both gram-positive and gram-negative species. These plasmid-encoded enzymes were initially divided into a number of classes based principally on their biochemical profiles. More recently sequence analysis has been used to study these proteins and thus the classification of dihydrofolate reductases now also takes into account sequence information. The number of plasmid-mediated dihydrofolate reductases has increased markedly in recent years. Whilst this probably results from the continuing evolution of resistance it can also be partly attributed to more discriminatory methods for studying these enzymes.

High prevalence of extended spectrum beta-lactamase production among Klebsiella pneumoniae strains isolated at a University Hospital in Turkey.

Beta-lactam susceptibility and beta-lactamase patterns of a random sample of 44 Klebsiella pneumoniae strains that had been isolated from nosocomial infections at Dokuz Eylül University Hospital in Izmir, were investigated. All strains were amoxycillin resistant but in the presence of clavulanic acid 26 became sensitive. Similarly 39 of the strains were resistant to ceftazidime and cefotaxime; clavulanic acid restored sensitivity to ceftazidime in 28 and to cefotaxime in 25 of these resistant strains. Extended spectrum beta-lactamase (ESBL) production was positive in 84% of the isolates as determined by the double disk synergy test. Isoelectric focusing revealed that each strain produced one to four beta-lactamases, pI 7.6 enzymes being the most prevalent. Other enzymes with pIs of 8.4, 8.2, 5.4, 7.8 were also detected. Resistance to ceftazidime was transferred from 18 of the 44 isolates to the recipient Escherichia coli K-12 at 37 degrees C. The transconjugants were examined for their plasmid content and the plasmids were characterized by their size and resistance profile. Fourteen different restriction pattern groups were identified with Eco R1. The results indicate a high prevalence of ESBL production in nosocomial K. pneumoniae isolates in Izmir and have major implications concerning the clinical use of later generation cephalosporins.

Moxifloxacin sensitivity of respiratory pathogens in the United Kingdom.

The in vitro activity of moxifloxacin and comparator agents against respiratory isolates from a range of geographically distinct centres around the United Kingdom was investigated in the following study. Clinical isolates of Streptococcus pneumoniae (n = 257), Haemophilus influenzae (n = 399) and Moraxella catarrhalis (n = 253) were obtained between March 1998 and April 1999 from nine centres in the United Kingdom. Sensitivity was determined by testing each isolate for its minimum inhibitory concentration (MIC) by agar dilution. Against Streptococcus pneumoniae moxifloxacin and grepafloxacin were the most active (MIC90 = 0.25 mg/l). Trovafloxacin and sparfloxacin were the next most active (MIC90 = 0.5 mg/l) followed by levofloxacin and ciprofloxacin. MIC90 values of the six fluoroquinolones versus H. influenzae ranged from <0.0039 mg/l to 0.0625 mg/l and from <0.0039 mg/l to 0.5 mg/l for M. catarrhalis. The rank order of activity of the fluoroquinolones versus H. influenzae was moxifloxacin = trovafloxacin = grepafloxacin = sparfloxacin > ciprofloxacin > levofloxacin. Against M. catarrhalis the lowest MIC90 was that of grepafloxacin at 0.0625 mg/l followed by moxifloxacin, sparfloxacin, levofloxacin and ciprofloxacin. Trovafloxacin demonstrated the highest MIC90 at 0.5 mg/l. These results demonstrate that moxifloxacin has superior in vitro activity against respiratory tract pathogens than any other comparator quinolones available for clinical use.