Antibiotic resistance

Microb Drug Resist, 2004 Winter, 10(4), 280 - 5
Isoniazid resistance and the future of drug-resistant tuberculosis; Cohen T et al.; Bacterial chromosomal mutations that confer antibiotic resistance often have deleterious effects that impose costs on reproductive fitness . This observation has led to the generalization that in the absence of the selection pressure exerted through treatment, the frequency of resistance will decrease . This model implies that the prudent use of antibiotics will eventually result in a decline in the prevalence of drug resistance . Recent work, however, suggests that some resistance-conferring mutations may not significantly impair fitness and that others may be accompanied by compensatory mutations that restore the organisms' reproductive potential . Thus drug resistance, once introduced, may persist unless specific measures are implemented to target prevalent drug-resistant cases . Here we present ecological evidence to support the hypothesis that mutations at the 315 position of katG confer isoniazid resistance for Mycobacterium tuberculosis without diminishing virulence or transmissibility.

New Microbiol, 2004 Oct, 27(4), 353 - 60
Morphological changes and outer membrane protein patterns in Helicobacter pylori during conversion from bacillary to coccoid form; Citterio B et al.; Conversion from bacillary to fully coccoid form via an intermediate U-and V-shaped form has been described in prolonged cultures of H . pylori . This morphological transformation may be the expression of transitory adaptation to a particular environment and may play an important role in antibiotic resistance and the difficulty to eradicate the pathogen . The aim of this study was to evaluate morphological and outer membrane protein changes in H . pylori during ageing-induced conversion to coccoid morphology . We used two H . pylori strains (the reference NCTC 11639 and a fresh clinical isolate) cultivated in microaerophilic environment at 37 degrees C, monitoring their morphological and biochemical evolutions for 11 days . Microscopic examination revealed the passage from spiral to U- and V-shaped form after 5-8 days of incubation, the conversion to coccoid form and the entry into viable but non-culturable state (VBNC) between days 9 and 11 . Protein pattern difference appeared at 97.4 to 45 and 30 kDa molecular weight . Biochemical tests demonstrated not only a modification of outer membrane protein profiles, but also an intra-specific variability by comparison between the two analysed strains . Our findings suggest that structural and outer membrane changes associated with coccoid transformation represent a typical response in H . pylori and may constitute a survival strategy in adverse environmental conditions.

World J Gastroenterol, 2005 Jan 28, 11(4), 584 - 6
Breath and string test: A diagnostic package for the identification of treatment failure and antibiotic resistance of Helicobacter pylori without the necessity of upper gastrointestinal endoscopy; Leodolter A et al.; AIM: Helicobacter pylori (H pylori) resistance after failed eradication has a major impact on the outcome of a further treatment regimen . The aim of this study was to assess the validity of a non-invasive strategy using the (13)C-urea breath test (UBT) and the gastric string test in identifying post-treatment resistance of H pylori . METHODS: The UBT was routinely performed 4 to 6 wk after H pylori eradication therapy . Forty-two patients (24 females, 18 males, mean age 48 years) with a positive UBT were included in the study . A gastric string test using a capsule containing a 90 cm-long nylon fiber was performed . Before the capsule was swallowed, the free end of the string was taped to the cheek . After one hour in the stomach, the string was withdrawn . The distal 20 cm of the string was inoculated onto an agar plate and processed under micro-aerophilic conditions . Following the string test, upper gastrointestinal endoscopy was performed to obtain gastric biopsies for conventional culture . RESULTS: H pylori was successfully cultured from the gastric string in 34 patients (81%), but not in 5 patients due to contamination with oropharyngeal flora . H pylori was cultured from the gastric biopsies obtained at endoscopy in 39 patients (93%) . CONCLUSION: The UBT followed by the gastric string test in the case of treatment failure is a valid diagnostic strategy with the aim of determining the post-therapeutic antibiotic resistance of H pylori with little inconvenience to the patient . Upper GI-endoscopy can be avoided in several cases by applying consequently this diagnostic package.

APMIS, 2004 Dec, 112(11-12), 728 - 52
Molecular genetic methods for diagnosis and antibiotic resistance detection of mycobacteria from clinical specimens; Shamputa IC et al.; Shamputa IC, Rigouts L, Portaels F . Molecular genetic methods for diagnosis and antibiotic resistance detection of mycobacteria from clinical specimens . APMIS 2004;112:728-52.Mycobacteria comprise a diverse group of bacteria that are widespread in nature, some of which cause significant disease in humans . Members of the Mycobacterium tuberculosis complex (MTBC) are the most important human pathogens of the genus Mycobacterium . Traditional methods for detection and identification of mycobacteria include microscopy, culture and phenotypic tests . These methods either lack sensitivity, specificity, or are time consuming . Advances in the field of molecular biology have provided rapid diagnostic tools that have reduced the turnaround times for detecting MTBC and drug resistance in cultures and directly in clinical specimens from weeks to days . This review discusses the molecular genetic techniques for detecting and identifying MTBC as well as drug resistance of mycobacteria in clinical specimens.

Klin Oczna, 2004, 106(3 Suppl), 521 - 4
{The use of antibiotics in patients undergoing ocular surgery and prevalence of infectious complications, according to data from some ocular surgery centers in Poland}; Szaflik J et al.; The aim of the study was to evaluate effectiveness of prophylactic antibiotics usage and the prevalence of infectious complications, after ocular surgery . Authors analyzed data from 53 ocular surgery centers in Poland: 28674 cases of cataract surgery, 6518 cases of complicated cataract surgery, 1387 cases of contemporary cataract and glaucoma surgery and 2978 cases of glaucoma surgery . The prevalence of endophthalmitis in this group of patients ranged from 0.29% after cataract surgery to 0.93% after complex surgery (cataract and glaucoma), and was higher than other studies show . This is because the study group was much smaller . The study is the first step to create optimal schema of antibiotics usage during ocular surgery . Rational antibiotics usage is very important because of worldwide growing antibiotic resistance.

Virology, 2005 Jan 20, 331(2), 247 - 59
Construction and applications of yellow fever virus replicons; Jones CT et al.; Subgenomic replicons of yellow fever virus (YFV) were constructed to allow expression of heterologous reporter genes in a replication-dependent manner . Expression of the antibiotic resistance gene neomycin phosphotransferase II (Neo) from one of these YFV replicons allowed selection of a stable population of cells (BHK-REP cells) in which the YFV replicon persistently replicated . BHK-REP cells were successfully used to trans-complement replication-defective YFV replicons harboring large internal deletions within either the NS1 or NS3 proteins . Although replicons with large deletions in either NS1 or NS3 were trans-complemented in BHK-REP, replicons that contained deletions of NS3 were trans-complemented at lower levels . In addition, replicons that retained the N-terminal protease domain of NS3 in cis were trans-complemented with higher efficiency than replicons in which both the protease and helicase domains of NS3 were deleted . To study packaging of YFV replicons, Sindbis replicons were constructed that expressed the YFV structural proteins in trans . Using these Sindbis replicons, both replication-competent and trans-complemented, replication-defective YFV replicons could be packaged into pseudo-infectious particles (PIPs) . Although these results eliminate a potential role of either NS1 or full-length NS3 in cis for packaging and assembly of the flavivirus virion, they do not preclude the possibility that these proteins may act in trans during these processes.

J Microbiol Methods, 2005 Feb, 60(2), 281 - 3
Promoter-probe cassettes with the gusA (beta-glucuronidase) reporter gene and several different antibiotic resistance markers; He X et al.; A series of gene cassettes with promoterless gusA reporter genes encoding beta-glucuronidase (GUS), linked to different antibiotic resistance markers, have been developed . The gusA-antibiotic resistance gene cassettes can be used to generate transcriptional fusions useful for expression studies.

Pharmacotherapy, 2004 Dec, 24(12 Pt 2), 216S - 23S
Treating serious infections: focus on cefepime; Wong-Beringer A; Antibiotic resistance among medically important pathogens is curtailing the therapeutic value of many of the standard broad-spectrum antibiotics used for empiric treatment of serious infections . This article presents a brief overview of the etiology of serious infections and the threat of emerging resistance, and reviews the adverse impact of inadequate empiric treatment on patient outcomes and hospital ecology . The treatment of two clinically important infections with cefepime--pneumonia in hospitalized patients and febrile neutropenia--also is discussed.

Appl Environ Microbiol, 2004 Dec, 70(12), 7018 - 23
Comparison of two alternative dominant selectable markers for wine yeast transformation; Cebollero E et al.; Genetic improvement of industrial yeast strains is restricted by the availability of selectable transformation markers . Antibiotic resistance markers have to be avoided for public health reasons, while auxotrophy markers are generally not useful for wine yeast strain transformation because most industrial Saccharomyces cerevisiae strains are prototrophic . For this work, we performed a comparative study of the usefulness of two alternative dominant selectable markers in both episomic and centromeric plasmids . Even though the selection for sulfite resistance conferred by FZF1-4 resulted in a larger number of transformants for a laboratory strain, the p-fluoro-DL-phenylalanine resistance conferred by ARO4-OFP resulted in a more suitable selection marker for all industrial strains tested . Both episomic and centromeric constructions carrying this marker resulted in transformation frequencies close to or above 10(3) transformants per microg of DNA for the three wine yeast strains tested.

Acta Microbiol Immunol Hung, 2004, 51(3), 333 - 49
Infectious plasmid resistance and efflux pump mediated resistance; Molnar J et al.; Various bacterial plasmids can be eliminated from bacterial species cultured as pure or mixed bacterial cultures by non-mutagenic heterocyclic compounds at subinhibitory concentrations . For plasmid curing, the replication should be inhibited at three different levels simultaneously: the intracellular replication of plasmid DNA, partition and intercellular transconjugal transfer . The antiplasmid action of the compounds depends on the chemical structure . The targets for antiplasmid compounds were analysed in detail . It was found that amplified extrachromosomal DNA in the superhelical state binds more drug molecules than does the linear or open-circular form of the plasmid or the chromosome, without stereospecificity which leads to functional inactivation of the extrachromosomal genetic code . Plasmid elimination also occurs in ecosystems containing numerous bacterial species simultaneously, but the elimination of antibiotic resistance-encoding plasmids from all individual cells of the population is never complete . The medical significance of plasmid elimination in vitro is, it provides a method to isolate plasmid-free bacteria for biotechnology without any risk of mutations, and it opens up a new perspective in rational drug design against bacterial plasmids . Hypothetically, the combination of antiplasmid drugs and antibiotics may improve the effectivity of antibiotics against resistant bacteria; therefore, the results cannot be exploited until the curing efficiency reaches 100% . Inhibition of the conjugational transfer of antibiotic resistance plasmids can be exploited to reduce the spreading of these plasmids in ecosystems.

Nat Genet, 2005 Jan, 37(1), 73 - 6 Epub 2004 Nov 28.
The 'evolvability' of promiscuous protein functions; Aharoni A et al.; How proteins with new functions (e.g., drug or antibiotic resistance or degradation of man-made chemicals) evolve in a matter of months or years is still unclear . This ability is dependent on the induction of new phenotypic traits by a small number of mutations (plasticity) . But mutations often have deleterious effects on functions that are essential for survival . How are these seemingly conflicting demands met at the single-protein level? Results from directed laboratory evolution experiments indicate that the evolution of a new function is driven by mutations that have little effect on the native function but large effects on the promiscuous functions that serve as starting point . Thus, an evolving protein can initially acquire increased fitness for a new function without losing its original function . Gene duplication and the divergence of a completely new protein may then follow.

Antimicrob Agents Chemother, 2004 Dec, 48(12), 4843 - 7
Distribution of antibiotic MICs for Helicobacter pylori strains over a 16-year period in patients from Seoul, South Korea; Kim JM et al.; Recently, the development of antibiotic resistance emerged as a significant clinical problem in the eradication of Helicobacter pylori . We investigated the MICs of antibiotics for 135 H . pylori isolates from adults in Seoul, South Korea, over the past 16 years . The MICs of amoxicillin, clarithromycin, metronidazole, tetracycline, azithromycin, and ciprofloxacin increased from 1987 to 2003 . Rates of primary resistance to clarithromycin increased from 2.8% in 1994 to 13.8% in 2003 . The A2144G mutation was frequently observed in the 23S rRNA gene in clarithromycin-resistant isolates . The increase in resistance to clarithromycin seems to result in a decrease in eradication efficacy for H . pylori . These results suggest that the MICs of several antibiotics for H . pylori have increased over the past 16 years in Seoul.

Anal Biochem, 2004 Dec 15, 335(2), 305 - 15
Selecting transpositions using phage P1 headful packaging: new markerless transposons for functionally mapping long-range regulatory sequences in bacterial artificial chromosomes and P1-derived artificial chromosomes; Chatterjee PK et al.; New Tn10 minitransposons were constructed to functionally map long-range transcription regulatory sequences in bacterial artificial chromosomes (BACs) and P1-derived artificial chromosomes (PACs) . Each contained a wild-type loxP site but, significantly, contained no mammalian or bacterial genes and/or promoter elements within the transposed portion of DNA . In contrast to loxP transposons described previously, the new ones do not introduce transcription regulatory elements capable of interfering with those endogenous to the BAC clone in functional mapping studies . Progressive deletions from the loxP end of genomic DNA were efficiently generated using these transposons, and a series of truncations generated in a green fluorescence protein (GFP)-BAC fusion clone unambiguously identified three new long-range enhancer sequences functionally in the Nkx2-5 gene in transgenic mice . Insertions of these new transposons lacking antibiotic resistance genes into a BAC or PAC were indirectly selected by their ability to delete enough DNA from the clone so as to enable its packaging within a P1 phage head with both loxP sites intact for subsequent recovery of the large plasmid . The outcome of such an indirect mode of selection is both desirable and undesirable . First, because the screen is not antibiotic resistance marker dependent, the same transposon can be used to generate nested deletions efficiently in both BACs and PACs . Second, deletions through intrainsert recombinations unrelated to loxP/Cre also get packaged and recovered, and size analyses of the BAC/PAC vector band after NotI digestion is indispensable to identify authentic loxP/Cre deletions . The procedure nevertheless offers a potential approach to map recombinogenic sequences in BACs and PACs.

Ann Ig, 2004 Jan-Apr, 16(1-2), 157 - 62
Non compliance to therapeutic prescriptions in paediatric patients: role of social communication; Agodi A et al.; The aim of this pilot study was to identify prevalent non-compliance behaviour in paediatric antibiotic therapy and to investigate the possible role of its social correlates . Patients' parents were surveyed at two paediatric practices in Catania, Italy, using an interviewer-administered questionnaire . The two practices were chosen for their location, in two different urban areas, to represent different sets of parents in terms of social status . Anticipated suspension of prescribed antibiotic therapy was the most frequently encountered form of non-compliance, shown by 41.2% of the parents . After partitioning the sample by mothers' occupational status--housewives versus working women--anticipated suspension revealed a positive association with educational level among the former group and a negative one, although not statistically significant, among the latter . Exposition to mass media messages about bacterial antibiotic resistance appeared to be a key intervening variable in interpreting these results, especially among more educated parents . Non-compliance was also associated with perceived characteristics of doctor-parents communication, particularly, with parents' perceived understanding of prescribed therapy.

Clin Gastroenterol Hepatol, 2004 Nov, 2(11), 997 - 1002
A 10-day levofloxacin-based therapy in patients with resistant Helicobacter pylori infection: a controlled trial; Bilardi C et al.; BACKGROUND & AIMS: Antibiotic resistance is a major issue in anti- Helicobacter pylori treatment . This study was aimed at assessing the efficacy of 2 therapies in patients with resistant H pylori infection . METHODS: Patients who had failed 1 or more eradication regimens underwent upper gastrointestinal endoscopy and 2 antral and 2 corpus biopsy specimens were taken for histology and culture . Metronidazole, clarithromycin, and amoxicillin resistance were determined by E-test . Patients were randomly assigned to 2 therapies: 1 group received pantoprazole 40 mg, amoxicillin 1 g, levofloxacin 250 mg, all twice daily for 10 days, and the other group was treated with omeprazole 20 mg twice daily for the first week and omeprazole 20 mg twice daily, tetracycline 250 mg 4 times daily, metronidazole 500 mg twice daily, and bismuth subcitrate 240 mg twice daily for the second week . Therapeutic success was evaluated by 13C urea breath test after 4 weeks of treatment . RESULTS: We enrolled 44 patients in the levofloxacin-based regimen and 46 patients in the quadruple therapy . The former was successful in 31 of 44 (70%; 95% confidence interval: 53-87) and the latter in 17 of 46 (37%; 95% confidence interval: 23-47) patients, using intention-to-treat (ITT) analysis (P < .001) . The rates of H pylori resistance to metronidazole, clarithromycin, and amoxicillin were 46%, 12%, and 0%, respectively . Resistance to both metronidazole and clarithromycin was found in 10% of cases . CONCLUSIONS: Triple therapy containing levofloxacin was better than quadruple therapy . The 70% success rate observed indicates that 10 days of pantoprazole, amoxicillin, and levofloxacin should be considered in patients who had failed 1 or more eradication regimens.

Nat Rev Microbiol, 2004 Dec, 2(12), 989 - 94
Antibiotic resistance: a view from the prescriber; Finch RG; Antibiotic resistance is increasingly affecting the management of infectious diseases . The prescribing clinician is not only important to the development of the problem but also central to its solution . This article addresses the current weaknesses in the information systems and the evidence base that support prescribing . Remedies necessary for improvements in prudent prescribing include better guidance in managing specific diseases where resistance is of prognostic significance and also increasing diagnostic precision.

Int J Syst Evol Microbiol, 2004 Nov, 54(Pt 6), 2269 - 73
Methylobacterium nodulans sp . nov., for a group of aerobic, facultatively methylotrophic, legume root-nodule-forming and nitrogen-fixing bacteria; Jourand P et al.; Data on 72 non-pigmented bacterial strains that specifically induce nitrogen-fixing root nodules on the legume species Crotalaria glaucoides, Crotalaria perrottetii and Crotalaria podocarpa are reviewed . By SDS-PAGE analysis of total protein patterns and by 16S rRNA PCR-RFLP, these strains form a homogeneous group that is separate from other legume root-nodule-forming bacteria . The 16S rRNA gene-based phylogeny indicates that these bacteria belong to the genus Methylobacterium . They can grow on C(1) compounds such as methanol, formate and formaldehyde but not methylamine as sole carbon source, and carry an mxaF gene, encoding methanol dehydrogenase, which supports their methylotrophic metabolism . Presence of a nodA nodulation gene, and ability to nodulate plants of Crotalaria species and to fix nitrogen are features that separate the strains currently included in this group from other members of the genus Methylobacterium . The present study includes additional genotypic and phenotypic characterization of this novel Methylobacterium species, i.e . nifH gene sequence, morphology, physiology, enzymic and carbon source assimilation tests and antibiotic resistance . The name Methylobacterium nodulans sp . nov . (type strain, ORS 2060(T)=CNCM I 2342(T)=LMG 21967(T)) is proposed for this group of root-nodule-forming bacteria.

Mol Biotechnol, 2004 Nov, 28(3), 201 - 4
An efficient bipartite PCR technique to introduce specific changes in large plasmids; Davis K et al.; Amplifying an entire double-stranded plasmid by an inverse polymerase chain reaction (PCR) using a pair of tail-to-tail primers is a particularly efficient approach for introducing changes into DNA sequences . However, the approach generally works best for plasmids less than 5 Kb and it can be difficult to amplify the large multicomponent vectors that are used for protein expression in various eukaryotic cells . We have therefore adopted an alternative approach in which two smaller PCR products are generated and then ligated to produce the complete plasmid . A mutagenic primer is used to introduce the desired change and each reaction includes one of a pair of tail-to-tail primers from within an antibiotic resistance gene contained on the plasmid so that the two PCR products contain complementing parts of the complete gene . Ligating the two products generates various combinations but only the correctly ligated molecules recreate the antibiotic resistance gene and are able to replicate in Escherichia coli . When combined with methods to minimize the carryover of template plasmid, this can be an efficient way of introducing mutations into large plasmids.

Intensive Care Med, 2004 Dec, 30(12), 2145 - 56 Epub 2004 Nov 05.
Antibiotic pharmacokinetic and pharmacodynamic considerations in critical illness; Mehrotra R et al.; BACKGROUND: Many factors over which there may be little control may influence the response of a patient to therapy . However, therapy with antibiotics can be readily optimised.DISCUSSION: Concentration-dependent agents such as aminoglycosides appear effective and to entail fewer side effects when given in large, infrequent doses . There is also evidence that time-dependent antibiotics often fail to reach adequate concentrations throughout the treatment period . To date no randomised controlled prospective trial has demonstrated improvement in clinical outcome following infusion rather than intermittent boluses of time-dependent antibiotics . Critical illness alters antibiotic pharmacokinetics principally through increases in volume of distribution . Other than glycopeptides and aminoglycosides, antibiotic blood concentrations are rarely monitored and therefore adequate concentrations can only be inferred from clinical response.CONCLUSIONS: Failure to respond within the first few days of empirical treatment may be due to antibiotic resistance or inadequate doses . Therefore the same rigor should be applied to achieving adequate antibiotic concentrations as is applied to inotropes, which are titrated to achieve predetermined physiological targets.

Prev Med, 2005 Feb, 40(2), 162 - 9
Appropriate antibiotic use: variation in knowledge and awareness by Hispanic ethnicity and language; Corbett KK et al.; BACKGROUND: Recent campaigns are informing the public that antibiotics are inappropriate for viral respiratory infections . As little is known about their effect on populations challenged by less access to care, lower education, low income, low English proficiency, or non-mainstream cultural backgrounds, this study assessed knowledge, attitudes, and awareness in an ethnically diverse community . METHODS: A telephone survey in English or Spanish of a cross-sectional, random sample of 692 non-Hispanic whites (NHWs) and 300 Hispanics in Colorado . RESULTS: For all respondent groups, knowledge of appropriate antibiotic use for colds and bronchitis was low . Hispanics surveyed in Spanish, compared with non-Hispanic whites, had significantly lower knowledge about antibiotics for colds, higher knowledge for bronchitis, lower awareness about antibiotic resistance, and greater dissatisfaction if an antibiotic were not prescribed . In all comparisons, English-language Hispanics tended to reflect non-Hispanic white response patterns . Independent predictors of awareness were ethnicity, education, and age . Independent predictors of dissatisfaction were ethnicity, knowledge about antibiotic use for colds, and bronchitis . Ethnicity was an independent predictor of knowledge about the inappropriateness of antibiotics for colds and bronchitis . CONCLUSIONS: To bridge knowledge gaps, educational campaigns for all segments of the population are needed . Content should be responsive to heterogeneity within populations.

J Antimicrob Chemother, 2005 Jan, 55(1), 6 - 9 Epub 2004 Nov 05.
Antibiotic cycling or rotation: a systematic review of the evidence of efficacy; Brown EM et al.; Of the interventions designed to reduce antibiotic resistance rates in hospitals, one that is currently attracting considerable interest, particularly in the intensive care unit setting, is antibiotic cycling or rotation . Cycling is the scheduled rotation of one class of antibiotics with one or more different classes exhibiting comparable spectra of activity; in order to fulfil the definition, the cycle must be repeated . Following a search of the literature we identified 11 articles in which the authors claimed to have evaluated the efficacy of this intervention . Only four were suitable for review, but, owing to multiple methodological flaws and a lack of standardization, the results of these studies do not permit reliable conclusions regarding the efficacy of cycling . Further studies are therefore required in order to resolve this question . However, before such studies can be undertaken, there are a great many issues relating to cycling which must be addressed . For the time being, we advise against the routine implementation of this measure as a means of reducing antibiotic resistance rates.

Cell Mol Life Sci, 2004 Oct, 61(19-20), 2632 - 45
Single-copy T-DNAs integrated at different positions in the Arabidopsis genome display uniform and comparable beta-glucuronidase accumulation levels; De Buck S et al.; This study aimed at determining whether transgene expression variability is observed in single-copy T-DNA plants and whether it can be correlated with the T-DNA integration position . Among a population of 135 Arabidopsis thaliana transformants, selected on the basis of antibiotic resistance marker expression, 21 single-copy T-DNA transformants were identified and characterized . In 19 of these 21 lines, 35S-beta-glucuronidase transgene expression, measured in two subsequent generations, was similar . This observation means that the intra-transformant variability was as high as the inter-transformant variability . Integration into an intergenic or genic region, into an exon or intron, in sense or antisense orientation, did not result in differential transgene expression . Remarkably, single-copy transformants were not always the highest expressers, implying that low transgene expression is not always induced by multicopy transformants . In only 2 of the 21 single-copy plants was the transgene expression more than 20-fold lower . However, characteristics of the insertion position in one of these lines did not differ significantly when compared to high-expressing lines . In the remaining line, methylation of the transgene was clearly demonstrated . In conclusion, screening for single-copy T-DNA transformants greatly enriches for stable and high transgene expression, because the integration position is not a major determinant of transgene expression variability in Arabidopsis.

Med Sci (Paris), 2004 Nov, 20(11), 999 - 1003
{Immunoprophylaxis of respiratory infections}; Dreffier C et al.; Anti-infective antibody-based immunotherapy has gained renewed interest since the crisis of antibiotic resistance and because there is no therapy against various viral infections . The immunoprophylaxis of respiratory infections aims to utilize the ability of local antibodies to neutralize inhaled micro-organisms and their cytopathic products . Immunoglobulins for intravenous use (i.v.i.g.) have a wide spectrum of specificities . Hyperimmune i.v.i.g . containing high titers of specific antibodies have demonstrated efficacy in clinical trials, notably against the respiratory syncytial virus . Monoclonal antibodies have the advantage to be homogenous and specific for one selected epitope and several studies have demonstrated their efficacy to neutralize several infectious agents . Moreover, antibodies can be administered topically and are effective at lower doses than those needed for systemic administration . The mechanism of action could be the agglutination of bacteria or viruses at the epithelial surfaces of the respiratory tract inhibiting the early steps of the infectious process . Thanks to new technologies of humanized monoclonal antibodies, immunotherapy offers real promising perspectives for prophylactic and therapeutic therapies against a variety of current or emerging infectious diseases.

Ned Tijdschr Geneeskd, 2004 Sep 4, 148(36), 1765 - 9
{The increase in pneumonia-related morbidity and mortality among adults in the Netherlands and possible explanations for it}; Oosterheert JJ et al.; As seen in several national registration systems, the morbidity and mortality due to pneumonia-related disease has increased in the Netherlands in the past 10 years . It is unlikely that the observed increase in mortality due to pneumonia is purely the result of the registration system used or misclassification . An increase in the elderly population is the most likely, albeit only partial, explanation . An increase in underlying diseases such as chronic obstructive pulmonary disease and diabetes mellitus is also a possible explanation . Increased antibiotic resistance, inadequate treatment or a shift in the spectrum of micro-organisms causing pneumonia are less likely explanations . Reducing the morbidity and mortality due to pneumonia demands an integrated approach aimed at elderly patients or patients with co-existing disease and executed by specialists in the fields of geriatrics, internal medicine, pulmonary disease and infectious diseases.

Turk J Gastroenterol, 2004 Sep, 15(3), 159 - 163
The effect of Helicobacter pylori eradication in patients with functional dyspepsia: Assessment of different diagnostic tests; Cinar K et al.; BACKGROUND/AIMS: Helicobacter pylori infection, is seen in more than 80% of adult population in Turkey . The aims of this study were 1) to evaluate the importance of the diagnostic tests 2) to investigate the relationship between Hp infection and functional dyspepsia . METHODS: A total 75 patients with functional-dyspepsia were involved into the study . Hp infection was diagnosed by histopathological examination . CLO, cytology, culture, stool antigen and breath test . Symptom score using ROME II criteria was also evaluated to all patients . All patients were taken ranitidine-bismuth-citrate (400mg bid/day), clarithromycin (500mg bid/day) and amoxicillin (1000mg bid/day) for 14 days . All tests and symptom score analyses were re-applied at month 1and 6 . RESULTS: The eradication rate was 95.9% . The baseline specifity of breath test, CLO, cytology, culture and stool antigen were 87.3%, 95.4%, 95.4% 94.5% and 86.4%, respectively . The sensitivity of such tests at first month after stopping the treatment were 86.1%, 100%, 100%, 100%, 84.7%, respectively, and were 91.6%, 100%, 100%, 100%, 87.5%, respectively at six months after treatment . Symptom scores were 29.6+/-5.4, 15.8+/-4.7 and 17.9+/-5.3 at baseline, first month and six months after treatment, respectively (p<0.001) . CONCLUSIONS: The success of eradication may be related to use of bismuth which prevents antibiotic resistance development . Stool antigen and breath tests are less effective than invasive diagnostic-tests . The finding of improved symptomscores after eradication suggests that Hp may play a role in functional dyspepsia.

Expert Rev Anti Infect Ther, 2004 Apr, 2(2), 227 - 34
Optimization of antibiotic dosing schedules in the light of increasing antibiotic resistance; Smith SV et al.; The development of antibiotic-resistant bacteria is an increasing problem throughout the world and is, without doubt, due to the increasing use of antibiotics themselves . As organisms become more resistant, treatment options become more limited and treatment failures increasingly likely . The need to reverse, or at least minimize this pattern of increasing resistance is therefore essential . Numerous strategies to achieve this have been postulated and there is no doubt that a combination of these will ultimately prove to be most effective . Among them, using antibiotic dosing regimens that may be less likely to promote resistance is one measure which could be beneficial, and this will be the focus of this review . Individual antibiotics will be discussed under the headings of the patterns of killing activity that they produce . Namely whether they exhibit concentration-dependent killing with prolonged persistent effects, time-dependent killing with minimal or no persistent effects, or time-dependent killing with prolonged persistent effects . The available evidence for optimal dosing regimens, as far as minimizing antibiotic resistance is concerned, will be reviewed . Where possible, recommendations relating to clinical practice will be made, or failing that, an indication of where further research would be of benefit.

Expert Rev Anti Infect Ther, 2003 Aug, 1(2), 253 - 60
Antibiotic resistance in the intensive care unit setting; Johnson AP; Antibiotic resistance is an important factor influencing clinical outcome for patients in intensive care units . It is also associated with increased healthcare costs resulting from prolonged patient stays . The problem of antibiotic resistance is particularly acute in intensive care units because they house seriously ill patients who are predisposed to infection, as a result of which, antibiotic use is extremely common . Strategies for controlling resistance in intensive care units have focused on attempting to reduce unnecessary antibiotic use, while at the same time ensuring adequate antibiotic cover is provided . The formulation of policies for the effective use of antibiotics in individual intensive care units requires a multidisciplinary approach, entailing regular epidemiological surveillance, together with input from critical care specialists, infectious disease specialists and pharmacists.

Yi Chuan Xue Bao, 2004 May, 31(5), 525 - 32
{Site-directed mutagenesis and promoter functional analysis of RM07 DNA fragment from Halobacterium halobium in Escherichia coli}; Yang Y et al.; The RM07 DNA fragment from Halobacterium halobium was inserted upstream of chloramphenicol acetyltransferase (cat) reporter gene in pKK232-8 in two different orientations (positively or negatively), generating the RM07-cat fusion plasmids pRM07-1 (+) and pRM07-1 (-) . These two plasmids and pKK232-8 were transformed into Escherichia coli HB101 respectively, then the antibiotic resistance level and the chloramphenicol acetyltransferase protein concentration of different transformants were detected . The research results revealed that HB101/pRM07-1 (+) was resistant to chloramphenicol and could grow on the plate containing chloramphenicol, but HB101/pRM07-1 (-) and HB101/pKK232-8 were sensitive to it . Therefore, the results suggested that the positive RM07 fragment had promoter activity in Escherichia coli while the negative RM07 fragment did not . Site-directed mutagenesis of RM07 was performed by PCR mutagenesis method . The effect of specific nucleotide mutations on the chloramphenicol resistance level of different transformants was detected . The different transformants containing different nucleotide mutations were inoculated on the plates containing various concentrations of chloramphenicol and then incubated at the same time . The change of chloramphenicol resistance level reflected the change of promoter activity . By using this method, the important nucleotide responsible for the promoter function of RM07 in Escherichia coli was determined . The promoter activity of RM07 in Escherichia coli was also improved greatly by modifying the nucleotide component.

J Perinat Neonatal Nurs, 2004 Jul-Sep, 18(3), 241 - 58
Strategic use of antibiotics in the neonatal intensive care unit; Tom-Revzon C; Infection is one of the most common causes of infant morbidity and mortality in the neonatal intensive care unit, despite the availability of various therapeutic medical interventions, such as mechanical ventilation, that allow premature infants a better chance of survival . In fact, many of these therapeutic interventions violate the natural protective barriers for infection . Fortunately, maternal and neonatal risk factors have been identified for early- and late-onset sepsis and the prevalence of highly suspected pathogens for each type of infection influences the empiric selection of antibiotics . In many institutions, the common pathogens have developed resistance to antibiotics . In others, the predominance of pathogens has shifted . Both challenges may warrant modification of traditional antibiotic regimens . Continual monitoring of pathogen shifts within an individual neonatal intensive care unit and application of various antibiotic characteristics are key elements of strategic, safe, and effective use of antibiotics in this patient population . Development of a protocol incorporating infection control measures and guidelines for judicious use of antibiotics can minimize the threat of antibiotic resistance.

Rev Med Chil, 2004 Aug, 132(8), 1001 - 6
{Antibiotics and aquaculture in Chile: implications for human and animal health}; Cabello FC; Industrial antibiotic usage in agribusinesses and aquaculture is the force that drives the evolution of antibiotic resistant bacteria that produce human and animal disease in many countries . Several studies have demonstrated that most of the industrial use of antibiotics is unnecessary, and that modernization and hygienic changes can reduce this use of antibiotics without negative economic impact . In Chile, industrial aquaculture of salmon has expanded rapidly in the last 20 years becoming a major export business . The exponential growth of this industry has been accompanied by an unrestricted heavy usage of antibiotics in the aquatic environments of lakes, rivers and the ocean, and its impact is being felt in the emergence of antibiotic-resistant bacteria around aquaculture sites and a decrease in the plancktonic diversity in the same areas . The passage of antibiotic resistance genes from aquatic bacteria to human and animal pathogens has been demonstrated, indicating that industrial use of antibiotics in aquaculture affects negatively the antibiotic therapy of human and animal bacterial infections . The Chilean situation triggers important concerns because it includes the use of fluoroquinolones in aquaculture, that are not biodegradable and are able to remain in the environment for years as well as being still effective in treating human infections . The use of large volumes of a wide spectrum of antibiotics in an aquatic environment heavily contaminated with human and animal pathogens also amplifies the opportunities for gene transfer among bacteria, facilitating the emergence of antibiotic resistance and more pathogenic bacterial recombinants . The detection of residual antibiotics in salmons marketed for human consumption that can modify the normal flora of the population also suggests the need for controls on this antibiotic usage and on the presence of residual antibiotics in aquaculture food products . This important problem of public health demands an active dialogue between government officials responsible for protecting public health, aquaculture industry representatives, politicians, consumers and professionals dealing with these matters.

N Z Med J . 2004 Aug 20;117(1200):U1022.
Antibiotic resistance in Helicobacter pylori: is it a problem in New Zealand?
Ahmed D, Brooks H, McConnell M, Barbezat G.
AIMS: Infection with Helicobacter pylori requires antibiotic treatment when associated with upper gastrointestinal symptoms . Antibiotic susceptibility tests on this pathogen are rarely carried out and there is little information available on the incidence of antibiotic resistant strains in New Zealand . The aim of this study was to assess the susceptibility of H . pylori (cultured from gastric biopsies in Dunedin) to antibiotics that are commonly used in treatment . METHODS: Over a 13-month period, gastric biopsies were obtained from selected patients undergoing endoscopy at Dunedin Public Hospital because of upper gastrointestinal symptoms . Biopsies were cultured for H . pylori and examined histologically . Fifty isolates from 50 patients with histological findings compatible with H . pylori infection were tested for sensitivity to metronidazole, clarithromycin, amoxycillin, and tetracycline using the epsilometer (E-) test . RESULTS: Metronidazole resistance was detected in 10 (20%) isolates . These isolates were highly resistant to metronidazole and were not inhibited by 250 mg/L . Resistance to clarithromycin, amoxycillin, and doxycycline was not detected . CONCLUSIONS: Clinicians should be aware that resistance to metronidazole appears to be common in H . pylori in New Zealand and that treatment regimens including this antibiotic may be less effective as a result . Although clarithromycin resistance was not detected, it is becoming increasingly problematical overseas and has been recorded in Auckland.

Diagn Microbiol Infect Dis, 2004 Oct, 50(2), 153 - 5
Antibiotic resistance may affect alkali decontamination of specimens containing mycobacteria; Yesilkaya H et al.; The influence of genetic modification on the decontamination process of mycobacteria was investigated using a wild type Mycobacterium bovis BCG Pasteur and its isogenic mutant strain . The result showed that the killing effect of sodium hydroxide (NaOH) progressively increased with prolonged incubation (up to 20 minutes), and the deteriorating effect of NaOH was more severe on the genetically modified strain.

Appl Environ Microbiol, 2004 Oct, 70(10), 5947 - 54
High-level genotypic variation and antibiotic sensitivity among Escherichia coli O157 strains isolated from two Scottish beef cattle farms; Vali L et al.; Escherichia coli O157:H7 is a human pathogen that is carried and transmitted by cattle . Scotland is known to have one of the highest rates of E . coli O157 human infections in the world . Two hundred ninety-three isolates were obtained from naturally infected cattle and the environment on two farms in the Scottish Highlands . The isolates were typed by pulsed-field gel electrophoresis (PFGE) with XbaI restriction endonuclease enzyme, and 19 different variations in patterns were found . There was considerable genomic diversity within the E . coli O157 population on the two farms . The PFGE pattern of one of the observed subtypes matched exactly with that of a strain obtained from a Scottish patient with hemolytic-uremic syndrome . To examine the stability of an individual E . coli O157 strain, continuous subculturing of a strain was performed 110 times . No variation from the original PFGE pattern was observed . We found three indistinguishable subtypes of E . coli O157 on both study farms, suggesting common sources of infection . We also examined the antibiotic resistance of the isolated strains . Phenotypic studies demonstrated resistance of the strains to sulfamethoxazole (100%), chloramphenicol (3.07%), and at a lower rate, other antibiotics, indicating the preservation of antibiotic sensitivity in a rapidly changing population of E . coli O157.

BMC Surg . 2004 Sep 29;4(1):12.
Probiotic prophylaxis in patients with predicted severe acute pancreatitis (PROPATRIA): design and rationale of a double-blind, placebo-controlled randomised multicenter trial {ISRCTN38327949}; Besselink MG et al.; BACKGROUND: Infectious complications are the major cause of death in acute pancreatitis . Small bowel bacterial overgrowth and subsequent bacterial translocation are held responsible for the vast majority of these infections . Goal of this study is to determine whether selected probiotics are capable of preventing infectious complications without the disadvantages of antibiotic prophylaxis; antibiotic resistance and fungal overgrowth . METHODS/DESIGN: PROPATRIA is a double-blind, placebo-controlled randomised multicenter trial in which 200 patients will be randomly allocated to a multispecies probiotic preparation (Ecologic 641) or placebo . The study is performed in all 8 Dutch University Hospitals and 7 non-University hospitals . The study-product is administered twice daily through a nasojejunal tube for 28 days or until discharge . Patients eligible for randomisation are adult patients with a first onset of predicted severe acute pancreatitis: Imrie criteria 3 or more, CRP 150 mg/L or more, APACHE II score 8 or more . Exclusion criteria are post-ERCP pancreatitis, malignancy, infection/sepsis caused by a second disease, intra-operative diagnosis of pancreatitis and use of probiotics during the study . Administration of the study product is started within 72 hours after onset of abdominal pain . The primary endpoint is the total number of infectious complications . Secondary endpoints are mortality, necrosectomy, antibiotic resistance, hospital stay and adverse events . To demonstrate that probiotic prophylaxis reduces the proportion of patients with infectious complications from 50% to 30%, with alpha 0,05 and power 80%, a total sample size of 200 patients was calculated . CONCLUSION: The PROPATRIA study is aimed to show a reduction in infectious complications due to early enteral use of multispecies probiotics in severe acute pancreatitis.

Int J Syst Evol Microbiol, 2004 Sep, 54(Pt 5), 1459 - 63
Mycobacterium psychrotolerans sp . nov., isolated from pond water near a uranium mine; Trujillo ME et al.; An acid-fast, rapidly growing, psychrotolerant short rod was isolated from pond water near a uranium mine . Phylogenetic analysis of the 16S rRNA gene sequence grouped this strain with the rapidly growing mycobacteria . The 16S rRNA gene sequence of isolate WA101T showed highest similarity to that of Mycobacterium sphagni DSM 44076T; however, DNA-DNA relatedness between the two strains was less than 30 % . Chemotaxonomic analyses, which included fatty acid and mycolic acid patterns, confirmed the classification of strain WA101T in the genus Mycobacterium . Physiological data, including antibiotic resistance, NaCl tolerance, carbon sources, temperature growth range and enzyme activities, were also determined . Based on the genotypic and phenotypic results it is proposed that isolate WA101T represents a novel Mycobacterium species . The name Mycobacterium psychrotolerans sp . nov . is proposed, with type strain WA101T (= DSM 44697T = LMG 21953T).

Antimicrob Agents Chemother, 2004 Oct, 48(10), 3996 - 4001
Mechanisms of resistance in multiple-antibiotic-resistant Escherichia coli strains of human, animal, and food origins; Saenz Y et al.; Seventeen multiple-antibiotic-resistant nonpathogenic Escherichia coli strains of human, animal, and food origins showed a wide variety of antibiotic resistance genes, many of them carried by class 1 and class 2 integrons . Amino acid changes in MarR and mutations in marO were identified for 15 and 14 E . coli strains, respectively.

Pharmacoepidemiol Drug Saf, 2004 Dec, 13(12), 863 - 70
Predictors of receipt of a fluoroquinolone versus trimethoprim-sulfamethoxazole for treatment of acute pyelonephritis in women in Manitoba, Canada; Carrie AG et al.; PURPOSE: The increasing and comparatively high proportion of uropathogens in Canada resistant to trimethoprim-sulfamethoxazole (TMP-SMX) may be partially responsible for the increasing use of fluoroquinolones . A number of patient-specific variables have been identified as risk factors for infections caused by antibiotic-resistant pathogens . However, variables unrelated to need, have also been associated with receipt of broad-spectrum antibiotics . We identified patient variables associated with receipt of a fluoroquinolone versus TMP-SMX for treatment of acute pyelonephritis . METHODS: Healthcare claims from the province of Manitoba, Canada for the period February 1996 to March 1999 were examined to identify episodes of pyelonephritis in non-pregnant females between 18 and 65 years of age treated with TMP-SMX or a fluoroquinolone . Patient variables were identified based on healthcare claims review and data from Statistics Canada . Logistic regression was used to model the probability of receipt of a fluoroquinolone . RESULTS: A total of 1084 women met inclusion criteria; 653 treated with TMP-SMX and 431 treated with a fluoroquinolone . Age, income, rural residence, recent antibiotic use, recent hospitalization and presentation to an emergency room (ER) were positively associated with receipt of a fluoroquinolone . CONCLUSIONS: Patient variables reportedly associated with an increased probability of resistant organisms (e.g., age, recent antibiotic use and recent hospitalization) were significantly associated with an increased probability of receipt of fluoroquinolones . However, variables unrelated to antibiotic resistance (e.g., income, rural residence and presentation to an ER) were also significantly associated with receipt of a fluoroquinolone .

Curr Opin Crit Care, 2004 Oct, 10(5), 364 - 8
Prevention of infection in the intensive care unit; Bonten MJ; PURPOSE OF REVIEW: Infections remain an important threat for critically ill patients, and the emergence of antibiotic resistance is increasingly hampering successful treatment . In this review, new aspects of the diagnosis and prevention of ventilator-associated pneumonia and of strategies of antibiotic use to limit the development and spread of resistance are described . RECENT FINDINGS: Prevention of ventilator-associated pneumonia is cost effective, but the most optimal preventive measure (or set of measures) remains controversial . There is growing evidence that antibiotic prophylaxis reduces the occurrence of ventilator-associated pneumonia and improves patient outcome . Moreover, antibiotic use can be reduced by increasing the specificity of diagnosing ventilator-associated pneumonia and by limiting the duration of antibiotic treatment . SUMMARY: Recent developments in diagnosis, treatment, and prevention of ventilator-associated pneumonia and strategies to reduce emergence of antibiotic resistance have been reviewed . Whether changes in antibiotic policy will reduce the emergence of antibiotic resistance remains to be determined . In this area, methodologic problems that have been overlooked in many studies have been addressed recently . These issues must be clarified to provide reliable data on the effects of interventions in hospital settings .

Infect Immun, 2004 Oct, 72(10), 5555 - 64
The vimE gene downstream of vimA is independently expressed and is involved in modulating proteolytic activity in Porphyromonas gingivalis W83; Vanterpool E et al.; Regulation/activation of the Porphyromonas gingivalis gingipains is poorly understood . A unique 1.3-kb open reading frame downstream of the bcp-recA-vimA transcriptional unit was cloned, insertionally inactivated with the ermF-ermAM antibiotic resistance cassette, and used to create a defective mutant by allelic exchange . In contrast to the wild-type W83 strain, the growth rate of the mutant strain (designated FLL93) was reduced, and when plated on Brucella blood agar it was nonpigmented and nonhemolytic . Arginine- and lysine-specific gingipain activities were reduced by approximately 90 and 85%, respectively, relative to activities of the parent strain . These activities were unaffected by the culture's growth phase, in contrast to the vimA-defective mutant P . gingivalis FLL92, which has increased proteolytic activity in stationary phase . Expression of the rgpA, rgpB, and kgp gingipain genes was unaltered in P . gingivalis FLL93 compared to that of the wild-type strain . Further, in extracellular protein fractions a 64-kDa band was identified that was immunoreactive with the RgpB-specific proenzyme antibodies . Active-site labeling with dansyl-glutamyl-glycyl-arginyl chloromethyl ketone or immunoblot analysis showed no detectable protein band representing the gingipain catalytic domain . In vitro protease activity could be slightly induced by a urea denaturation-renaturation cycle in an extracellular protein fraction, in contrast to the vimA-defective mutant P . gingivalis FLL92 . Expression of flanking genes, including recA, vimA, and Pg0792, was unaltered by the mutation . Taken together, these results suggest that the vimA downstream gene, designated vimE (for virulence-modulating gene E), is involved in the regulation of protease activity in P . gingivalis.

J Mol Microbiol Biotechnol, 2004, 7(4), 197 - 203
Effect of plasmid copy number and lac operator sequence on antibiotic-free plasmid selection by operator-repressor titration in Escherichia coli; Cranenburgh RM et al.; The Escherichia coli strain DH1lacdapD enables plasmid selection and maintenance that is free from antibiotics and selectable marker genes . This is achieved by using only the lac operator sequence as a selectable element . This strain is currently used to generate high copy number plasmids with no antibiotic resistance genes for use as DNA vaccines and for expression of recombinant proteins . Until now these have been limited to pUC-based plasmids containing a high copy number pMB1-derived origin of replication, and the principle lacO(1) and auxiliary lacO(3) operators . In this study we have shown that this system can also be used to select and maintain pBR322-based plasmids with the lower copy number pMB1 origin of replication, and that lacO(1) alone or a palindromic version of lacO(1) can provide a sufficient level of repressor titration for plasmid selection . This is advantageous for recombinant protein production, where low copy number plasmids are often used and plasmid maintenance is important . The degree of repressor titration due to these plasmids was measured using the natural lactose operon in E . coli DH1 as a model .

J Mol Biol, 2004 Oct 8, 343(1), 213 - 22
The reconstitution and activity of the small multidrug transporter EmrE is modulated by non-bilayer lipid composition; Curnow P et al.; The ability of multidrug transport proteins within biological membranes to recognise a diverse array of substrates is a fundamental aspect of antibiotic resistance . Detailed information on the mechanisms of recognition and transport can be provided only by in vitro studies in reconstituted bilayer systems . We describe the controlled, efficient reconstitution of the small multidrug transporter EmrE in a simple model membrane and investigate the effect of non-bilayer lipids on this process . Transport activity is impaired, in line with an increase in the lateral pressure within the bilayer . We demonstrate the potential of this lateral pressure modulation method as a general approach to the folding and assembly of membrane proteins in vitro, by recovering functional transporter from a partly denatured state . Our results highlight the importance of optimising reconstitution procedures and bilayer lipid composition in studies of membrane transporters . This is particularly pertinent for multidrug proteins, and we show that the use of a sub-optimal lipid bilayer environment or reconstitution method could lead to incorrect information on protein activity.

J Virol Methods, 2004 Nov, 121(2), 137 - 43
Coupling generation of cytomegalovirus deletion mutants and amplification of viral BAC clones; Wang W et al.; Human cytomegalovirus (HCMV) genome manipulation has always been difficult . Recently, the introduction of full-length HCMV DNA into Escherichia coli as an artificial bacterial chromosome (BAC) clone has allowed reliable targeted mutagenesis . Here, we show the next generation of improvement in designing recombinant HCMV, which will also be applicable to other viral BAC clones . An inducible origin of replication linked with an antibiotic resistance marker was used as a cassette for targeted replacement of sequences within a HCMV BAC clone, TowneBAC . The origin of replication allowed for the induction of increased amounts BAC DNA that improved recovery, ease of use and transfections for mutant viruses . By specific deletion of UL147 and the recombinant GFP gene, we have shown that targeted deletion of a gene and selection for a recombinant genome are coupled with the ability to amplify the BAC clone DNA . These HCMV BAC clones were amplified approximately 10-fold . In the case of the removal of GFP from the clone TowneBAC shown in this study, the resulting BAC DNA preparation following amplification was used for successful primary cell transfection . Both parental and deletion BAC clone transfections gave similar levels of recombinant HCMV, and the GFP deletion virus replicated the same as the TowneBAC in a multi-step growth curve analysis.

Clin Pharmacol Ther, 2004 Sep, 76(3), 201 - 9
CYP2C19 polymorphism is a major predictor of treatment failure in white patients by use of lansoprazole-based quadruple therapy for eradication of Helicobacter pylori; Schwab M et al.; OBJECTIVE: Proton pump inhibitors, metabolized by the polymorphic enzyme cytochrome P450 (CYP) 2C19, are essential drugs for Helicobacter pylori eradication . It was reported that patients with CYP2C19 wild type in Asia had lower eradication rates . This study tests the hypothesis that CYP2C19 wild type ( wt/wt ) in white patients is also associated with a higher probability of treatment failure . METHODS: This was a cohort study involving 131 H pylori -positive white (German) patients treated by quadruple therapy including lansoprazole (30 mg twice daily for 5 days) . Eradication success, as well as lansoprazole trough steady-state serum concentrations, was determined according to different CYP2C19 genotypes . RESULTS: We found 3 homozygous variant patients (2.3%) ( mt/mt, CYP2C19*2/*2 ), 42 heterozygous patients (32.1%) ( wt/mt, CYP2C19*1/*2 ), and 86 wild-type individuals (65.6%) . Significant differences in eradication success could be found between wt/wt patients (80.2%) versus combined mt/mt (100%) and wt/mt patients (97.8%) ( P <.01; odds ratio, 10.8 {confidence interval, 1.4-84}), which were associated with corresponding changes in the serum levels of lansoprazole (median, 753 ng/mL for mt/mt, 59 ng/mL for wt/mt, and 21 ng/mL for wt/wt; P <.001) . Apart from antibiotic resistance, CYP2C19 polymorphism was the most important influencing factor for eradication success on multivariate analysis ( P <.0001) . CONCLUSION: Eradication rates of H pylori highly depend on CYP2C19 in white patients if standard doses of lansoprazole (30 mg twice daily) are administered within a quadruple regimen . Because wt/wt individuals have lower eradication rates and lower serum concentrations of lansoprazole, these patients might benefit from a higher proton pump inhibitor dosage.

Trends Microbiol, 2004 Sep, 12(9), 412 - 6
Human intestinal bacteria as reservoirs for antibiotic resistance genes; Salyers AA et al.; Human intestinal bacteria have many roles in human health, most of which are beneficial or neutral for the host . In this review, we explore a more sinister side of intestinal bacteria; their role as traffickers in antibiotic resistance genes . Evidence is accumulating to support the hypothesis that intestinal bacteria not only exchange resistance genes among themselves but might also interact with bacteria that are passing through the colon, causing these bacteria to acquire and transmit antibiotic resistance genes.

Biol Reprod, 2004 Dec, 71(6), 1755 - 65 Epub 2004 Dec.
Self-renewal vs . differentiation of mouse embryonic stem cells; O'Shea KS; Embryonic stem (ES) cells are typically derived from the inner cell mass of the preimplantation blastocyst and can both self-renew and differentiate into all the cells and tissues of the embryo . Because they are pluripotent, ES cells have been used extensively to analyze gene function in development via gene targeting . The embryonic stem cell is also an unsurpassed starting material to begin to understand a critical, largely inaccessible period of development . If their differentiation could be controlled, they would also be an important source of cells for transplantation to replace cells lost through disease or injury or to replace missing hormones or genes . Traditionally, ES cells have been differentiated in suspension culture as embryoid bodies, named because of their similarity to the early postimplantation-staged embryo . Unlike the pristine organization of the early embryo, differentiation in embryoid bodies appears to be largely unpatterned, although multiple cell types form . It has recently been possible to separate the desired cell types from differentiating ES cells in embryoid bodies by using cell-type-restricted promoters driving expression of either antibiotic resistance genes or fluorophores such as EGFP . In combination with growth factor exposure, highly differentiated cell types have successfully been derived from ES cells . Recent technological advances such as RNA interference to knock down gene expression in ES cells are also producing enriched populations of cells and elucidating gene function in early development.

Int J Antimicrob Agents, 2004 Sep, 24(3), 205 - 12
Animal growth promoters: to ban or not to ban? A risk assessment approach; Kelly L et al.; The use of antibiotics for animal growth promotion has been controversial because of the potential transfer of antibiotic resistance from animals to humans . Such transfer could have severe public health implications in that treatment failures could result . We have followed a risk assessment approach to evaluate policy options for the streptogramin-class of antibiotics: virginiamycin, an animal growth promoter, and quinupristin/dalfopristin, a antibiotic used in humans . Under the assumption that resistance transfer is possible, models project a wide range of outcomes depending mainly on the basic reproductive number (R(0)) that determines the potential for person-to-person transmission . Counter-intuitively, the benefits of a ban on virginiamycin were highest for intermediate values of R(0), and lower for extremely high or low values of R(0).

Curr Med Res Opin, 2004 Aug, 20(8), 1301 - 7
The efficacy, safety and tolerability of pantoprazole-based one-week triple therapy in H . pylori eradication and duodenal ulcer healing; Sivri B et al.; OBJECTIVE: Recently, proton pump inhibitor (PPI)-based triple therapy has been recommended as a first line treatment in the eradication of Helicobacter pylori . The aim of this open, multicentre trial was to investigate the efficacy, safety, tolerability and the ulcer healing rate of a triple regimen consisting of pantoprazole 40 mg, clarithromycin 500 mg and amoxicillin 1000 mg twice daily for 7 days, in the eradication of H . pylori in patients with duodenal ulcer in Turkey . RESEARCH DESIGN AND METHODS: H . pylori infection was assessed by histological examination and rapid urease test at baseline and 4 weeks after the completion of the therapy . Seventy-seven patients were enrolled, 5 were excluded due to various reasons and 72 completed the entire course of the trial . RESULTS: H . pylori eradication was confirmed in 49 of these patients; the eradication rate was 68% by per-protocol analysis and 63.6% by intention-to-treat analysis . The ulcers were completely healed in 61 patients (85%) at the second endoscopic examination . Drug compliance was excellent (97.3%) and there were no serious adverse events . CONCLUSION: Pantoprazole-based 1-week triple therapy was well tolerated and the ulcer healing rate was high (85%) . Relatively low H . pylori eradication rates may be attributed to rising antibiotic resistance over recent years . A large scale, comparative study with other PPI-based regimens is warranted based on the results of this open study with the pantoprazole-based regimen.

Environ Microbiol, 2004 Sep, 6(9), 981 - 9
Uncultured soil bacteria are a reservoir of new antibiotic resistance genes; Riesenfeld CS et al.; Antibiotic resistance genes are typically isolated by cloning from cultured bacteria or by polymerase chain reaction (PCR) amplification from environmental samples . These methods do not access the potential reservoir of undiscovered antibiotic resistance genes harboured by soil bacteria because most soil bacteria are not cultured readily, and PCR detection of antibiotic resistance genes depends on primers that are based on known genes . To explore this reservoir, we isolated DNA directly from soil samples, cloned the DNA and selected for clones that expressed antibiotic resistance in Escherichia coli . We constructed four libraries that collectively contain 4.1 gigabases of cloned soil DNA . From these and two previously reported libraries, we identified nine clones expressing resistance to aminoglycoside antibiotics and one expressing tetracycline resistance . Based on the predicted amino acid sequences of the resistance genes, the resistance mechanisms include efflux of tetracycline and inactivation of aminoglycoside antibiotics by phosphorylation and acetylation . With one exception, all the sequences are considerably different from previously reported sequences . The results indicate that soil bacteria are a reservoir of antibiotic resistance genes with greater genetic diversity than previously accounted for, and that the diversity can be surveyed by a culture-independent method.

Rev Prat, 2004 May 15, 54(9), 963 - 7
{Antibiotic treatment of bacterial meningitis}; Stahl JP; Community-acquired bacterial meningitis are due, in most of the cases, to S . pneumoniae and N . meningitidis . These two pathogenes are concerned by antibiotic resistance phenomenons . S . pneumoniae is developing resistance in some countries, including France . The recommended empirical treatment is nowadays a third-generation cephalosporin, combined with vancomycin if there is a risk of Penicillin resistant pneumococcus . Non-antibiotic drugs are now validated and very useful adjunctive treatment.

Asia Pac J Clin Nutr . 2004 Aug;13(Suppl):S92.
The role of oligosaccharides and Helicobacter pylori-specific antibodies in disease prevention; Campbell MA et al.; Background- Problems with current therapies for management of Helicobacter pylori-associated disease are patient compliance and increasing antibiotic resistance . Therefore, prevention of disease via nutritional intervention is a practical alternative approach for management of Helicobacter pylori (H . pylori) infection . Objective- This study was carried out to assess the ability of nutritional components, namely oligosaccharides and H . pylori-specific antibodies, to prevent acquisition of disease . Design- The interventions investigated were 0.25 % (w/v) hyperimmune bovine colostrum (HBC), 25% (w/v) acidic oligosaccharides (AOS) and 30% (w/v) fructo- and galacto-oligosaccharides (FOS/GOS), administered alone or in combination twice daily by orogastric gavage (0.2 ml per mouse on each occasion) from day 1 onwards . Control animals were given water by the same method . Mice were then challenged on day 14 with 1 x 10(8) viable H . pylori, Sydney strain 1 (SS1; 1 x 10(9)/ml) in sterile saline (0.1 ml) by orogastric gavage . After challenge, intervention was continued for a further 21 days . The next day (day 36), blood was withdrawn for antibody testing then mice were killed and the stomach removed for histology and bacterial culture . Outcomes- None of the interventions studied prevented colonisation by H . pylori . However, gastritis scores were lower in treatment versus control mice . Antibody titres were also different between groups, lower in all oligosaccharide-treated mice and higher in mice treated with HBC alone, compared with control mice . Conclusions- Nutritional components may be beneficial in H . pylori infection and in preventing progress of disease.

Int J Nurs Stud, 2004 Sep, 41(7), 745 - 53
Development of the choices and acquisition of antibiotics model from a descriptive study of a lay Honduran population; Crigger NJ et al.; Antibiotic resistance is a global public health problem that is accelerated by overuse and misuse of antibiotics . In today's world of increasing international travel and exchange of goods, the spread of antibiotic resistant organisms is a growing threat . Despite significant antibiotic use in developing nations, research to describe and curtail inappropriate use is limited . In this study, the investigators developed a model of antibiotic use, choices and acquisition of antibiotics model, from a study of a lay population in Honduras . A representative sample of 939 rural and urban Hondurans completed the Preguntas Para El Uso de Antibiotics questionnaire to determine how the participant made choices about antibiotic use . The study indicated that the rural participants used significantly fewer antibiotics than the urban participants and that the demographic indicators did not show a significant difference in antibiotic use in those of lower socioeconomic status . In addition, the participants reported that they seek out professional advice and care rather than self-prescribing . Implications for educational and empowerment programs based on the model are discussed.

Klin Padiatr, 2004 Jul-Aug, 216(4), 215 - 24
{Acute otitis media in children: an evidence-based practice guideline}; Koneczny N et al.; Acute Otitis media is one of the most common acute respiratory infections managed in primary care and the most common infection among in children . Diagnostic criteria, however, do not always correspond to scientific evidence . They often differ depending on individual preferences and competences . Treatment, also, is controversial . In Germany, most children attending their pediatrician or primary care physician will be prescribed antibiotics . Evidence from several randomized studies and systematic reviews suggests that routine usage of antibiotics provides only modest benefit . The benefit of prescribing antibiotics should not only be balanced against the increased likelihood of side effects such as diarrhoea but also against the potential to contribute to longterm antibiotic resistance.

Antimicrob Agents Chemother, 2004 Aug, 48(8), 3141 - 6
Lack of evidence that DNA in antibiotic preparations is a source of antibiotic resistance genes in bacteria from animal or human sources; Lau SK et al.; Although DNA encoding antibiotic resistance has been discovered in antibiotic preparations, its significance for the development of antibiotic resistance in bacteria is unknown . No phylogenetic evidence was obtained for recent horizontal transfer of antibiotic resistance genes from antibiotic-producing organisms to bacteria from human or animal sources.

Methods Mol Biol, 2004, 267, 135 - 43
Plasmid vectors for marker-free chromosomal insertion of genetic material in Escherichia coli; Le Borgne S et al.; A method to achieve the insertion of genetic material into the chromosome of Escherichia coli is described . The method is based on the use of integration vectors from the pBRINTs-rAnbR family . These vectors offer the choice of using the antibiotics chloramphenicol, gentamycin, or kanamycin to select for chromosomal integration events . In addition, it is possible to eliminate these chromosomal antibiotic resistance markers, after integration has taken place . The overall insertion strategy is as follows: a fragment containing the gene(s) to be integrated in the chromosome is inserted into the multiple cloning site of a pBRINTs-rAnbR vector and the resulting plasmid is used to transform E . coli cells . The plasmid is first allowed to replicate in the cell at the permissive temperature of 30 degrees C . Next, the temperature of the culture is raised to 44 degrees C to inhibit plasmid replication and to select for the integrants in the presence of the appropriate antibiotic . Chromosomal excision of the AnbR gene can then be catalyzed by the Cre recombinase that is transiently expressed in the cell from the temperature-sensitive pJW168 plasmid . This plasmid is finally eliminated from the cells by increasing the temperature of the culture to 44 degrees C.

Biotechnol Lett, 2004 Jul, 26(14), 1107 - 13
Construction of a novel shuttle vector based on an RCR-plasmid from a haloalkaliphilic archaeon and transformation into other haloarchaea; Zhou M et al.; The pNB101 is the first plasmid to be isolated from an haloalkaliphilic archaea . With insertion of the ColE1 replicon of Escherichia coli, as well as two antibiotic resistance genes at its unique Hin dIII site, a novel shuttle vector between haloarchaea and E . coli was developed . This vector, named pNB102, was successfully transformed into two non-alkaliphilic haloarchaea, Halobacterium salinarum SNOB and Haloarcula hispanica ATCC33960 . The presence and stability of pNB102 in the transformants were confirmed by PCR identification, Southern blotting and restriction endonuclease digestion . Results also indicated that the presence of restriction-modification (R-M) systems in some Halobacterium species prevented this transformation . It is the first report that the replicon of pNB101 has such a wide host range, and has taken the first step for construction of the vector/host system in haloalkaliphilic archaea.

CMAJ, 2004 Jul 20, 171(2), 133 - 8
Decrease in antibiotic use among children in the 1990s: not all antibiotics, not all children; Kozyrskyj AL et al.; BACKGROUND: Decreases in antibiotic use were widely reported in the 1990s . This study was undertaken to determine trends in the use of antibiotics from fiscal year (FY) 1995 (April 1995 to March 1996) to FY 2001 in a complete population of Manitoba children . METHODS: Using Manitoba's health care databases, we determined annual population-based rates of antibiotic prescription among children by antibiotic class (narrow-spectrum and broader-spectrum antibiotics), age group, physician diagnosis (e.g., otitis media or bronchitis) and neighbourhood income in urban areas (derived from the 1996 census) . Antibiotic prescription rates were generated within a generalized linear model framework with general estimating equations, and differences between FY 2001 and FY 1995 were tested . Differences in antibiotic use over time were compared across antibiotic classes, age groups, diagnoses and income neighbourhoods . RESULTS: The overall antibiotic prescription rate decreased by almost one-third, from 1.2 prescriptions per child in FY 1995 to 0.9 prescriptions in FY 2001 . Total antibiotic use declined for all respiratory tract infections; decreases were greatest for the sulfonamides (decrease to less than one-third the FY 1995 rate) and narrow-spectrum macrolides (decrease to less than half the FY 1995 rate) . In contrast, the FY 2001 rate for broader-spectrum macrolides was as much as 12.5 times the FY 1995 rate . Otitis media accounted for one-quarter of the use of the latter agents . Preschool children and low-income children received the greatest number of antibiotic prescriptions . Declines in antibiotic prescriptions were of a lesser magnitude for low-income children (for whom rates in FY 2001 were four-fifths the rates in FY 1995) than for higher-income children (for whom rates in FY 2001 were about two-thirds the rates in FY 1995) . INTERPRETATION: Overall, antibiotic use declined over the late 1990s in this population of Canadian children, but the increasing use of broader-spectrum macrolides and higher rates of antibiotic use among preschool and low-income children may have implications for antibiotic resistance.

Appl Microbiol Biotechnol, 2004 Oct, 65(5), 566 - 75 Epub 2004 Jul 10.
Chestnut bur-shaped aggregates of chrysotile particles enable inoculation of Escherichia coli cells with plasmid DNA; Yoshida N et al.; In the present study, Escherichia coli cells exhibited antibiotic resistance after transformation with exogenous plasmid DNA adsorbed onto chrysotile particles during agar-exposure . We previously demonstrated penetration of E . coli by chrysotile particles during agar-exposure . To further investigate the mechanism by which transformation of E . coli is achieved through the use of chrysotile fibers, the interaction between E . coli cells and chrysotile was examined during agar-exposure . Dispersion of chrysotile particles within the chrysotile solution was analyzed by flow cytometry . A suspension containing E . coli cells expressing blue fluorescence protein and chrysotile particles was exposed to agar using stirring apparatus, which allowed a constant vertical reaction force to be applied to the surface of the gel . Fluorescence microscopy was then used to illustrate the adsorption of fluorescein isothiocyanate-conjugated DNA oligomers to chrysotile . Larger aggregates were observed when increasing concentrations of chrysotile were added to the solution . With prolonged exposure, during which surface moisture diffused into the agar gel, greater concentrations of chrysotile were observed on the agar surface . In addition, chrysotile aggregates exceeding 50 microm developed on the agar surface . They were shaped like a chestnut bur . The chrysotile aggregates penetrated the cell membranes of adherent E . coli cells during agar-exposure due to sliding friction forces generated at the interface of the agar and the stirring stick . E . coli cells thus acquired plasmid DNA and antibiotic resistance, since the plasmid DNA had been adsorbed onto the chrysotile particles . The inoculation of plasmid DNA into E . coli cells demonstrates the usefulness of chrysotile for E . coli transformation.

Anal Biochem, 2004 Aug 1, 331(1), 153 - 60
A one-step approach to obtain cell clones expressing tetracycline-responsive transactivators; Munoz I et al.; Despite the wide application of the tetracycline-regulated gene expression system, several drawbacks in establishing the system in in vitro-cultured cells have been described . Most of the problems are related to obtaining a reliable tetracycline-regulated cell clone, which often results in arduous labor . We describe here a new approach to facilitate the screening and selection of such cell clones . We have constructed a tetracycline-responsive plasmid that harbors an antibiotic resistance gene fused to the enhanced green fluorescent protein (EGFP) gene and the luciferase gene, both under the control of a bidirectional promoter . We demonstrate that the selection of tetracycline-regulated clones is highly simplified by using this plasmid . Only clones expressing the system in a functional manner are able to survive under antibiotic selection . In addition, a quick characterization of the responsiveness of the clones is possible by monitoring GFP expression in vivo.

Curr Opin Ophthalmol, 2004 Aug, 15(4), 316 - 20
Fourth-generation fluoroquinolones: new topical agents in the war on ocular bacterial infections; Mah FS; PURPOSE OF REVIEW: The fourth-generation fluoroquinolones, moxifloxacin and gatifloxacin, were introduced in 2003 promising improved spectrum of activity and delayed development of resistance . Although these topical agents have recently been introduced in commercial form, there is already a growing body of evidence showing excellent potency in the war on ocular infections . The purpose of this review is to discuss the literature to date regarding these two agents . RECENT FINDINGS: Since their introduction in 1990 in the United States, fluoroquinolones have rapidly become the standard of care in the topical antibiotic arena . Unfortunately, recent evidence has shown the widespread use of fluoroquinolones, not only in eye care, but also in agriculture, and general medical and surgical use, has lead to decreasing susceptibilities of important ocular bacterial pathogens . Moxifloxacin and gatifloxacin have improved potency and are able to overcome resistant isolates . These agents also provide improved penetration into ocular tissues . SUMMARY: Moxifloxacin and gatifloxacin offer improved spectrum of activity, increased penetration into ocular tissues, and delayed propensity to the development of bacterial antibiotic resistance.

Mol Microbiol, 2004 Jul, 53(1), 183 - 91
Transcriptional and translational regulation of the marRAB multiple antibiotic resistance operon in Escherichia coli; Martin RG et al.; The marRAB multiple antibiotic resistance operon of Escherichia coli is autorepressed by MarR . MarR binds to two palindromic sequences in vitro: site I lies between and overlaps the -35 and -10 hexamers for RNA polymerase binding; site II lies between the transcription start site and the GTG initiation codon of marR . To assess the importance of these sites in vivo, the effects of mutant sites on transcription were analysed using fusions to lacZ in the presence and absence of wild-type MarR . When both sites were wild type, transcription in the derepressed marR-deleted strain was 19-fold that of the wild-type strain; when only site I or site II was wild type, this ratio was reduced to 4.3- and 2.6-fold, respectively, showing that full repression requires both sites, but some repression can occur at one site independently of the other . Translational fusions of the wild-type promoter to lacZ demonstrated that marR translation proceeds at only 4.5% of the transcription rate . Analysis of translational fusions with mutant leader sequences demonstrated that the principal reason for inefficient translation is a weak Shine-Dalgarno (SD) sequence, AGG(G) . Although the SD sequence is located within the potential stem-loop structure of site II, no evidence for occlusion of the SD sequence was found in the wild-type strain . However, a single basepair mutation that strengthens the stem-loop structure drastically reduced the translational efficiency . Substitution of ATG for GTG as the initiation codon increased translational efficiency by 50% . Increasing the 5 bp spacing between the SD sequence and the GTG codon by one to four bases reduced the translational efficiency by 50-75% . Inefficient translation of marR may help to sensitize the cell to environmental signals.

Chemistry, 2004 Jul 5, 10(13), 3308 - 14
Aminoglycoside microarrays to explore interactions of antibiotics with RNAs and proteins; Disney MD et al.; RNA is an important target for drug discovery efforts . Several clinically used aminoglycoside antibiotics bind to bacterial rRNA and inhibit protein synthesis . Aminoglycosides, however, are losing efficacy due to their inherent toxicity and the increase in antibiotic resistance . Targeting of other RNAs is also becoming more attractive thanks to the discovery of new potential RNA drug targets through genome sequencing and biochemical efforts . Identification of new compounds that target RNA is therefore urgent, and we report here on the development of rapid screening methods to probe binding of low molecular weight ligands to proteins and RNAs . A series of aminoglycosides has been immobilized onto glass microscope slides, and binding to proteins and RNAs has been detected by fluorescence . Construction and analysis of the arrays is completed by standard DNA genechip technology . Binding of immobilized aminoglycosides to proteins that are models for study of aminoglycoside toxicity (DNA polymerase and phospholipase C), small RNA oligonucleotide mimics of aminoglycoside binding sites in the ribosome (rRNA A-site mimics), and a large (approximately 400 nucleotide) group I ribozyme RNA is detected . The ability to screen large RNAs alleviates many complications associated with binding experiments that use isolated truncated regions from larger RNAs . These studies lay the foundation for rapid identification of small organic ligands from combinatorial libraries that exhibit strong and selective RNA binding while displaying decreased affinity to toxicity-causing proteins.

Oral Microbiol Immunol, 2004 Aug, 19(4), 233 - 9
Alkyl hydroperoxide peroxidase subunit C (ahpC) protects against organic peroxides but does not affect the virulence of Porphyromonas gingivalis W83; Johnson NA et al.; The cloned Porphyromonas gingivalis alkyl hydroperoxide reductase (ahpC) gene complemented an ahpC defect in Escherichia coli . To study the role of ahpC in protecting against oxidative stress in P . gingivalis a 1.8 kb fragment containing the ahpC gene was amplified from the chromosome of P . gingivalis W83 . This gene was insertionally inactivated using the ermF-ermAM antibiotic resistance cassette and used to create a ahpC-deficient mutant by allelic exchange . One mutant strain, designated FLL141, demonstrated no change in the growth rate, black pigmentation, beta-hemolysis or level of proteolytic activity compared to the parent strain . Although P . gingivalis FLL141 was more sensitive to hydrogen peroxide than the parent strain, there was no change in its virulence potential in the mouse model compared to the wild-type strain . These findings suggest that the ahpC gene plays a role in peroxide resistance in P . gingivalis but does not contribute significantly to virulence.

Bundesgesundheitsblatt Gesundheitsforschung Gesundheitsschutz, 2004 Apr, 47(4), 345 - 51
{SARI: surveillance of antibiotic use and bacterial resistance in German intensive care units . Correlation between antibiotic use and the emergence of resistance}; Meyer E et al.; Intensive care units (ICUs) are considered to be high-risk areas for the emergence and spread of multiresistant bacterial pathogens . In Germany, there are no representative epidemiological data on antibiotic resistance, on the use of antibiotics in ICUs, or on the correlation between antibiotic use and the emergence of resistance . Project SARI (surveillance of antibiotic use and bacterial resistance in ICUs), which as a part of the epidemiological network Spread of Nosocomial Infections and Resistant Pathogens (SIR) is supported by the German Ministry of Science and Education, started in February 2000 and meanwhile includes data on antibiotic use and resistance rates in 38 medical, surgical, and interdisciplinary ICUs . To date (February 2000-June 2003), a total of 1142 months, 413,065 patient days, and 550,288 defined daily doses (DDDs in accordance with the WHO) have been covered with a mean antibiotic usage density (AD) of 1335 DDDs/1000 patient days and resistance data on 37,612 isolates from ICUs . Ciprofloxacin use and MRSA correlate significantly, as do imipenem use and the rate of imipenemresistant P . aeruginosa.The genodiversity of P . aeruginosa is lower in ICUs with high resistance rates and low use of imipenem than in ICUs with high resistance rates and high use . This is an indirect parameter of transmission of identical strains.The epidemiological data of SARI form a basis for improved antibiotic and infection control management in ICUs .

J Biotechnol, 2004 Jul 1, 111(1), 17 - 30
A host/plasmid system that is not dependent on antibiotics and antibiotic resistance genes for stable plasmid maintenance in Escherichia coli; Hagg P et al.; Uneven distribution of plasmid-based expression vectors to daughter cells during bacterial cell division results in an increasing proportion of plasmid free cells during growth . This is a major industrial problem leading to reduction of product yields and increased production costs during large-scale cultivation of vector-carrying bacteria . For this reason, a selection must be provided that kills the plasmid free cells . The most conventional method to obtain this desired selection is to insert some gene for antibiotic resistance in the plasmid and then grow the bacteria in the presence of the corresponding antibiotic . We describe here a host/plasmid Escherichia coli system with a totally stable plasmid that can be maintained without the use of antibiotic selection . The plasmid is maintained, since it carries the small essential gene infA (coding for translation initiation factor 1, IF1) in an E . coli strain that has been deleted for its chromosomal infA gene . As a result only plasmid carrying cells can grow, making the strain totally dependent on the maintenance of the plasmid . A selection based on antibiotics is thus not necessary during cultivation, and no antibiotic-resistance genes are present neither in the final strain nor in the final plasmid . Plasmid-free cells do not accumulate even after an extended period of continuous growth . Growth rates of the control and the plasmid harboring strains are indistinguishable from each other in both LB and defined media . The indicated approach can be used to modify existing production strains and plasmids to the described concept . The infA based plasmid stability system should eliminate industrial cultivation problems caused by the loss of expression vector and use of antibiotics in the cultivation medium . Also environmental problems caused by release of antibiotics and antibiotic resistance genes, that potentially can give horizontal gene transfer between bacterial populations, are eliminated.

J Am Pharm Assoc (Wash DC), 2004 May-Jun, 44(3), 387 - 95; quiz 395-6
Using buzz marketing to promote ideas, services, and products; Holdford DA; OBJECTIVE: To (1) discuss buzz marketing, contrast it with traditional forms of promotional communications, and provide guidelines for use and (2) describe a successful buzz-marketing program used by Sentara Healthcare to decrease overuse and inappropriate use of antibiotic medications . DATA SOURCES: An English-language-only literature search of ABI Inform, Lexus-Nexus, InfoTrac, and university library databases from 1980 to the present using the keywords buzz, word of mouth, opinion leader, and thought leader . Articles and books were cross referenced for other works of interest . STUDY SELECTION: Performed by the author for their contribution to an exploratory analysis of this topic . DATA EXTRACTION: Performed by the author . DATA SYNTHESIS: Buzz marketing is an indirect communications method that has been used successfully in the promotion of a wide variety of products, services, and ideas . By identifying and cultivating nonmedia opinion leaders, the technique generates word-of-mouth communications between these early adopters of products and services and the early and late majority of people who tend to follow their lead . Opinion leaders can be categorized as ordinary or extraordinary, technical or social, and specialist or generalist, depending on the nature of their communications, expertise, and range of knowledge . Buzz marketing is most useful for ideas that are memorable, produce small changes in behavior that have big effects over time, and have the potential to reach a "tipping point" in terms of momentum among a target population . Pharmacists can use buzz marketing for promoting innovative services such as pharmaceutical care . A case study is presented on the use of buzz marketing by a health system for decreasing antibiotic resistance through lessening of public demand for antibiotics and support of physicians in prescribing the agents appropriately . CONCLUSION: Buzz marketing is a potent force in the promotion of pharmaceuticals and can be used by pharmacists . It works best when patients perceive the benefits of innovations . Providing samples and demonstrations of the innovation will foster positive perceptions . Innovations also spread better when they are compatible with the needs, desires, and preferences of individuals and can be adapted to the unique situation of the adopter.

Rev Prat, 2003 Sep 30, 53(14), 1561 - 5
{How to prescribe fewer antibiotics in children?}; Cohen R; During the last years, there has been an alarming worldwide increase in antibiotic resistance among bacterial pathogens of the respiratory tract, and particularly pneumococcus . This is generally attributed to extensive use of antibiotics prescribed for respiratory tract infection and the selection pressure they exert on bacterial strains of nasopharyngeal flora . In our opinion, five actions have to be coordinated in order to control this phenomenon . To improve the continuing education of practitioners and the diagnosis between viral and bacterial respiratory infections, to restrict the use for the situations where antibiotics are really necessary, to prescribe in these situations at the right dosage and duration the more potent antibiotics, to plan straight-a-way an alternative treatment in case of failures, finally achieve a prompt implementation of the pneumococcal conjugate vaccine.

Rev Prat, 2003 Sep 30, 53(14), 1527 - 32
{Antibiotic use: excesses and consequences}; Choutet P; Antibiotic use in France is excessive . This statement is regularly expressed . The comparison with usage in other European countries, the frequency of use in viral infections, the misuse, specifically in the duration of treatment is easy proof of this . However, does this excessive use really have negative consequences in terms of public health? Yes, through the development of antibiotic resistance in hospital bacteria and also in the community . The phenomenon of selection has been amplified as much in the hospital as in the community due to the increase in antibiotic use . It has been a problem that has been neglected for a long time due to the regular release of new antibiotics with better performance onto the market . More than ever, as with any side effect, it has to be taken into account with each prescription of an antibiotic . It encourages the development of a reservoir of resistant bacteria and creates a collective medical risk of the appearance of infections for which the choice of treatment is at best restricted and costly, and at worst inexistent.

Health Econ, 2004 Jun, 13(6), 575 - 83
How property rights and patents affect antibiotic resistance; Horowitz JB et al.; Antibiotic resistance tends to increase when a patent on an antibiotic expires . Since other companies can now sell the antibiotic, more of the antibiotic is produced and prices fall . Because the benefits of reducing current production go to other firms, pharmaceutical companies will have little concern about future resistance . This 'open-access' problem causes excessive antibiotic use and resistance problems in the future . Extending patents is one solution . However, a pharmaceutical company that has patent protection on a drug that is cross-resistant may have little concern about future resistance . This is because when people use completely different antibiotics which cause bacteria to become resistant to the original antibiotic, then the benefits of reducing current production go to other companies . A single buyer such as national health insurance or private health insurance may also have an incentive to reduce antibiotic resistance since they bear the future cost of future resistance . However, insurance coverage reduces the price that patients pay at the margin and thus the patients are likely to use more antibiotics . National health insurance policies may even set the price of antibiotics so low that resistance problems are created even when the patent is in effect .

J Vasc Interv Radiol, 2004 Jun, 15(6), 547 - 56
Antibiotic prophylaxis in interventional radiology; Ryan JM et al.; Despite several decades of advances in both minimally invasive techniques and antibiotic therapy, infection remains one of the more common complications of invasive procedures . Interventional radiology (IR) has traditionally been believed to be associated with lower infection rates than surgery . However, new interventional techniques, as well as more aggressive therapeutic interventions, have presented new challenges in relation to pharmacological management of postprocedural infection and pain . The risk of infection associated with IR procedures can never be completely eliminated, and the reasons for this are manifold, including more virulent organisms, ongoing and newly emerging antibiotic resistance, increased numbers of immunocompromised patients, and the adoption into everyday interventional practice of more aggressive interventional techniques such as chemoembolization, uterine fibroid embolization, and complex biliary intervention . Despite the widespread use of prophylactic antibiotics in IR, and the widely held belief that they are beneficial and are the standard of care, randomized controlled clinical trials have never validated the use of antibiotics in this setting . As such, an argument could be made not to use antibiotics at all for prophylaxis in IR . The purpose of this article is to discuss some of the issues relating to the use of prophylactic antibiotics, and what choice of antibiotics physicians make when they decide to use prophylaxis for IR procedures.

Clin Lab Med, 2004 Jun, 24(2), 329 - 41
Antibiotic-resistant infections in the critically ill adult; Silveira F et al.; Intensive care units (ICUs) frequently are the epicenter of nosocomial infections with antibiotic-resistant bacteria . Optimization of antibiotic therapy for seriously ill patients with bacterial infections appears to have a strong influence on outcome . Laboratories can aid in provision of appropriate antibiotic therapy by providing clinicians with "antibiograms" to aid empiric antibiotic choice and by providing minimal inhibitory concentrations of key antibiotics so that antibiotic dosing is optimized to key pharmacodynamic targets . Laboratories also play a crucial role in the prevention of antibiotic resistance in the ICU . Molecular epidemiologic evidence of an oligoclonal outbreak of infections orients prevention measures toward investigation of common environmental sources of infection and prevention of patient-to-patient transmission . In contrast, evidence of polyclonality shifts prevention of antibiotic resistance to antibiotic management strategies.

J Biochem Biophys Methods, 2004 Jun 30, 59(3), 217 - 27
A quantitative real-time PCR assay for the detection of tetR of Tn10 in Escherichia coli using SYBR Green and the Opticon; Morsczeck C et al.; Bacteria of implant infections are extremely resistant to antibiotics . One reason for this antibiotic resistance are transposons; the well-known transposon Tn10, for example, mediates tetracycline resistance to Escherichia coli . Two genes of Tn10, tetA and tetR, are essential for the mechanism of resistance . These genes encode a drug-specific efflux protein and a tetracycline repressor protein, respectively . Tn10 is also widely used in molecular biology . For example, tTA, a recombinant derivate of tetR, has been utilised for a highly efficient gene regulation system in mammalian cells . We have examined E . coli isolates from implant infections for tetracycline resistance and for the presence of tetR . A real-time PCR assay was developed for detection of tetR with SybrGreen using the Opticon PCR machine of MJ Research . This method offers a quick, sensitive, efficient, and reliable approach to the detection and quantification of genes . Clinical isolates of E . coli were examined successfully for tetracycline resistance and for the presence of tetR . The real-time PCR is effective using a variety of templates including isolated E . coli DNA, pure colonies, or liquid culture sources . Using quantified standard DNA, this assay can accurately detect as few as 15 copies . Moreover, this assay has the ability to quantify the number of tetR genes in the presence of contaminating mammalian DNA . In conclusion, the tetR real-time PCR offers new methods for detection and quantification of tetracycline-resistant bacteria and tTA in transfected cell-lines or transgenic animals.

FEMS Microbiol Lett, 2004 Jun 1, 235(1), 89 - 94
Generation of Leishmania mutants lacking antibiotic resistance genes using a versatile hit-and-run targeting strategy; Denise H et al.; The development of a method to create defined mutants of Leishmania parasites lacking foreign genes conferring resistance to antibiotics has both experimental and practical applications . Mutants deficient in specific virulence genes have potential as attenuated live vaccines, but these can only be of clinical relevance if the antibiotic resistance genes used for selection of the mutants are subsequently removed . In addition, the limited number of antibiotic resistance genes that can be used for genetic manipulation of Leishmania means that a system for recycling them for subsequent use would be highly beneficial when multiple genetic modifications are wanted . In the method we report here, a cassette carrying in tandem the hygromycin resistance gene as a positive marker and thymidine kinase gene as a negative marker is first integrated into the locus of interest and then replaced by a null targeting fragment containing no exogenous DNA . The application of this hit-and-run strategy for removal of one allele of the CPB cysteine peptidase gene array of Leishmania infantum is described.

Bull Entomol Res, 2004 Apr, 94(2), 179 - 88
An interspersed refuge for Sitodiplosis mosellana (Diptera: Cecidomyiidae) and a biocontrol agent Macroglenes penetrans (Hymenoptera: Pteromalidae) to manage crop resistance in wheat; Smith MA et al.; An interspersed refuge of susceptible plants in a resistant, spring-sown wheat crop was tested as a strategy to protect crop resistance against evolution of virulence by the wheat midge Sitodiplosis mosellana (Gehin), and also to conserve a biocontrol agent Macroglenes penetrans(Kirby) . Eight replicated field experiments were conducted using seed mixtures of 0, 5, 10, 15 and 100% or 0, 5 and 100% susceptible wheat with an agronomically similar wheat expressing the antibiotic resistance gene Sm1 . The frequencies of eggs, mature larvae and parasitized larvae in susceptible and resistant wheat spikes, and midge-affected seeds in the harvest, were recorded for each plot . In susceptible wheat, insect densities and seed damage were typical of those in commercial wheat . In resistant wheat, few larvae completed development, 2% or less compared with about 80% in susceptible wheat, when larvae were sampled at maturity . This resistant wheat also deterred midge oviposition, reducing egg densities by 65% compared with susceptible wheat . The wheat midge and its parasitoid oviposited throughout the plots, and parasitism was density independent . The densities of mature midge larvae and parasitoids were in proportion to the size of the refuge . A 5% susceptible refuge produced about 41 mature larvae for each mature larva from the resistant wheat, and provided effective control of damage . An interspersed refuge of susceptible plants in resistant wheat is a promising strategy for sustaining resistance conferred by Sm1 and biocontrol of the wheat midge.

Aliment Pharmacol Ther, 2004 Jun 1, 19(11), 1137 - 46
Review article: Deoxyribonucleic acid-based diagnostic techniques to detect Helicobacter pylori; Ruzsovics A et al.; Helicobacter pylori is an important cause of many gastrointestinal disorders, ranging from chronic gastritis to gastric lymphoma and adenocarcinoma . The deoxyribonucleic acid-based assays have the potential to be a powerful diagnostic tool given its ability to specifically identify H . pylori deoxyribonucleic acid . Markers used to include general H . pylori structures and pathogenetic factors like ureaseA, cagA, vacA, iceA . Deoxyribonucleic acid or bacterial ribonucleic acid for polymerase chain reaction assays can be collected from gastric biopsy, gastric juice, stool, buccal specimens . Polymerase chain reaction can yield quantitative and genotyping results with sensitivity and specificity that approaches 100% . A clear trend in the direction of the determination of quantitative H . pylori infection by real-time polymerase chain reaction can be observed . Fluorescent in situ hybridization is suggested for routine antibiotic resistance determination . To identify the organism, deoxyribonucleic acid structure and its virulence factors may be feasible by using oligonucleotide microarray specifically recognizing and discriminating bacterial deoxyribonucleic acid and various virulence factors . Deoxyribonucleic acid-based H . pylori diagnosis yields higher sensitivity, however, specificity requires sophisticated labour environment and associated with higher costs.

J Biol Chem, 2004 Aug 13, 279(33), 34665 - 73 Epub 2004 May 19.
The importance of a critical protonation state and the fate of the catalytic steps in class A beta-lactamases and penicillin-binding proteins; Golemi-Kotra D et al.; Beta-lactamases and penicillin-binding proteins are bacterial enzymes involved in antibiotic resistance to beta-lactam antibiotics and biosynthetic assembly of cell wall, respectively . Members of these large families of enzymes all experience acylation by their respective substrates at an active site serine as the first step in their catalytic activities . A Ser-X-X-Lys sequence motif is seen in all these proteins, and crystal structures demonstrate that the side-chain functions of the serine and lysine are in contact with one another . Three independent methods were used in this report to address the question of the protonation state of this important lysine (Lys-73) in the TEM-1 beta-lactamase from Escherichia coli . These techniques included perturbation of the pK(a) of Lys-73 by the study of the gamma-thialysine-73 variant and the attendant kinetic analyses, investigation of the protonation state by titration of specifically labeled proteins by nuclear magnetic resonance, and by computational treatment using the thermodynamic integration method . All three methods indicated that the pK(a) of Lys-73 of this enzyme is attenuated to 8.0-8.5 . It is argued herein that the unique ground-state ion pair of Glu-166 and Lys-73 of class A beta-lactamases has actually raised the pK(a) of the active site lysine to 8.0-8.5 from that of the parental penicillin-binding protein . Whereas we cannot rule out that Glu-166 might activate the active site water, which in turn promotes Ser-70 for the acylation event, such as proposed earlier, we would like to propose as a plausible alternative for the acylation step the possibility that the ion pair would reconfigure to the protonated Glu-166 and unprotonated Lys-73 . As such, unprotonated Lys-73 could promote serine for acylation, a process that should be shared among all active-site serine beta-lactamases and penicillin-binding proteins.

Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi, 2003 Mar, 19(2), 132 - 5
{Construction and identification of the E.coli-BCG shuttle vector expressing lipoprotein Der p2 on cell wall of mycobacterium vaccae}; Shi JR et al.; AIM: To construct the E.coli-BCG shuttle vector carrying and expressing dust mite antigen gene Der p2 on cell wall of mycobacterium vaccae . METHODS: The gene fragment encoding 19 kDa antigen and the upstream control element (19-ss) was amplified by PCR from the mycobacteria tuberculosis H37Rv.Subsequently, the 19-ss gene was cloned into the E.coli-BCG shuttle vector pOLYG, which can schlepped and expressed exogenous antigen gene on cell wall of mycobacteria and containing the Der p2 gene . The expression of Der p2 gene in mycobacterium vaccae determined by indirect immunofluorescence staining . RESULTS: Sequencing proved that the cloned sequence of 19-ss gene was correct . The constructed E.coli-BCG shuttle vector (pCW) containing 19-ss gene had function of shuttle between E.coli and mycobacteria, and mediated the expression of antibiotic resistance gene . Indirect immunofluorescence staining indicated that the Der p2 gene was expressed in the form of lipoprotein on surface of the mycobacterium vaccae . CONCLUSION: E.coli-BCG shuttle vector has been constructed successfully, which can express exogenous antigen gene as a chimeric protein on cell wall.

J Gastroenterol Hepatol, 2004 Jun, 19(6), 626 - 31
Genetic and phenotype changes following in vitro interactions between Helicobacter pylori strains; Yakoob J et al.; BACKGROUND: The purpose of the present paper was to determine whether in vitro interaction between different Helicobacter pylori strains leads to changes in antibiotic susceptibility, cagA, vacAM2 and DNA fingerprint patterns . METHODS: Three H . pylori strains with known antibiotic susceptibility, cagA, vacAM2 status and polymerase chain reaction-random amplified polymorphic DNA (PCR-RAPD) fingerprint analysis were suspended in phosphate buffered saline (PBS pH 7.0), and the suspensions were mixed in equal proportion prior to culture on chocolate agar plates . Subcultures were performed five times every 3 days . As a control, each of the three strains was also subcultured separately . Antibiotic susceptibility testing, PCR for cagA, vacAM2 and PCR-RAPD analysis were done . RESULTS: Surviving strain of the two H . pylori strains in each combination demonstrated change in resistance to both antibiotics but no change in sensitivity . CagA status of the surviving strain varied as compared to the vacAM2 status, which did not change . The PCR-RAPD fingerprint showed unique band pattern . CONCLUSION: DNA transformation follows in vitro interaction . Helicobacter pylori strain with antibiotic resistance is likely to dominate in such in vitro interactions between various strains.

Microb Drug Resist, 2004 Spring, 10(1), 27 - 30
Integron-associated antibiotic resistance in enteroaggregative and enteroinvasive Escherichia coli; Gassama A et al.; Ten enteroinvasive (EIEC) and 25 enteroaggregative (EaggEC) E . coli strains isolated from Senegalese patients were analyzed for their integron content . All strains were resistant to at least two antibiotics . Four EIEC and 15 EaggEC were found to carry a class 1 integron . An identical integron carrying a single dfrA5 cassette, conferring resistance to trimethoprim, was identified in all four EIEC strains . Five EaggEC strains harbored an integron with a single cassette, dfrA7, while the remaining 10 strains carried two integrons, one with a single cassette, aadA1a conferring resistance to streptomycin and spectinomycin, and the second one bearing two cassettes, dfrA13 and oxa5, the later being a beta-lactam resistance cassette . The presence of these integrons is worrying, because trimethoprim is largely used for diarrheal disease therapy in Africa . Thus, the presence of integrons in diarrheagenic strains is of public health importance because a limited number of antibiotics are available in developing countries.

Chest, 2004 May, 125(5), 1913 - 9
Empiric therapy of community-acquired pneumonia: guidelines for the perplexed?
Cunha BA.
This article discusses the key clinical aspects of empiric therapy of community-acquired pneumonia (CAP) . Antibiotic selection, severity of CAP, single vs multiple pathogens, pharmacokinetic considerations, antibiotic resistance, i.v . vs oral antibiotic therapy for CAP, oral therapy for non-ICU hospitalized patients with CAP, beta-lactams, macrolides, ketolides, doxycycline, respiratory quinolones, and pharmacoeconomic implications are discussed.

Cas Lek Cesk, 2004, 143(3), 178 - 83
{Antibiotic resistance in invasive pneumococci and their serotypes in the Czech Republic}; Urbaskova P et al.; BACKGROUND: Among strains of S . pneumoniae (pneumococci), isolated from blood or cerebrospinal fluid of 483 patients between January 2001 and October 2003 as many as 5.2% strains were penicillin-non-susceptible . METHODS AND RESULTS: Incidence of penicillin-non-susceptible S . pneumoniae (PNSP) strains was highest in children of the youngest age group (12.7%) . Cefotaxim-non-susceptibility was detected in 2.7% of strains . Pneumococci tested were resistant to co-trimoxazol (9.9%), tetracycline (8.9%), chloramphenicol (4.6%), erythromycin (2.3%), clindamycin (1.4%), levofloxacin (0.6%) and rifampicin (0.2%); none of the strains was resistant to linezolid . Comparison of the results from 1996-1999 shows a permanent occurrence of invasive PNSP strains ranging about 5% . The PNSP strains were of 7 serotypes, with serotype 9V being the most frequent . The Czech collection of invasive pneumococcal strains is five times smaller than that of any other European country with comparable populations over the same period of time . CONCLUSIONS: Low incidence of invasive S . pneumoniae strains seems to be related to the underestimated significance of blood culturing . Valid data necessary for the safety of antibiotic therapy in terms of efficacy and prevention of resistance development cannot be obtained from some localities of the Czech Republic.

Southeast Asian J Trop Med Public Health, 2003 Sep, 34(3), 620 - 6
Antibiotic resistance, plasmid profile and RAPD-PCR analysis of enteropathogenic Escherichia coli (EPEC) clinical isolates; Wan KF et al.; Enteropathogenic Escherichia coli (EPEC) is a leading cause of diarrhea among infants in developing countries . A total of 38 EPEC isolates, obtained from diarrhea patients of Hospital Miri, Sarawak, were investigated through plasmid profile, antibiotic resistance and randomly amplified polymorphic DNA (RAPD) analysis . From the 8 types of antibiotics used, all isolates were 100% resistant to furoxime, cephalothin and sulphamethoxazole and showed high multiple antibiotic resistant (MAR) indexes, ranging from 0.5 to 1.0 . In plasmid profiling, 22 isolates (58%) showed the presence of one or more plasmids in the range 1.0 to 30.9 mDa . The dendrogram obtained from the results of the RAPD-PCR discriminated the isolates into 30 single isolates and 3 clusters at the level of 40% similarity . The EPEC isolates were highly diverse, as shown by their differing plasmid profiles, antibiotic resistance patterns and RAPD profiles.

Methods Mol Biol, 2004, 269, 15 - 30
Construction and isolation of recombinant vaccinia virus using genetic markers; Lorenzo MM et al.; The standard approach for the isolation of vaccinia virus recombinants involves homologous recombination between a transfected plasmid and the replicating viral DNA . In a typical infection/transfection experiment, recombinant viruses only account for a tiny proportion (10-4 to 10-3) of the progeny virus; thus, genetic markers are often included in the transfected plasmid to facilitate the selection of recombinant viruses . This chapter describes in detail two different selection procedures: one relies on plaque formation phenotype using the vaccinia virus gene F13L; the other relies on antibiotic resistance using the Escherichia coli xanthine-guanine phosphoribosyl transferase gene.

J Antimicrob Chemother, 2004 Jun, 53(6), 958 - 63 Epub 2004 Apr 21.
Enhancement of host fitness by the sul2-coding plasmid p9123 in the absence of selective pressure; Enne VI et al.; OBJECTIVES: Despite a 97% reduction in clinical sulphonamide usage, the prevalence of sulphonamide resistance among Escherichia coli has remained constant in the UK . Genetic linkage of sulphonamide resistance to other resistances is thought important for this maintenance, but the finding also implies that sulphonamide resistance exerts little fitness cost . To test this hypothesis, we examined the fitness impact of four naturally occurring sul2-coding plasmids upon their hosts . METHODS: The fitness impact of the plasmids upon E . coli was determined by pairwise growth competition in a minimal medium . The DNA sequence of plasmid p9123 was obtained by primer walking and PCR . RESULTS: Three of the four sul2-coding plasmids studied imposed fitness costs on their hosts . The fourth plasmid, a 6.2 kb resistance element carrying sul2, strA and strB designated p9123, conferred a 4% fitness advantage upon its original clinical host and also on E . coli K12 JM109 . The complete sequence of p9123 revealed eight open reading frames, including five of unknown function . There was no obvious gene to which the fitness advantage might be attributed . CONCLUSIONS: The novel finding that p9123 can improve host fitness may explain why this plasmid and its close relatives are so widespread among enteric bacteria . In addition to other factors such as co-selection of sulphonamide resistance by other agents, the fitness advantage conferred by plasmids such as p9123 may have contributed to the maintenance of sulphonamide resistance in the UK in the absence of clinical selection pressure . These data indicate that once antibiotic resistance has been established on mobile genetic elements, it may be difficult to eliminate.

J Clin Gastroenterol, 2004 Apr, 38(4), 326 - 31
A national survey of primary care physicians' perceptions and practices related to Helicobacter pylori infection; Sharma VK et al.; OBJECTIVE: Our aim was to assess current perceptions and practices of primary care physicians in the United States concerning Helicobacter pylori infection . METHODS: We mailed a structured questionnaire to approximately 2500 primary care physicians chosen at random from a national database . We asked about personal and practice demographics and practices relating to testing for and treating H . pylori infection . RESULTS: We received 424 responses from 2349 questionnaires (18%) . Only 3% each had used either the C- or C-urea breath test; 5% had used the stool antigen test; 92% and 91% recommended testing-and 90% and 82% treatment-for H . pylori in patients with active duodenal and gastric ulcer, respectively . However, only 64% would test for and only 59% would treat H . pylori infection in a patient with past history of duodenal ulcer . Almost half would test patients with gastroesophageal reflux disease being started or maintained on a proton pump inhibitor . The most frequent treatment regimens used were combinations of a proton pump inhibitor, clarithromycin, and either amoxicillin or metronidazole . Most respondents had inaccurate information on antibiotic resistance rates for H . pylori . In the absence of symptoms, 26% would personally undergo testing for H . pylori; 30% would be treated if infected . CONCLUSIONS: Primary care physicians usually test for and treat H . pylori infection in patients with active ulcer, but fail to do so in patients with a prior history of ulcer . Some test for H . pylori in gastroesophageal reflux disease patients . Most use efficacious treatment regimens, but have inaccurate information on resistance rates.

Pharm World Sci, 2004 Apr, 26(2), 90 - 5
Methodological validation of monitoring indicators of antibiotics use in hospitals; Mandy B et al.; BACKGROUND: For several years now, the French national recommendations have been trying to set up a surveillance system in hospitals to link data on antibiotic resistance and data on the use of antibiotics, particularly for certain 'micro-organism/antibiotic' pairs . The indicators recommended in the lastest newsletter of the Direction Generale de la Sante (French Public Health Department) for monitoring the consumption of antibiotics were the number of days of treatment or the number of defined daily doses (DDD), both (in)directly related to the number of days of hospitalisation and/or the number of patients hospitalised . OBJECTIVE: The aim of this study was to compare the actual number of days of treatment, which is an observed indicator, with two indicators calculated on the basis of the DDD and the DPD (daily prescribed dose), both in terms of feasibility of collection and the relevance of the information generated . MATERIALS AND METHODS: For several hospital care units, the 'length of exposure' to a given antibiotic was determined by four different indicators: two actual observed indicators {the patient's medical file (reference) and the named-patient based, computerised dispensing system from the central pharmacy} and two derived calculated indicators {obtained by dividing the number of grams prescribed by the DDD or by the DPD} . RESULTS: The average incidence density of antibiotic treatment (length of exposure per 1000 days of hospitalisation) obtained by the calculated indicators was higher than that obtained with the observed reference (+52% for the DDD and +33% for the DPD) but lower than that obtained with the second observed indicator (computerised system) (-10%) . The differences were large and random (high variability depending on the hospital department, the antibiotic and the administration route; variations in both directions: actual length of treatment longer or shorter than the calculated length of treatment) . CONCLUSION: The question which indicator should be chosen is inconclusive for the evaluation of the selection pressure exerted by an antibiotic . The two indicators proposed in the newsletter (observed indicator and calculated indicator) seem to be complementary for use in a regional or national network to monitor resistance and consumption of antibiotics . Each hospital should validate the indicators and define for itself which indicator is most appropriate for estimating the actual length of antibiotic exposure . This may imply different indicators for different units, antibiotics or even administration routes within one particular hospital setting . Once validated the hospital has a powerful tool generating data that can be linked to resistance data.

Curr Opin Pulm Med, 2004 May, 10(3), 166 - 70
Community-acquired pneumonia in casualty: etiology, clinical features, diagnosis, and management (or a look at the "new" in pneumonia since 2002); Alves DW et al.; PURPOSE OF REVIEW: Community-acquired pneumonia, because of its substantial treatment costs, incidence, and mortality, is an aggressively researched diagnosis . In this review, we highlight new developments in the diagnosis, etiology, pathophysiology, treatment, and prevention of community-acquired pneumonia published since April 2002 . RECENT FINDINGS: The combined end points of improved patient care and conservation of health care resources have prompted several studies examining current professional society community-acquired pneumonia guidelines . In general, patients treated with the recommended third-generation cephalosporin and macrolide or an antipneumococcal fluoroquinolone when indicated have fared better, including reduced overall costs, inpatient days, and mortality, than those receiving alternative treatments . Etiologic identification efforts by traditional methods, blood and sputum cultures, are being questioned owing to poor success rates and, even when positive, are being underused or ignored in antibiotic selection and patient management . Newer diagnostic tests are becoming commercially available, along with tests for biologic markers that have been only recently identified as contributors to, or prognosticators of, community-acquired pneumonia . Because antibiotic resistance remains a major obstacle to successful patient treatment, prevention or mitigation of community-acquired pneumonia is gaining increasing popularity through more aggressive pneumococcal and influenza vaccination of at risk groups, even before hospital discharge from a community-acquired pneumonia admission . SUMMARY: Although prevention is our best defense, current community-acquired pneumonia treatment guidelines are effective for treatment and cost containment . However, they should be scrutinized in light of clinical utilization data now entering the literature regarding their testing recommendations . Providers should consider encouraging focused culturing of sicker patients and those with significant comorbidities.

Vaccine, 2004 Apr 16, 22(13-14), 1709 - 16
DNA immunisation with minimalistic expression constructs; Moreno S et al.; The low efficacy obtained in large animals makes plasmid-based DNA vaccines commercially unviable . Another concern is the presence of antibiotic resistance markers on virtually all conventional plasmids . Here we describe the use of minimalistic, immunogenically defined gene expression (MIDGE) vectors for DNA vaccination . MIDGE are linear, covalently-closed vectors containing all the essential information for gene expression and none of the non-essential and potentially dangerous plasmid backbone sequences . MIDGE vectors can also be chemically modified on both ends at defined positions allowing targeting of the DNA to specific cell types or cellular compartments . Immunisation of mice with simple and end-modified MIDGE vectors showed that they are efficacious tools to generate and/or manipulate antigen-specific immune responses.

FEBS Lett, 2004 Apr 9, 563(1-3), 7 - 12
Resolving the evolutionary paradox of genetic instability: a cost-benefit analysis of DNA repair in changing environments; Breivik J et al.; Loss of genetic stability is a critical phenomenon in cancer and antibiotic resistance, and the prevailing dogma is that unstable cells survive because instability provides adaptive mutations . Challenging this view, we have argued that genetic instability arises because DNA repair may be a counterproductive strategy in mutagenic environments . This paradoxical relationship has also been confirmed by explicit experiments, but the underlying evolutionary principles remain controversial . This paper aims to clarify the issue, and presents a model that explains genetic instability from the basic perspective of molecular evolution and information processing.

FEMS Microbiol Lett, 2004 Apr 15, 233(2), 201 - 4
Genetic transformation and mutant isolation in Ganoderma lucidum by restriction enzyme-mediated integration; Kim S et al.; A white-rot basidiomycete Ganoderma lucidum has long been used as a medicinal mushroom in Asia, and it has an array of enzymes important for wood degrading activity . There have been many reports about the ingredients which show health aiding effects . In order to analyze gene functions and introduce foreign genes into this fungus, genetic transformation is required . We have successfully transformed G . lucidum to geneticin resistance using pJS205-1 which has the antibiotic resistance genes against geneticin and phosphinothricin . Many different mutants have been generated during the transformation by restriction enzyme mediated integration, and the transformation yield was 4-17 transformants (microg plasmid DNA)(-1) . The plasmid was integrated stably into the recipient chromosome, which was confirmed by PCR with the plasmid-specific primers.

Otolaryngol Head Neck Surg, 2004 Mar, 130(3 Suppl), S89 - 94
The development of antibiotic resistant organisms with the use of ototopical medications; Weber PC et al.; OBJECTIVE: There is growing concern over the use of systemic antibiotics and the development of bacterial resistance . The question remains as to whether ototopical medications may also promote antibiotic-resistant organisms, either on a local level (in the ear) or in other areas of the aerodigestive tract . We performed an evidence-based review to answer the following clinical question, "Do antibiotic ototopical medications induce antibiotic resistant organisms?" STUDY DESIGN: We performed a MEDLINE search of the published literature from 1966 to the present . We used appropriate search terms such as "ototopical antibiotics," "ototopical drops," "antibiotic resistance," "topical antibiotics and otitis externa," "otitis externa and treatment," "otitis externa and antibiotic drops," "otitis externa and ototopical drops," "otitis media," "otitis media and treatment," "otitis media and antibiotic drops," "chronic suppurative otitis media," "chronic suppurative otitis media and treatment," "chronic suppurative otitis media and antibiotic drops," " otitis externa and resistant organisms," "otitis media and resistant organisms," "chronic suppurative otitis media and resistant organisms," "ophthalmic antibiotic drops," "draining ear," "P.E . tube otorrhea," "pressure equalizing tube otorrhea," "pressure equalizing tube otorrhea and treatment," and "pressure equalizing tube otorrhea and ototopical therapy" to identify pertinent articles . These articles were reviewed and graded according to the evidence quality . RESULTS: After an initial screening of over 2,500 articles, 38 articles were analyzed further; of these, 11 were determined to warrant extensive review . Eight articles evaluated chronic suppurative otitis media; 2, otitis externa; and 1, post-tympanostomy tube otorrhea, whereas 3 others studied systemic absorption . Of the 8 chronic suppurative otitis media studies, there were thought to be 5 grade 2B studies, 1 grade 1B study, and 1 grade 2C study . These studies did not demonstrate a propensity for the development of resistant organisms . No study answered the question as to whether resistance to systemic antibiotics might occur in otitis externa . CONCLUSIONS: Overall grade B evidence seems to indicate that no significant antibiotic resistance develops from the use of ototopical antibiotic treatment.

Otolaryngol Head Neck Surg, 2004 Mar, 130(3 Suppl), S79 - 82
Ototoxicity of ototopical antibiotic drops in humans; Matz G et al.; OBJECTIVE: We sought to determine the incidence of injury to the cochlea or vestibular system from the use of ototopic antibiotic ear drops . STUDY DESIGN: We conducted a MEDLINE search of the published literature from 1966 to the present using the search items of "ototopic antibiotics," "ototopic drops," "antibiotic resistance," and others to identify pertinent articles . These articles were reviewed and graded according to evidence quality . Forty-four articles were found; 27 articles were considered appropriate for review and 14 of these articles were thought to warrant further extensive review . The latter 14 articles dealt directly with the evaluation of either hearing loss or changes in vestibular function after the use of ototopic ear drops . We eliminated studies that did not consider hearing or vestibular function . We considered only articles in the English language, eliminating articles that not address the ototoxicity of ototopical ear drops . RESULTS: Most of these articles were in the level 3 to 3b category . Two of the articles were in the 1b category, evaluating evidence-based studies . An article that reviewed case reports of ototopical ototoxicity and a survey article are included in this study, as case reports of ototopical ototoxicity and survey articles . CONCLUSION: We found a total of 54 cases of gentamicin vestibular toxicity and, in 24 of these patients, cochlear toxicity was also documented . Our review also found 11 cases of cochlear and 2 cases of vestibular toxicity in neomycin-based ear drops.

Can J Microbiol, 2004 Feb, 50(2), 69 - 77
Biosensors for the detection of bacteria; Deisingh AK et al.; This review will consider the role of biosensors towards the detection of infectious bacteria, although non-infectious ones will be considered where necessary . Recently, there has been a heightened interest in developing rapid and reliable methods of detection . This is especially true for detection of organisms involved in bioterrorism, food poisoning, and clinical problems such as antibiotic resistance . Biosensors can assist in achieving these goals, and sensors using several of the different types of transduction modes are discussed: electrochemical, high frequency (surface acoustic wave), and optical . The paper concludes with a discussion of three areas that may make a great impact in the next few years: integrated (lab-on-a-chip) systems, molecular beacons, and aptamers.

J Virol, 2004 Apr, 78(8), 4314 - 22
Rubella virus capsid protein modulates viral genome replication and virus infectivity; Chen MH et al.; The structural proteins (SP) of the Togaviridae can be deleted in defective interfering RNAs . The dispensability of viral SP has allowed construction of noninfectious viral expression vectors and replicons from viruses of the Alphavirus and Rubivirus genera . Nevertheless, in this study, we found that the SP of rubella virus (RUB) could enhance expression of reporter genes from RUB replicons in trans . SP enhancement required capsid protein (CP) expression and was not due to RNA-RNA recombination . Accumulation of minus- and plus-strand RNAs from replicons was observed in the presence of SP, suggesting that SP specifically affects RNA synthesis . By using replicons containing an antibiotic resistance gene, we found 2- to 50-fold increases in the number of cells surviving selection in the presence of SP . The increases depended significantly on the amount of transfected RNA . Small amounts of RNA or templates that replicated inefficiently showed more enhancement . The infectivity of infectious RNA was increased by at least 10-fold in cells expressing CP . Moreover, virus infectivity was greatly enhanced in such cells . In other cells that expressed higher levels of CP, RNA replication of replicons was inhibited . Thus, depending on conditions, CP can markedly enhance or inhibit RUB RNA replication.

J Intensive Care Med, 2003 Jul-Aug, 18(4), 175 - 88
Nosocomial pneumonia in the intensive care unit: controversies and dilemmas; Mehta RM et al.; Nosocomial pneumonia (NP), and its most serious form, ventilator-associated pneumonia (VAP), is a major cause of morbidity and mortality in the ICU . Numerous controversies exist, from diagnostic criteria to prevention and treatment, including the issues of attributable mortality of VAP, differences in the approach to early and late VAP, and the best diagnostic methods . Initial, accurate therapy is one of the most important factors determining outcome in VAP . Antibiotic monotherapy versus combination therapy is not clearly defined, as clinicians struggle with the dual risk of inadequate therapy negatively affecting outcome and overtreatment promoting antibiotic resistance . The role of airway and gastrointestinal colonization and innovative preventive strategies such as noninvasive ventilation, antibiotic rotation, and aerosolized antibiotics are discussed . No uniform standards exist for the approach to VAP . The authors highlight the major controversies and dilemmas in the clinical approach to VAP, with recommendations for the bedside management of these patients.

J Food Prot, 2004 Mar, 67(3), 486 - 92
Characterization of colicinogenic Escherichia coli strains inhibitory to enterohemorrhagic Escherichia coli; Schamberger GP et al.; A previously identified set of anti-Escherichia coli O157:H7 colicinogenic E . coli were characterized to assess the suitability of these isolates as a preharvest food safety intervention in cattle . This collection of 23 E . coli strains were screened for virulence factors, antibiotic resistance, type of colicin(s) present, and their ability to inhibit other pathogenic E . coli . With the use of PCR, pathogen genes were detected in six of the 23 colicinogenic E . coli . When the nonpathogenic strains were assessed for antibiotic resistance, four strains showed resistance to at least one antibiotic . The remaining set of 14 strains were evaluated for the presence of previously identified colicins . Seven colicins (B, El, E2/E7, E7, Ia/Ib, K, and M) were detected . One half of the strains possessed multiple types of colicins . The most commonly detected colicins were B, E2/E7, and M, which were found in six strains each . DNA sequencing was also performed in order to classify the E2/E7 colicins separately from E7 colicins . The 14 colicinogenic E . coli also were evaluated for their ability to inhibit 10 different non-O157 pathogenic E . coli . Six of the colicinogenic E . coli were capable of inhibiting all 10 pathogens, and the remaining eight strains could each inhibit between six to eight of the pathogenic E . coli . This strain collection has great potential for inhibiting E . coli O157:H7 in cattle.

Nat Rev Microbiol, 2004 Jan, 2(1), 73 - 8
Can better prescribing turn the tide of resistance?
Livermore D.
In the wake of concerns about the level of antibiotic resistance, governments worldwide are pressing for reduced antibiotic use, hoping thereby to reverse resistance trends . Is success likely? The evidence is mixed, and expectations should be tempered by the growing realization that many resistant bacteria are biologically fit, making them difficult to displace . If resistance is unlikely to be reduced significantly by changing prescription practices, how can clinicians outpace increased resistance, particularly when much of 'big pharma' is abandoning antibiotic development?

Asia Pac J Clin Nutr . 2003;12 Suppl:S58.
Can nutritional manipulation enhance immune competence in broiler chickens?
Digby SN, Revell DK, Hughes BJ.
Background - Antibiotic use for disease control in animal production systems, such as the chicken meat broiler industry, has been implicated in the development of antibiotic resistance in humans . As a result, there is increasing interest in the manipulation of nutrition and the development of new vaccination programs to enhance immune competence . Objective - To determine whether nutrition could be used to enhance immune competence through supplementation of broiler diets with vitamins A, C and E and the minerals zinc, copper and selenium . Design - To vary the extent of challenge to the immune system, two different housing environments (optimal and suboptimal) were used . Two diets were fed in each environment (nutritionally enriched vs control) and half of the birds within each diet group and environment were vaccinated with a live vaccine for coccidiosis (containing various Eimeria strains) whilst the other half were not . The trial was conducted over a six-week period during which time body weight, weekly feed intake and indicators of immune competence (white blood cell counts and bursa and thymus weights) were measured . Outcomes - Results showed increased bursa and thymus weights in chickens in the suboptimal environment suggesting the level of infection was greater . Eosinophil counts were significantly greater (P<0.05) in birds fed the enriched diet and in vaccinated birds, suggesting an enhanced response to the coccidiosis parasites . Conclusions - These results are promising for the use of dietary manipulation as an alternative to antibiotic treatment in the future.

Mol Microbiol, 2004 Mar, 51(6), 1801 - 16
Proteolytic degradation of Escherichia coli transcription activators SoxS and MarA as the mechanism for reversing the induction of the superoxide (SoxRS) and multiple antibiotic resistance (Mar) regulons; Griffith KL et al.; In Escherichia coli, the SoxRS regulon confers resistance to redox-cycling compounds, and the Mar regulon provides a defence against multiple antibiotics . The response regulators, SoxS and MarA, are synthesized de novo in response to their inducing signals and directly activate transcription of a common set of target genes . Although the mechanisms of transcription activation by SoxS and MarA have been well studied, little is known about how the systems are shut-off once the inducing stress has subsided, except that de novo synthesis of the regulators is known to cease almost immediately . Here, we induced the SoxRS regulon and determined that, upon removal of the inducer, expression of the regulon's genes quickly returns to the preinduced level . This rapid shut-off indicates that the system is reset by an active process . We found that SoxS is unstable and infer that SoxS degradation is responsible for the rapid return of the system to the ground state upon removal of the inducing signal . We also found that MarA is unstable and that the instability of both proteins is intrinsic and unregulated . We used null mutations of protease genes to identify the proteases involved in the degradation of SoxS and MarA . Among single protease mutations, only lon mutations increased the half-life of SoxS and MarA . In addition, SoxS appeared to be nearly completely stable in a lon ftsH double mutant . Using hexahistidine tags placed at the respective ends of the activators, we found that access to the amino-terminus is essential for the proteolytic degradation.

Infect Dis Clin North Am, 2003 Dec, 17(4), 773 - 84
Prevention of hospital-acquired pneumonia: European perspective; Bonten MJ; Several preventive measures for VAP have been empirically tested . There is clear evidence that antibiotic-containing preventive strategies, such as SDD and oropharyngeal decontamination, are very effective in different patient populations . Selection of antibiotic resistance remains the major disadvantage of these strategies, however, limiting its applicability in settings with high levels of antibiotic resistance . This probably precludes the use of these strategies in many American settings, but may allow their use in European countries with much lower endemic levels of resistance . There is little evidence that systemic prophylaxis is effective for the prevention of VAP, and initial studies were associated with resistance problems . Of the non antibiotic-containing preventive strategies, subglottic aspiration was effective in several studies, whereas other strategies, such as immunonutrition with glutamine or the semirecumbent patient position, were effective in single studies . All these studies were executed in European ICUs . For these interventions, more data are needed on the generalizability, feasibility, and cost effectiveness . Few data support the use of sucralfate for stress ulcer prophylaxis and modulation of enteral nutrition practices as preventive measures for VAP.

Anaesthesist, 2004 May, 53(5), 427 - 33
{Quality assurance in intensive care medicine . SARI-surveillance on antibiotic use and bacterial resistance in intensive care units}; Meyer E et al.; Intensive care units (ICUs) are high risk areas for emergence and spread of multiresistant bacterial pathogens . In Germany, there are no representative epidemiological data on antibiotic resistance, prophylactic or therapeutic use of antibiotics in ICUs, or on the correlation between antibiotic use and emergence of resistance . Supported by the German Ministry of Science and Education, project SARI (Surveillance on antibiotic use and bacterial resistance in ICUs) started in 02/2000 and now includes data on antibiotic use and resistance rates in 35 medical, surgical and interdisciplinary ICUs . To date (2/2000-12/2002), a total of 939 participant months, 339,461 patient days and 452,282 defined daily doses (DDD) have been covered with a mean antibiotic usage density (AD) of 1,332 DDDs/1,000 patient days and resistance data on 31,189 isolates from ICUs . The design of the project and first results of SARI are presented . The epidemiological data of SARI form a basis for improved antibiotic and infection control management in ICUs .

Orthopade, 2004 Mar, 33(3), 316 - 26
{Postoperative and bacterial osteitis . New possibilities for therapy}; Heppert V et al.; Current therapy concepts for post-trauma osteitis include radical debridement, stabilization of bone and soft tissue care using early flap transfers . With this more radical approach, the long term results are improved, however, total treatment time still is very long . In addition to the physical, mental and social consequences for the patient, treatment costs are also very high . Antibiotic therapy as an adjuvant is still standard and effective although the development of worldwide antibiotic resistance has increased treatment difficulties . One of the main aims of research is to reduce these problems or to prevent the development of osteitis entirely . However, research has not yet elucidated the complex immunologic changes involved . In addition, most results are obtained from animal studies and clinical investigations are required.

Proc Natl Acad Sci U S A, 2004 Mar 16, 101(11), 3786 - 91 Epub 2004 Mar 04.
Isolation of antibiotic resistance mutations in the rRNA by using an in vitro selection system; Cochella L et al.; Genetic, biochemical, and structural data support an essential role for the ribosomal RNA in all steps of the translation process . Although in vivo genetic selection techniques have been used to identify mutations in the rRNAs that result in various miscoding phenotypes and resistance to known ribosome-targeted antibiotics, these are limited because the resulting mutant ribosomes must be only marginally disabled if they are able to support growth of the cell . Furthermore, in vivo, it is not possible to control the environment in precise ways that might allow for the isolation of certain types of rRNA variants . To overcome these limitations, we have developed an in vitro selection system for the isolation of functionally competent ribosomal particles from populations containing variant rRNAs . Here, we describe this system and present an example of its application to the selection of antibiotic resistance mutations . From a pool of 4,096 23S rRNA variants, a double mutant (A2058U/A2062G) was isolated after iteration of the selection process . This mutant was highly resistant to clindamycin in in vitro translation reactions and yet was not viable in Escherichia coli . These data establish that this system has the potential to identify mutations in the rRNA not readily accessed by comparable in vivo systems, thus allowing for more exhaustive ribosomal genetic screens.

Plant J, 2004 Mar, 37(6), 906 - 13
A novel approach to plastid transformation utilizes the phiC31 phage integrase; Lutz KA et al.; Thus far plastid transformation in higher plants has been based on incorporation of foreign DNA in the plastid genome by the plastid's homologous recombination machinery . We report here an alternative approach that relies on integration of foreign DNA by the phiC31 phage site-specific integrase (INT) mediating recombination between bacterial and phage attachment sites (attB and attP, respectively) . Plastid transformation by the new approach depends on the availability of a recipient line in which an attB site has been incorporated in the plastid genome by homologous recombination . Plastid transformation involves insertion of an attP vector into the attB site by INT and selection of transplastomic clones by selection for antibiotic resistance carried in the attP plastid vector . INT function was provided by either expression from a nuclear gene, which encoded a plastid-targeted INT, or expressing INT transiently from a non-integrating plasmid in plastids . Transformation was successful with both approaches using attP vectors with kanamycin resistance or spectinomycin resistance as the selective marker . Transformation efficiency in some of the stable nuclear INT lines was as high as 17 independently transformed lines per bombarded sample . As this system does not rely on the plastid's homologous recombination machinery, we expect that INT-based vectors will make plastid transformation a routine in species in which homologous recombination rarely yields transplastomic clones.

Proc Natl Acad Sci U S A, 2004 Mar 9, 101(10), 3709 - 14 Epub 2004 Feb 25.
Persistent colonization and the spread of antibiotic resistance in nosocomial pathogens: resistance is a regional problem; Smith DL et al.; Infections with antibiotic-resistant bacteria (ARB) in hospitalized patients are becoming increasingly frequent despite extensive infection-control efforts . Infections with ARB are most common in the intensive care units of tertiary-care hospitals, but the underlying cause of the increases may be a steady increase in the number of asymptomatic carriers entering hospitals . Carriers may shed ARB for years but remain undetected, transmitting ARB to others as they move among hospitals, long-term care facilities, and the community . We apply structured population models to explore the dynamics of ARB, addressing the following questions: (i) What is the relationship between the proportion of carriers admitted to a hospital, transmission, and the risk of infection with ARB? (ii) How do frequently hospitalized patients contribute to epidemics of ARB? (iii) How do transmission in the community, long-term care facilities, and hospitals interact to determine the proportion of the population that is carrying ARB? We offer an explanation for why ARB epidemics have fast and slow phases and why resistance may continue to increase despite infection-control efforts . To successfully manage ARB at tertiary-care hospitals, regional coordination of infection control may be necessary, including tracking asymptomatic carriers through health-care systems.

Tidsskr Nor Laegeforen, 2004 Feb 19, 124(4), 484 - 7
{Treatment of serious infectious diseases in children}; Krogvold L et al.; BACKGROUND: Serious systemic infections represent a major challenge to all paediatric departments, hence the importance of having treatment guidelines available . MATERIALS AND METHODS: Based on a review of the literature and a meeting of Norwegian paediatricians with an interest in the field, guidelines for the treatment of serious infectious diseases are proposed . RESULTS: The two main new proposals include once daily dosage of aminoglycosides and increased dosage of aciclovir in the treatment of herpes simplex encephalitis . INTERPRETATION: The main objective of the meeting was to maintain a conservative and environmental-friendly antibiotics policy in order to contribute to the prevention of antibiotic resistance.

FEMS Microbiol Rev, 2004 Feb, 28(1), 79 - 100
A classification scheme for mobilization regions of bacterial plasmids; Francia MV et al.; Transmissible plasmids can be classified according to their mobilization ability, as being conjugative (self-transmissible) or mobilizable (transmissible only in the presence of additional conjugative functions) . Naturally occurring mobilizable plasmids carry the genetic information necessary for relaxosome formation and processing, but lack the functions required for mating pair formation . Mobilizable plasmids have a tremendous impact in horizontal gene transfer in nature, including the spread of antibiotic resistance . However, analysis of their promiscuity and diversity has attracted less attention than that of conjugative plasmids . This review will focus on the analysis of the diversity of mobilizable plasmids . For this purpose, we primarily compared the amino acid sequences of their relaxases and, when pertinent, we compared these enzymes with conjugative plasmid relaxases . In this way, we established phylogenetic relationships among the members of each superfamily . We conducted a database and literature analysis that led us to propose a classification system for small mobilizable plasmids in families and superfamilies according to their mobilization regions . This review outlines the genetic organization of each family of mobilization regions, as well as the most relevant properties and relationships among their constituent encoded proteins . In this respect, the present review constitutes a first approach to the characterization of the global gene pool of mobilization regions of small mobilizable plasmids.

Andrologia, 2004 Feb, 36(1), 1 - 23
Update on the impact of Chlamydia trachomatis infection on male fertility; Gonzales GF et al.; With approximately 90 million cases annually, infection with Chlamydia trachomatis is the most prevalent sexually transmitted bacterial disease in the world . Considering that these infections are often asymptomatic and cause major complications like acute pelvic inflammatory disease, ectopic pregnancy, infertility or infant pneumonia, the estimated costs for diagnosis and treatment in the USA amounts to 2.2 million US dollars for each 500 cases . Therefore, there is a high need for correct, quick and cost-effective diagnosis and treatment of this urogenital tract infection . New innovative therapies provide good results with regard to efficacy and patients' compliance . The success rates of treatments are at least 95% . However, the occurrence of antibiotic resistance should not be ignored and new treatment schemes must be developed . The state-of-the-art of diagnosis and treatment of chlamydial infections as well as the pathophysiology is discussed in this review . In conclusion, infections with C . trachomatis is an important public health problem, especially in third world and developing countries, and more socio-economic studies linking secondary prevention of chlamydial infections, infertility and adverse pregnancy outcome are needed to understand more of its aetiology . In addition, diagnosis and treatment should be improved . Data in men revealed that past infections but not present infections are more related to male infertility . There is still controversial results . In future studies, function of the seminal vesicles and evaluation of the antioxidant capacity should be taken into account when role of C . trachomatis infection on male fertility is assessed.

Med J Malaysia, 2003 Aug, 58(3), 465 - 9
Reevaluating the use of antibiotics in acute otitis media in children; Elango S; Increasing concern about the antibiotic resistance in acute otitis media (AOM) has led to debate over use of antibiotic in AOM and duration of therapy . Many studies have proved that watchful waiting should be used more often for acute otitis media . In children over two years, the most appropriate treatment was found to be initial observation followed by 5 days of an antibiotic if the child failed to improve spontaneously . In children less than 2 years or one with severe symptoms antibiotic can be started after 24 hours if there is no improvement with symptomatic treatment . Physician should be more selective in the prescription of antibiotics early in AOM.

Eur J Clin Pharmacol, 2004 Mar, 60(1), 37 - 43 Epub 2004 Jan 29.
Pharmacological treatment of acute otitis media in children . A comparison among seven locations: Tenerife, Barcelona and Valencia (Spain), Toulouse (France), Smolensk (Russia), Bratislava (Slovakia) and Sofia (Bulgaria); Sanz E et al.; OBJECTIVE: To describe patterns observed in the treatment of acute otitis media (AOM) in several locations of five countries . PATIENTS AND METHODS: Cross-sectional, descriptive study . Random sample of 12,264 paediatric outpatients seen by paediatricians or general practitioners (GPs) . Data on patient demographics, diagnoses and treatment were collected . Diagnoses were coded by ICD-9 and drugs by ATC classification . Patients diagnosed with AOM (ICD-9 codes: 381 and 382) were selected for analysis . RESULTS: Cases of AOM (873) accounted for 7.1% of the sample . There is a clear variation in the percentage of children diagnosed with AOM and treated with antibiotics in the different locations, antibiotic prescriptions being higher in Barcelona (93% of children), and lowest in Smolensk (56.4 % of children were treated without antibiotics) . The antibiotics used varied widely: ampicillin use is almost limited to Smolensk (26.7%) and Bratislava (13.8%), whereas amoxicillin plus clavulanic acid is the choice in Toulouse (33.8%), Valencia (30.2%) and Barcelona (28.9%), and cephalosporins are more frequently prescribed in Tenerife (51.7%) . Finally, macrolides are used in Barcelona (18.3%), Valencia (17.5%) and Tenerife (13.6%), but not prescribed in Toulouse or Sofia . Prescriptions of anti-inflammatory drugs were only relevant in Valencia (31.7%), Tenerife (27.2%) and Toulouse (17.4%) and of otological preparations in Sofia, where almost each child received ear drops (91.9%) . Nasal preparations are commonly used only in Sofia (41.9%), Bratislava (65.5%) and Smolensk (68.6%) . CONCLUSION: Despite the general agreement of most guidelines, wide differences in the treatment of uncomplicated AOM in children are observed . Non-antibiotic therapy for AOM and the use of first-choice antibiotics should be more actively encouraged in the primary care centres . Studies to measure prevailing rates of antibiotic resistance in these populations are needed.

Plant Cell Rep, 2004 Apr, 22(9), 660 - 7 Epub 2004 Jan 29.
Green fluorescent protein as a visual selection marker for papaya (Carica papaya L.) transformation; Zhu YJ et al.; Chemical-based selection for plant transformation is associated with a number of real and perceived problems that might be avoided through visual selection . We have used green fluorescent protein (GFP), as a visual selectable marker to produce transformed papaya ( Carica papaya) plants following microprojectile bombardment of embryogenic callus . GFP selection reduced the selection time from 3 months on a geneticin (G418) antibiotic-containing medium to 3-4 weeks . Moreover, GFP selection increased the number of transformed papaya plants by five-to eightfold compared to selection in the presence of antibiotics . Overall, the use of GFP for selecting transgenic papaya lines improved our throughput for transformation by 15- to 24-fold while avoiding the drawbacks associated with the use of antibiotic resistance-based selection markers.

J Antimicrob Chemother, 2004 Mar, 53(3), 432 - 4 Epub 2004 Jan 22.
Overexpression of the multidrug efflux pump SmeDEF impairs Stenotrophomonas maltophilia physiology; Alonso A et al.; OBJECTIVES: The use of antibiotics for the treatment of infectious diseases has led to important changes in the structure of pathogenic bacterial populations . However, these changes could be buffered if the expression of antibiotic resistance genes were to lead to the counter-selection of antibiotic-resistant strains in antibiotic-free environments . To test the effect of antibiotic resistance on bacterial fitness, we analysed the effect of the overproduction of the multidrug efflux pump SmeDEF on the physiology of Stenotrophomonas maltophilia . SmeDEF confers resistance to antibiotics belonging to different structural families, and its overexpression is associated with an antibiotic resistance phenotype in clinical isolates of S . maltophilia . RESULTS: Two S . maltophilia isogenic strains were analysed: the wild-type strain D457 and strain D457R, which is a SmeDEF overproducer . In co-culture experiments, under non-selective pressure the wild-type strain displaced the mutant strain D457R . Metabolic profiling showed that SmeDEF overproduction leads to several changes in S . maltophilia metabolism . Using a Dictyostelium discoideum model of bacterial virulence, we found overexpression of SmeDEF to be associated with a reduction in S . maltophilia virulence . CONCLUSIONS: Together, these data indicate that overexpression of the multidrug efflux pump SmeDEF causes a metabolic burden for S . maltophilia.

Ann Trop Paediatr, 2003 Dec, 23(4), 233 - 53
Antibiotic treatment for bacterial meningitis in children in developing countries; Fuller DG et al.; Bacterial meningitis causes 125,000 deaths each year in infants and young children and 96% of these occur in less developed countries where up to 50% of children with this disease die and 25-50% of survivors have neurological sequelae . Although 3rd-generation cephalosporins are optimal empirical therapy for bacterial meningitis, they are unaffordable in many developing countries . The majority of children worldwide are currently treated with cheaper alternatives . This paper reviews the challenges facing clinicians treating bacterial meningitis in developing countries, highlighting the problem of changing patterns of antibiotic resistance . In particular, it details the evidence for the use of chloramphenicol and 3rd-generation cephalosporins.

Plant Cell Rep, 2004 May, 22(10), 747 - 51 Epub 2004 Jan 21.
Dicistronic expression of the green fluorescent protein and antibiotic resistance genes in the plastid for selection and tracking of plastid-transformed cells in tobacco; Jeong SW et al.; A plastid transformation vector was constructed for dicistronic expression of the aminoglycoside 3'-adenyltransferase (aadA) and green fluorescent protein (gfp) genes under the control of the plastid rrn promoter . Gold particles coated with the vector DNA were bombarded onto tobacco leaf explants using a particle delivery system . Leaf explants produced adventitious shoots when cultured on shoot-inducing medium containing 500 mg l(-1) spectinomycin . Shoots that exhibited green fluorescence under UV light were selected . Southern blot analysis detected the presence of the aadA and gfp genes between trnA and trnI in the plastid genome . Northern blot analysis revealed that the aadA and gfp genes were both properly transcribed into a dicistronic transcriptional unit . The expression of the gfp gene in the plastid enabled separation of transformed chloroplasts from wild-type chloroplasts in the protoplast under a fluorescent microscope . The overall results indicate that dicistronic expression of the aadA and gfp genes in the plastid simplifies gene manipulation, facilitating selection and tracking of plastid-transformed cells .

J Antimicrob Chemother, 2004 Feb, 53(2), 371 - 4 Epub 2004 Jan 16.
BRO beta-lactamase alleles, antibiotic resistance and a test of the BRO-1 selective replacement hypothesis in Moraxella catarrhalis; Levy F et al.; OBJECTIVES: The hypothesis that BRO-1 selectively replaced the BRO-2 isoform of the Moraxella catarrhalis BRO beta-lactamase was tested by examining the temporal distribution, antibiotic resistance and epidemiological characteristics of isolates from a long-term collection at a single locale . METHODS: A rapid, one-step PCR assay conducted on 354 isolates spanning 1984-1994 distinguished bro alleles in over 97% of the beta-lactamase-producing isolates . Probes of dot blots were used to distinguish PCR failure from non-beta-lactamase-mediated penicillin resistance . RESULTS: BRO-2 isolates comprised 0-10% of the population per year with no evidence of a decline over time . All beta-lactamase producers exceeded the clinical threshold for penicillin resistance . Bimodality of penicillin MICs for beta-lactamase producers was caused by variation within BRO-1 rather than differences between BRO-1 and BRO-2 . Non-beta-lactamase factors also confer resistance to penicillin and may contribute to the BRO-1 bimodality . The 13 BRO-2 isolates were associated with diverse genotypes within which there was evidence of epidemiologically linked clusters . The exclusive association of BRO-2 with four unrelated genotypes suggested maintenance of BRO-2 by recurrent mutation or horizontal exchange . CONCLUSIONS: The relative rarity of BRO-2 throughout the study, the absence of a declining temporal trend, and genetic diversity within BRO-2 all failed to support the hypothesis that BRO-2 was more common in the past and has been selectively replaced by BRO-1.

AMIA Annu Symp Proc . 2003;:410-4.
A framework for infection control surveillance using association rules; Ma L et al.; Surveillance of antibiotic resistance and nosocomial infections is one of the most important functions of a hospital infection control program . We employed the association rule method for automatically identifying new, unexpected, and potentially interesting patterns in hospital infection control . We hypothesized that mining for low-support, low-confidence rules would detect unexpected outbreaks caused by a small number of cases . To build a framework, we preprocessed the data and added new templates to eliminate uninteresting patterns . We applied our method to the culture data collected over 3 months from 10 hospitals in the UPMC Health System . We found that the new process and system are efficient and effective in identifying new, unexpected, and potentially interesting patterns in surveillance data . The clinical relevance and utility of this process await the results of prospective studies.

Lancet Infect Dis, 2004 Jan, 4(1), 44 - 53
Educational interventions to improve antibiotic use in the community: report from the International Forum on Antibiotic Resistance (IFAR) colloquium, 2002; Finch RG et al.; National and international strategies for the control of antibiotic resistance recommend education for health-care professionals and the public to promote prudent antibiotic use . This paper, based on discussions at the 2002 colloquium of the International Forum on Antibiotic Resistance (IFAR), provides an international discourse between theoretical approaches to behaviour change and practical experience gained in large-scale antibiotic use educational campaigns . Interventions are more likely to be effective if their aim is to change behaviour, rather than provide information . They should target all relevant groups, especially parents, children, day-care staff, and health-care professionals . They should use clear and consistent messages concerning bacterial versus viral infection, prudent antibiotic use, symptomatic treatment, and infection-control measures (eg, handwashing) . Campaigns should use a range of communications using pilot-testing, strong branding, and sociocultural adaptation . Prime-time television is likely to be the most effective public medium, while academic detailing is especially useful for health-care professionals . Multifaceted interventions can improve antibiotic prescribing to some degree . However, there are few data on their effects on resistance patterns and patient outcomes, and on their cost-effectiveness . Current research aims include the application of behaviour-change models, the development and validation of prudent antibiotic prescribing standards, and the refinement of tools to assess educational interventions.

Am J Respir Med, 2003, 2(6), 491 - 8
Antibiotic prescribing patterns of French GPs for upper respiratory tract infections: impact of fusafungine on rates of prescription of systemic antibiotics; Fagnani F et al.; INTRODUCTION: Despite attempts to limit their use, systemic antibiotics are extensively prescribed for respiratory infections in France . This survey analyzed data from the Thales database, which contains information from 1010 representative French general practitioners (GPs) . The objective was to assess French GP prescribing patterns in upper respiratory tract infections (URTIs) including the rate of prescription of systemic antibiotics and anti-inflammatory drugs in the presence or absence of prescribing fusafungine (Locabiotal) an antibiotic with anti-inflammatory activity indicated for local use in URTIs . Drug costs to the French National Sickness Fund were also assessed . METHODS: This was a retrospective, longitudinal, case-control analysis . Prescribing patterns and costs were compared between patients who did and patients who did not receive fusafungine for a URTI (rhinopharyngitis, tonsillitis, or an influenza-like condition) . The fusafungine group consisted of all patients in the database who were prescribed fusafungine at least once between 1 December 1999 and 30 November 2000 . The control group was made up of randomly selected patients, matched for age and sex with the study group, who received at least one drug prescription (but not fusafungine) for a URTI during the same period . Patients were selected at the time of their first prescription, and their records for 1 year were analyzed . RESULTS: Each group contained 22 164 patients . For URTIs overall, systemic antibiotics were widely prescribed (at a rate of 54.6% and 67.8% in the fusafungine and control groups, respectively; p < 0.01) . The rate of prescription of systemic antibiotics, NSAIDs and corticosteroids per prescription and per episode was significantly lower in the fusafungine group than in the control group.The mean cost per prescription for the French National Sickness Fund was significantly lower for the three URTIs overall when fusafungine was prescribed (9.21 euros {euro} vs euro9.67; p < 0.01) . The mean cost to the National Sickness Fund per prescription of systemic antibiotics, NSAIDs, and corticosteroids was also significantly lower in the fusafungine group compared with the control group . The cost of nasal preparations was higher in the fusafungine group because Locabiotal is classified as a nasal preparation.The cost per prescription to the National Sickness Fund was increased by the presence of systemic antibiotics, NSAIDs, or corticosteroids among the prescribed drugs and decreased with the prescription of fusafungine . CONCLUSION: When fusafungine was prescribed for URTIs, fewer systemic antibiotics were prescribed, an important result in the current context of concern about emerging antibiotic resistance . The use of fusafungine was associated with a lower mean cost per prescription to the French National Sickness Fund.

Eur J Gen Pract, 2003 Sep, 9(3), 84 - 90
Public knowledge, attitudes and behaviour regarding antibiotics--a survey of patients in general practice; Emslie MJ et al.; OBJECTIVE: To assess public knowledge, attitudes and behaviour regarding antibiotics to provide information for a local health education policy . DESIGN: Postal survey to 605 patients registered with two general medical practices in Grampian, North-East Scotland (351 respondents, 59.3%) in February/ March 2000 . Questionnaire topics covered attitudes, knowledge and behaviour towards antibiotic use . RESULTS: 326 (93%) had experience of antibiotic use and 319 (81%) were happy to take antibiotics when necessary . 158 (45%) were concerned about antibiotic resistance . Few would expect antibiotics to treat a slight cold or heavy cold, but around half would expect antibiotics for very sore throats and influenza . 262 (75%) indicated they would ask a pharmacist for advice about the treatment of RTI . Most respondents experienced at least one episode of RTI during the review period . 280 (80%) provided details of their most recent episode; most self-treated, 57 consulted a doctor of whom 43 (75%) were prescribed antibiotics . CONCLUSIONS: Antibiotic resistance is a matter of concern for the public . Mixed opinions about the use of antibiotics for common respiratory tract infections were heard . Encouragingly the majority of people suffering an RTI during the review period treated themselves with paracetamol-based products for symptom relief . Those who consulted a doctor were in the minority but were likely to be prescribed antibiotics . Future local health education initiatives should target antibiotic use for sore throats as well as colds and 'flu.

Plasmid, 2004 Jan, 51(1), 12 - 23
Structural comparison of the integrative and conjugative elements R391, pMERPH, R997, and SXT; Boltner D et al.; R391 and SXT are members of a group of eleven chromosome-borne conjugative elements found in the gamma-proteobacteria, whose members carry different antibiotic resistance traits . Recent genomic analysis of R391 and SXT revealed a highly conserved 'backbone' encoding integration/excision, conjugative transfer, and regulation functions, augmented by an array of phenotypic traits and transposable elements . In this study, PCR amplification and sequence analysis were employed to investigate the genomic structure of two further MGE of the R391 family, pMERPH (HgR) and R997 (ApR, SmR, SuR) . R997 and pMERPH were found to be structurally related to R391 and SXT and share a number of virtually identical regions with them-including putative integration, conjugative transfer, and regulatory determinants-interrupted by variable DNA segments and transposable elements . The presence of a highly conserved backbone in the four elements strongly suggests their origin in a common ancestral element, which itself was a mosaic of sequences related to phages and plasmids . Subsequent genetic recombination and the acquisition of transposable elements resulted in the possession of variable phenotypic traits among the four MGE, and diversification into two distinct lineages, the first one including R391 and pMERPH, the second one containing SXT and R997.

Dent Update, 2003 Dec, 30(10), 570 - 4
Revisiting the role of dentists in prescribing antibiotics; Palmer NA; The purpose of this paper is to review the principles of therapeutic and prophylactic prescribing in light of the increasing problem of antibiotic resistance and the evidence of inappropriate use of antibiotics in dentistry . Recommendations based on a review of the evidence and good practice for prescribing therapeutic and prophylactic antibiotics will be given.

Eur J Clin Pharmacol, 2004 Mar, 60(1), 23 - 8 Epub 2003 Dec 19.
Respiratory tract infections in general practice: considerable differences in prescribing habits between general practitioners in Denmark and Spain; Bjerrum L et al.; OBJECTIVE: The prevalence of antibiotic resistance in a country reflects the local consumption of antibiotics . The majority of antibiotics are prescribed in general practice and most prescriptions are attributable to treatment of respiratory tract infections (RTIs) . The aim of this study was to compare general practitioners' (GPs') prescribing of antibiotics for respiratory tract infections in a country with a high prevalence of antibiotic resistance (Spain) with a country with a low prevalence of antibiotic resistance (Denmark) . METHODS: A group of GPs in Copenhagen and Barcelona registered all contacts ( n=2833) with patients with RTIs during a 3-week period between 1 November 2001 and 31 January 2002 . RESULTS: Overall, Spanish GPs treated a higher proportion of patients than Danish GPs . After adjusting for unequal distribution of age and sex, we found that Spanish GPs prescribed significantly more antibiotics to patients with focus of infection in tonsils and bronchi/lungs . Narrow-spectrum penicillin was the most used antibiotic in Denmark, representing 58% of all prescriptions issued, followed by macrolide and broad-spectrum penicillin . In Spain, prescriptions were distributed among a great number of compounds, with broad-spectrum penicillins and combinations of amoxicillin plus beta-lactamase inhibitors most frequently used . CONCLUSION: The substantial difference in the way GPs manage respiratory tract infections in Denmark and Spain cannot be explained by different patterns of RTIs in general practice . The discrepancies indicate variations in national recommendations, different treatment traditions or different impact of pharmaceutical marketing.

Int J Pediatr Otorhinolaryngol, 2004 Jan, 68(1), 29 - 36
International perspectives on management of acute otitis media: a qualitative review; Schilder AG et al.; BACKGROUND: Current opinions regarding the management of acute otitis media (AOM) in children vary across Western countries . With antibiotic resistance rising and more evidence regarding the limited clinical efficacy of antibiotics becoming available, interest in managing AOM other than with antibiotics is renewed . OBJECTIVES: To compare international rates of antibiotic prescription and surgery for AOM . To provide current evidence regarding the efficacy of various treatment options for AOM and their potential complications . METHODS: Qualitative (narrative) review . RESULTS: The percentage of patients given antibiotics for AOM varies from 31% in The Netherlands to more than 90% in most other Western countries . The 1998 prevalence of penicillin-resistant S . pneumoniae strains, on the other hand, ranges from 3% in The Netherlands to 53% in France . The surgical rate for tympanostomy tubes varies from 2 per 1000 children per year in the United Kingdom to 20 per 1000 in The Netherlands . The benefit of both antibiotic and surgical therapy in AOM appears to be limited, with numbers needed to treat ranging from 8 to 25 for antibiotic therapy, depending on the definition of outcome . CONCLUSIONS: International rates of antibiotic prescription and surgery for AOM vary strongly, which can be explained largely by the lack of uniform evidence-based guidelines . International debates with the aim of overcoming cultural differences regarding the management of otitis media, and of reaching agreement on guidelines on the basis of current evidence are necessary.

J Am Geriatr Soc, 2004 Jan, 52(1), 39 - 45
Antibiotic treatment of acute respiratory tract infections in the elderly: effect of a multidimensional educational intervention; Gonzales R et al.; OBJECTIVES: : To measure and improve antibiotic use for acute respiratory tract infections (ARIs) in the elderly . DESIGN: : Prospective, nonrandomized controlled trial . SETTING: : Ambulatory office practices in Denver metropolitan area (n=4 intervention practices; n=51 control practices) . PARTICIPANTS: : Consecutive patients enrolled in a Medicare managed care program who were diagnosed with ARIs during baseline (winter 2000/2001) and intervention (winter 2001/2002) periods . A total of 4,270 patient visits were analyzed (including 341 patient visits in intervention practices) . INTERVENTION: : Appropriate antibiotic use and antibiotic resistance educational materials were mailed to intervention practice households . Waiting and examination room posters were provided to intervention office practices . MEASUREMENTS: : Antibiotic prescription rates, based on administrative office visit and pharmacy data, for total and condition-specific ARIs . RESULTS: : There was wide variation in antibiotic prescription rates for ARIs across unique practices, ranging from 21% to 88% (median=54%) . Antibiotic prescription rates varied little by patient age, sex, and underlying chronic lung disease . Prescription rates varied by diagnosis: sinusitis (69%), bronchitis (59%), pharyngitis (50%), and nonspecific upper respiratory tract infection (26%) . The educational intervention was not associated with greater reduction in antibiotic prescription rates for total or condition-specific ARIs beyond a modest secular trend (P=.79) . CONCLUSION: : Wide variation in antibiotic prescription rates suggests that quality improvement efforts are needed to optimize antibiotic use in the elderly . In the setting of an ongoing physician intervention, a patient education intervention had little effect . Factors other than patient expectations and demands may play a stronger role in antibiotic treatment decisions in elderly populations.

Cell Commun Adhes, 2003 Jul-Dec, 10(4-6), 311 - 7
Development of a cell model for functional and structural analysis of connexin co-expression: achieving homogeneous and inducible expression of multiple connexins in stable transfectants; Halliday D et al.; We set out to develop an in vitro cell model in which connexins 43, 40 and 45 are co-expressed in the same combinations as found in different sub-types of cardiomyocyte in vivo, using inducible promoters of the Tet-Off and Ecdysone systems . In initial studies, a heterogeneous pattern of gene expression was observed . To achieve homogeneous expression, an Internal Ribosome Entry Site (IRES) sequence was employed, ensuring that a single mRNA coded for connexin and antibiotic resistance . We then constructed plasmids that combine the inducibility of the Tet-Off and Ecdysone systems with the homogeneous expression given by the IRES constructs . These were demonstrated to give inducible and homogeneous expression . By using the reporter gene, Enhanced Green Fluorescent Protein (EGFP), it was further shown in the Tet-Off system that expression of the transfected gene was modulated homogeneously in all cells when induction was repressed . The cell model is now at a suitable stage of development for investigation of the functional correlates of the distinctive connexin co-expression found in different regions of the heart.

Horm Res . 2003;60 Suppl 3:10.
Stem cells in diabetes: what has been achieved; Bonner-Weir S; Beta-cell replacement therapy via islet transplantation has received renewed interest due to the recent improved success . In order to make such a therapy available to more than a few of the thousands of patients with diabetes, new sources of insulin-producing cells must be readily available . The most promising sources are stem cells, with efforts of deriving new beta-cells from both embryonic and adult stem cells . Several groups have reported generating insulin-producing cells from mouse embryonic stem cells . The strategies in the first two acclaimed reports were very different . One strategy, used by Soria's group, is gene trapping in which an introduced antibiotic resistance under the control of the insulin promoter allowed the selection of insulin-expressing cells that had spontaneously differentiated within embryoid bodies . Another strategy, used by McKay's group, manipulated culture conditions in a multistep protocol used for generating neural cells but with changed final conditions . Since these reports, there have been modifications of the protocols in efforts to improve the yields and maturity of the resulting cells . While it is unclear if the insulin-producing cells in any of these studies are truly mature beta-cells, these studies show the clear potential of embryonic stem cells and support optimism that similar results will be possible with human embryonic stem cells . We know that new beta-cells are generated throughout adult life, but the identity of adult pancreatic stem cells has been elusive . The potential for expansion and differentiation of pluripotent adult stem cells, whether from bone marrow or as non-pancreas tissue resident SP cells, is being explored but has not yet yielded insulin-producing tissue . In contrast, insulin-producing cells have been generated in vitro from adult pancreatic tissues . We have been examining the hypothesis that the functional source for new beta-cells in the adult pancreas are mature duct epithelial cells that have regressed or lost their mature phenotype after replication . Others have isolated putative stem cells from islets and ducts . For adult cells the issue of expansion as well as of differentiation is a question . The field of generating new beta-cells from stem cells, either embryonic or adult, is still in its infancy . Each new report has been met with a mixture of excitement and skepticism . With continued efforts and rigorous assessments, hopefully the potential of generating enough new beta-cells from stem cells will be realized .

Int J Pediatr Otorhinolaryngol, 2003 Dec, 67 Suppl 1, S213 - 5
Is there a future for pediatric sinus surgery? An American perspective; Clary RA; The role of pediatric sinus surgery in chronic sinusitis continues to evolve . By the early 1990s, endoscopic sinus surgery was being touted as a primary intervention of chronic pediatric sinusitis . Around this same time these endoscopic techniques were being attempted in congenital nasosinus malformations, acute orbital complications of sinusitis and other areas . In 1996 the Brussels consensus paper on pediatric sinusitis recommended that the latter set of nasal disorders were "absolute" indications, and ironically, chronic sinusitis was a "possible indication" . This report reviews some of the changes which have impacted the use of sinus surgery during the last decade including the perception of nasal symptoms during the era of antibiotic resistance in bacteria, improved instrumentation, the role of adenoidectomy, research on the effects of sinus surgery on facial growth, and the research on surgical outcomes.

Hunan Yi Ke Da Xue Xue Bao, 2003 Jun, 28(3), 263 - 5
{Impact of non-target gene mutations and reduced permeability of outer membrane on quinolone resistance and multiple antibiotic resistance in Escherichia coli}; Zhao RH et al.; OBJECTIVE: To investigate the impact of non-target gene mutations and reduced permeability of the outer membrane on quinolone resistance and multiple antibiotic resistance (mar) in Escherichia coli . METHODS: Several high-level quinolone resistant isolates whose strains were accompanied by concurrent resistant to several antibiotics were selected . The PCR products of marOR region were sequenced to detect the possible gene changes and then the strains' outer membrane proteins (Omps) were extracted to analyze on the constitutive profiles . RESULTS: We found two site mutations in R410 isolate: 1,870(T-->C, Val-->Ala) and 1,879(A-->C, terminator-->Ser) . The OmpF deficiency was found in all 3 resistant strains . CONCLUSION: The increased expression of MarA induced by gene mutations in marOR and the constitutive profile changes of the Omps (OmpF deficient strains) may play some role in the quinolone resistance and multiple antibiotic resistance of Escherichia coli.

World J Surg, 2004 Jan, 28(1), 87 - 91 Epub 2003 Nov 26.
Predictors of surgical outcome for complicated pneumonia in children: impact of bacterial virulence; Margenthaler JA et al.; The charts of 110 children with community acquired bacterial pneumonia were reviewed . A subset of children who required surgical intervention for empyema or parapneumonic effusion was identified . Patients were divided into two treatment groups: antibiotics/tube thoracostomy alone (group 1) versus operative intervention (group 2) . Overall, 33 (30%) of the children required surgical intervention for complications . Seventeen (15%, group 1) were successfully treated with antibiotics/tube thoracostomy alone, while 16 (15%) in group 2 were treatment crossovers, failing this initial therapy . Of group 2 children, 4 (25%) underwent thoracotomy and lobectomy, while 12 (75%) underwent video-assisted thoracoscopic surgery (VATS) . Although group 2 children were younger than those in group 1 (4.4 +/- 3.6 versus 6.3 +/- 4.1 years, p<0.05) and had longer hospitalizations (20.1 +/- 10.1 versus 8.2 +/- 3.9 days, p<0.05), symptom duration, preoperative antibiotics, fibrinolytic use, and leukocytosis were similar (p>0.05) . Group 1 children had 13 (76%) positive cultures, 92% with pan-sensitivities, in contrast to group 2, which had 12 (75%) positive cultures, but only 33% were sensitive to first-line antibiotics (p<0.01) . Group 2 patients were also more likely to have complex multi-loculated empyemas, pneumatoceles, or pulmonary necrosis identified on imaging studies (100% versus 24%, p<0.01) . These data suggest that the natural history of pneumonia in children is heavily influenced by bacterial virulence . Tube thoracostomy and appropriate antibiotics remain effective for pan-sensitive, simple parapneumonic effusions and empyema . Complex parapneumonic effusions and empyema, however, which occur more frequently in the setting of first-line antibiotic resistance, often fail more conservative managements and may be best treated by earlier operative debridement.

Am J Gastroenterol, 2003 Nov, 98(11), 2403 - 8
Interleukin-1beta genetic polymorphism influences the effect of cytochrome P 2C19 genotype on the cure rate of 1-week triple therapy for Helicobacter pylori infection; Take S et al.; OBJECTIVES: Genetic polymorphism of interleukin (IL)-1beta is associated with differences in gastric acid suppression in response to Helicobacter pylori (H . pylori) infection . Thus, the polymorphism might affect H . pylori eradication therapy, as antibiotics used in treatment regimens may be acid sensitive . In this study, we examined the impact of IL-1beta genetic polymorphism on the cure rate of triple therapy for H . pylori in relation to cytochrome P (CYP) 2C19 genotype and antibiotic resistance . METHODS: A total 249 patients with peptic ulcer disease were randomized to receive one of the following regimens: amoxicillin and clarithromycin together with omeprazole, lansoprazole, or rabeprazole . CYP2C19 and IL-1beta-511 genetic polymorphisms were analyzed by polymerase chain reaction-restriction fragment length polymorphism . RESULTS: The intention-to-treat-based overall cure rate was 74.3% (95% CI=68-79%) . In the normal acid secretion IL-1beta genotype group, the cure rate among CYP2C19 poor metabolizers (93.3%, 95% CI=56-99%) was significantly higher than among subjects in the CYP2C19 homozygous (60.0%, 95% CI=38-78%) and heterozygous (63.6%, 95% CI=46-78%), i.e., extensive metabolizer, groups (p<0.05) . In the low acid secretion IL-1beta genotype group, there was no difference in the cure rate among the CYP2C19 genotype groups . Multiple logistic regression analysis identified susceptibility to clarithromycin (p<0.0001) and CYP2C19 genotype status (p=0.03) as significant independent factors for treatment failure . CONCLUSION: IL-1beta genetic polymorphism, although not an independent factor in treatment outcome, influences the impact of the CYP2C19 genotype on the cure rate of 1-wk triple therapy for H . pylori infection.

J Microbiol Methods, 2003 Dec, 55(3), 727 - 37
New disruption cassettes for rapid gene disruption and marker rescue in the yeast Yarrowia lipolytica; Fickers P et al.; Yarrowia lipolytica is one of the most extensively studied nonconventional yeasts . Unfortunately, few methods for gene disruption have been reported for this yeast, and all of them are time-consuming and laborious . The functional analysis of unknown genes requires powerful disruption methods . Here, we describe such a new method for rapid gene disruption in Y . lipolytica . This knockout system combines SEP method and the Cre-lox recombination system, facilitating efficient marker rescue . Versatility was increased by using both auxotrophic markers like ylURA3 and ylLEU2, as well as the antibiotic resistance marker hph . The hph marker, which confers resistance to hygromycin-B, allows gene disruption in a strain lacking any conventional auxothrophic marker . The disruption cassette was shown to integrate at the correct locus at an average frequency of 45% . Upon expression of Cre recombinase, the marker was excised at a frequency of 98%, by recombination between the two lox sites . This new method for gene disruption is an ideal tool for the functional analysis of gene families, or for creating large-scale mutant collections in general.

J Clin Microbiol, 2003 Nov, 41(11), 5011 - 4
Metronidazole-resistant Helicobacter pylori is more prevalent in patients with nonulcer dyspepsia than in peptic ulcer patients in a multiethnic Asian population; Lui SY et al.; The trend of increasing prevalence of antibiotic resistance among Helicobacter pylori strains has been suggested as a cause of the failure of treatment of H . pylori infections . In this study, 120 of 211 antral biopsy specimens from patients with dyspeptic symptoms were found to harbor H . pylori . The isolates from the 120 specimens were tested by the agar dilution method, and 38 (31.7%) were found to be metronidazole resistant . Among the 211 subjects, 81 of 115 (70.4%) patients with peptic ulcer (PU) were infected with H . pylori, whereas 39 of 96 (40.6%) patients with nonulcer dyspepsia (NUD) were infected with H . pylori . Interestingly, significantly more NUD patients than PU patients harbored metronidazole-resistant H . pylori (22 of 39 {56.4%} and 16 of 81 {19.8%}, respectively; P < 0.001) . A similar pattern was also observed among NUD patients of different ethnicities but not between male and female patients (23 of 78 {29.5%} and 15 of 42 {35.7%}, respectively; P = 0.54) . In the posttreatment follow-up, five of six patients who had positive urea breath test results, indicating treatment failure, were NUD patients . Of these, four harbored metronidazole-resistant H . pylori strains . This further illustrates the relevance of metronidazole-resistant H . pylori in NUD patients . The significantly higher percentage of metronidazole-resistant H . pylori isolates in NUD patients may be attributed to the protection offered by the mucus layer of the nonulcerated stomach to the bacteria that reside below it, resulting in organism exposure to sublethal concentrations of metronidazole and leading to the induction of metronidazole resistance . The results demonstrate that the H . pylori isolates colonizing NUD patients are more likely to be resistant to metronidazole . It will therefore be useful to reevaluate the use of metronidazole in the treatment of NUD patients infected with H . pylori.

J Mol Microbiol Biotechnol, 2003, 6(1), 29 - 40
New antibiotic resistance cassettes suitable for genetic studies in Borrelia burgdorferi; Elias AF et al.; In this report we describe two distinct approaches to develop new antibiotic resistance cassettes that allow for efficient selection of Borrelia burgdorferi transformants . The first approach utilizes fusions of borrelial flagellar promoters to antibiotic resistance markers from other bacteria . The AACC1 gene, which encodes a gentamicin acetyltransferase, conferred a high level of gentamicin resistance in B . Burfdorferi when expressed from these promoters . No cross-resistance occurred between this cassette and the kanamycin resistance cassette, which was previously developed in an analogous fashion . A second and different approach was taken to develop an efficient selectable marker that confers resistance to the antibiotic coumermycin A1 . A synthetic gene was designed from the GYRB301 allele of the coumermycin-resistant B . Burgdorferi strain B31-NGR by altering the coding sequence at the wobble position . The resulting gene, GYRB(SYN), encodes a protein identical to the product of GYRB301, but the genes share only 66% nucleotide identity . The nucleotide sequence of GYRB(SYN)is sufficiently divergent from the endogenous B . Burgdorferi GYRB gene to prevent recombination between them . The cassettes described in this paper improve our repertoire of genetic tools in B . Burgdorferi . These studies also provide insight into parameters governing recombination and gene expression in B . Burgdorferi .

Curr Opin Microbiol, 2003 Oct, 6(5), 452 - 6
Persistence of antibiotic resistant bacteria; Andersson DI; Bacterial adaptation to antibiotics has been very successful and over the past decade the increase in antibiotic resistance has generated considerable medical problems . Even though many drug resistances confer a fitness cost, suggesting that they might disappear by reducing the volume of antibiotic use, increasing evidence obtained from laboratory and epidemiological studies indicate that several processes will act to cause long-term persistence of resistant bacteria . Compensatory evolution that ameliorates the costs of resistance, the occurrence of cost-free resistances and genetic linkage between non-selected and selected resistances will confer a stabilization of the resistant bacteria . Thus, it is of importance that we forcefully implement strategies to reduce the rate of appearance and spread of resistant bacteria to allow new drug discovery to catch up with bacterial resistance development.

J Pediatr, 2003 Oct, 143(4), 438 - 44
Invasive pneumococcal infections in children with sickle cell disease in the era of penicillin prophylaxis, antibiotic resistance, and 23-valent pneumococcal polysaccharide vaccination; Adamkiewicz TV et al.; Rates and severity of pneumococcal infections in children with sickle cell disease were examined before licensure of pneumococcal-conjugated vaccine (PVC) . Rates of peak invasive infection rates in 1-year-old children with hemoglobin SS and mortality in those 0 to 10 years of age were 36.5 to 63.4 and 1.4 to 2.8 per 1000 person-years, respectively (>10 and 100 times as frequent as in the general population) . Overall, 71% of serotyped isolates (n=80) were PVC serotypes and 71% of nonvaccine serotype strains were penicillin-sensitive . Clinical presentation in children with hemoglobin SS (n=71; more with hypotension) and hemoglobin SC (n=18; more with acute chest syndrome, otitis media) differed . Penicillin nonsusceptibility (38% of isolates) varied between geographic study sites . Penicillin prophylaxis appeared less effective against intermediate and resistant strains . Of all infected children, meningitis developed in 20% and 15% died (hemoglobin SS, n=15 and 11; hemoglobin SC, n=1 each) . Factors associated with death included age >4 years (58%), serotype 19F, and not being followed by a hematologist (42% each) . The pneumococcal-polysaccharide vaccine was 80.4% effective within 3 years after vaccination (95% CI, 39.7, 93.6) . Children with sickle cell disease of all ages may benefit from PVC boosted with polysaccharide vaccination.

Dig Liver Dis, 2003 Aug, 35(8), 541 - 5
Primary and secondary antibiotic resistance of Helicobacter pylori strains isolated in central Italy during the years 1998-2002; Toracchio S et al.; BACKGROUND: Antibiotic resistance of Helicobacter pylori is the most common reason for failure in its eradication . AIM: To determine the incidence of primary and secondary resistance to tinidazole, clarithromycin and amoxycillin in Helicobacter pylori isolates from dyspeptic patients in central Italy and to evaluate the modifications of resistance over the period from 1998 to 2002 . METHODS: H . pylori strains were isolated from antral biopsies taken during upper endoscopy in 406 dyspeptic patients with no previous therapy against H . pylori, and in 96 patients who had already undergone one or more triple therapies . Antibiotic susceptibility test was performed using the screening agar method and agar dilution method . RESULTS: Overall primary resistance to tinidazole, clarithromycin and amoxycillin was 36.7, 23.4 and 0.2%, respectively . Secondary resistance rates were: tinidazole 69.8%, clarithromycin 82.3% and amoxycillin 1% . Resistance to clarithromycin was often associated with tinidazole resistance and was significantly higher in female patients (p<0.05) . Primary and secondary antibiotic resistance did not change during the 4 years of observation . CONCLUSIONS: The dyspeptic population with H . pylori infection in central Italy shows high levels of antibiotic resistance . Primary resistance to clarithromycin is most frequent in female patients . In patients with secondary resistance, dual resistance to clarithromycin and tinidazole is found in the majority of cases.

Internist (Berl), 2003 Jun, 44 Suppl 1, S44 - 58
{Community-acquired and nosocomial pneumonia}; Welte T; Mortality of pneumonia is low in the outside setting (1%) but rises up to 20% in hospital admitted patients . Early diagnosis and standardized therapy improve patient's prognosis . For community acquired pneumonia age, comorbidity and the setting of therapy (outside department, normal ward or intensive care unit) are the most important variables to choose an adequate antibiotic treatment . For nosocomial pneumonia risk stratification is according to severity of illness, length of hospital stay and antibiotic pretreatment . In the outpatient setting a 7-day monotherapy is mostly successful . In severe illness the combination of a betalactam antibiotic with a new fluorchinolon seems to be superior to an aminoglycosid therapy . Antibiotic resistance due to mistakes in antibiotic therapy is an increasing problem in the intensive care unit . Therefore, pneumonia preventive measures like influenza and pneumococcal vaccination become more important . Standardized hygienical procedures help to reduce nosocomial, mainly ventilator associated pneumonia.

Gene, 2003 Oct 16, 316, 33 - 8
DNA transfer from chloroplast to nucleus is much rarer in Chlamydomonas than in tobacco; Lister DL et al.; By transforming chloroplasts with an antibiotic-resistance gene under the control of a nuclear-specific promoter, we employed a selection scheme to detect the transfer of DNA from the chloroplast to the nucleus in the green alga Chlamydomonas reinhardtii . Among several billion homoplasmic cells tested, we were unable to detect any stable nuclear integration of chloroplast DNA under normal growth conditions or under stress conditions . This contrasts with results reported for the transfer of DNA from chloroplast to nucleus in higher plants and from mitochondrion to nucleus in Saccharomyces cerevisiae . Furthermore, we were unable to detect chloroplast DNA-derived sequences among nuclear genome data for C . reinhardtii, which also contrasts with the situation in higher plants . Taken together, these findings suggest that there is presently little, if any, movement of DNA from chloroplast to nucleus in C . reinhardtii, which may reflect the ultrastructure of the C . reinhardtii cell.

Biosens Bioelectron, 2003 Oct 30, 19(1), 1 - 8
DNA microarray for discrimination between pathogenic 0157:H7 EDL933 and non-pathogenic Escherichia coli strains; Wu CF et al.; The primary technique currently used to detect biological agents is based on immunoassays . Although sensitive and specific, currently employed immunoassays generally rely on the detection of a single epitope, and therefore often cannot discriminate subtle strain-specific differences . Since DNA microarrays can hybridize hundreds to thousands of genomic targets simultaneously and do not rely on phenotypic expression of these genetic features for identification purposes, they have enormous potential to provide inexpensive, flexible and specific strain-specific detection and identification of pathogens . In this study, pathogenic Escherichia coli O157:H7-specific genes, non-pathogenic K12-specific genes, common E . coli genes, and negative control genes were polymerase chain reaction-amplified and spotted onto the surface of treated glass slides . After labeled bacterial cDNA samples were hybridized with probes on the microarray, specific fluorescence patterns were obtained, enabling identification of pathogenic E . coli O157:H7 and non-pathogenic E . coli K12 . To test the utility of this microarray device to detect genetically engineered bacteria, E . coli BL21 (a B strain derivative with antibiotic resistance gene, ampR) and E . coli JM107 (a K12 strain derivative lacking the gene ompT) were also employed . The array successfully confirmed the strain genotypes and demonstrated that antibiotic resistance can also be detected . The ability to assess multiple data points makes this array method more efficient and accurate than a typical immunoassay, which detects a single protein product.

Proc Natl Acad Sci U S A, 2003 Oct 28, 100(22), 12944 - 9 Epub 2003 Oct 10.
Treatment of Helicobacter gastritis with IL-4 requires somatostatin; Zavros Y et al.; Fifty percent of the world's population is infected with Helicobacter pylori; however, treatment has been insufficient to eradicate the organisms due to rising antibiotic resistance . Helicobacter infection is characterized by induction of a T helper 1 lymphocyte (Th1) immune response, hypergastrinemia, and suppressed tissue somatostatin (SOM) levels . However, the mechanism by which the immune response regulates acid secretion is not known . We show here that treatment with IFN-gamma, a Th1 cytokine, was sufficient to induce gastritis, increase gastrin, and decrease SOM levels within 7 days . In contrast, the T helper 2 lymphocyte cytokine IL-4 increased SOM levels and effectively suppressed gastrin expression and secretion . This result demonstrated reciprocal regulation of acid regulatory peptides by immune modulators . IL-4 pretreatment prevented gastritis in infected wild-type but not in SOM null mice . Thus, the ability of IL-4 to oppose a Th1-mediated infection required SOM . Immunofluorescence was used to document the presence of IL-4 receptors on the gastric SOM-secreting cell (D cell) . Moreover, IL-4 stimulated SOM release from primary D cell cultures . Treatment of mice chronically infected with Helicobacter felis for 2 mo with the SOM analogue octreotide resolved the inflammation . Thus, a mechanism by which IL-4 resolves inflammation in the stomach is by stimulating the release of SOM from gastric D cells.

Biotechnol Adv, 2000 May, 18(3), 179 - 206
The impact of genetic modification of human foods in the 21st century: a review; Uzogara SG; Genetic engineering of food is the science which involves deliberate modification of the genetic material of plants or animals . It is an old agricultural practice carried on by farmers since early historical times, but recently it has been improved by technology . Many foods consumed today are either genetically modified (GM) whole foods, or contain ingredients derived from gene modification technology . Billions of dollars in U.S . food exports are realized from sales of GM seeds and crops . Despite the potential benefits of genetic engineering of foods, the technology is surrounded by controversy . Critics of GM technology include consumer and health groups, grain importers from European Union (EU) countries, organic farmers, environmentalists, concerned scientists, ethicists, religious rights groups, food advocacy groups, some politicians and trade protectionists . Some of the specific fears expressed by opponents of GM technology include alteration in nutritional quality of foods, potential toxicity, possible antibiotic resistance from GM crops, potential allergenicity and carcinogenicity from consuming GM foods . In addition, some more general concerns include environmental pollution, unintentional gene transfer to wild plants, possible creation of new viruses and toxins, limited access to seeds due to patenting of GM food plants, threat to crop genetic diversity, religious, cultural and ethical concerns, as well as fear of the unknown . Supporters of GM technology include private industries, research scientists, some consumers, U.S . farmers and regulatory agencies . Benefits presented by proponents of GM technology include improvement in fruit and vegetable shelf-life and organoleptic quality, improved nutritional quality and health benefits in foods, improved protein and carbohydrate content of foods, improved fat quality, improved quality and quantity of meat, milk and livestock . Other potential benefits are: the use of GM livestock to grow organs for transplant into humans, increased crop yield, improvement in agriculture through breeding insect, pest, disease, and weather resistant crops and herbicide tolerant crops, use of GM plants as bio-factories to yield raw materials for industrial uses, use of GM organisms in drug manufacture, in recycling and/or removal of toxic industrial wastes . The potential risks and benefits of the new technology to man and the environment are reviewed . Ways of minimizing potential risks and maximizing the benefits of GM foods are suggested . Because the benefits of GM foods apparently far outweigh the risks, regulatory agencies and industries involved in GM food business should increase public awareness in this technology to enhance worldwide acceptability of GM foods . This can be achieved through openness, education, and research.

Issue brief (Grantmakers Health), 2000 Oct 3, (7), 1 - 26
Progress and peril: examining antibiotic resistance and systemic contaminants; Grantmakers In Health et al.; As part of its ongoing mission to inform and support trustees and staff of health foundations and corporate giving programs, Grantmakers In Health (GIH), in conjunction with the Health and Environmental Funders Network (HEFN), held a day-long Issue Dialogue on two important environmental health issues: antibiotic resistance and systemic contaminants . Experts and advocates from many organizations came together to discuss concerns and opportunities, as well as current and potential roles for health philanthropy.This Issue Brief brings together key points from the day's discussion with factual information on environmental health drawn from a background paper prepared for the participants.

J Med Microbiol, 2003 Nov, 52(Pt 11), 981 - 7
Characterization of German penicillin non-susceptible serotype 23F pneumococci using multilocus sequence typing; Reinert RR et al.; Three nationwide multicentre studies (n = 5071) showed an increase in antibiotic resistance in pneumococci in Germany . Serotype 23F was the predominant serotype (n = 45, 22.4 %), followed by 6B (n = 30, 14.9 %) and 9V (n = 19, 9.5 %) . Multilocus sequence typing was used to characterize 45 serotype 23F strains with reduced penicillin susceptibility . The Spanish(23F)-1 clone {profile 4-4-2-4-4-1-1, sequence type (ST) 81} contributes significantly to the emergence of penicillin resistance in Germany (n = 21, 46.7 % of all penicillin non-susceptible serotype 23F isolates) . Isolates of ST 277 (profile 7-13-8-6-6-12-8), which has been found previously in the Netherlands, are also observed, particularly in western Germany (n = 8, 17.8 %) . A high proportion of strains (n = 11, 24.4 %) have sequence types that have not been reported to date from other countries (STs 353-362) . The major penicillin-resistant clones are present in Germany, a country with relatively low levels of beta-lactam resistance.

Biophys J, 2003 Oct, 85(4), 2323 - 32
Amphotericin B channels in the bacterial membrane: role of sterol and temperature; Venegas B et al.; Amphotericin B is an antibiotic that forms ion channels in the membrane of a host cell . The change in permeability produced by these channels is greatly improved by sterols; nevertheless, the single channel conductivity remains invariant . Hence, it is proposed that sterols do not act directly, but rather through the modulation of the membrane phase . We look at the formation of these channels in the bacterial membrane to determine the mechanism of its known antibiotic resistance . We found that channels can indeed be formed in this membrane, but a substantial amount of amphotericin B is required . We also study the effects of the antibiotic concentration needed for channel expression as well as the dynamics of channels affected by both sterol and temperature in phosphatidylcholine membranes . The results support the idea that membrane structure is a determining factor in the action of the antibiotic.

Clin Infect Dis, 2003 Oct 1, 37(7), 882 - 9 Epub 2003 Sep 05.
Effects of primary metronidazole and clarithromycin resistance to Helicobacter pylori on omeprazole, metronidazole, and clarithromycin triple-therapy regimen in a region with high rates of metronidazole resistance; Wong WM et al.; The aim of this study was to investigate the effect of metronidazole resistance (MtzR) and clarithromycin resistance (ClaR) on the eradication rate for omeprazole, clarithromycin, and metronidazole triple-therapy regimen and on the development of posttherapy drug resistance in a region of high rates of MtzR . One hundred ninety-six Helicobacter pylori isolates were recovered from patients with duodenal ulcer, gastric ulcer, or nonulcer dyspepsia during upper endoscopy . The prevalences of MtzR, ClaR, and dual resistance were 37.8%, 13.8%, and 8.7%, respectively . The intention-to-treat eradication rates for metronidazole-susceptible (87.2% vs . 67.6%; P=.001) and clarithromycin-susceptible (86.4% vs . 40.7%; P<.001) strains were significantly higher than the rates for resistant strains . Multiple logistic regression analysis implicated younger age (<40 years old), MtzR, ClaR, and the diagnosis of nonulcer dyspepsia as independent factors that predicted treatment failure . The rates of posttreatment MtzR, ClaR, and dual resistance were 88%, 88%, and 75%, respectively . MtxR and ClaR significantly affected the success of eradication therapy . Posttreatment rates of resistance were high and were related to the presence of pretreatment antibiotic resistance.

Science, 2003 Sep 12, 301(5639), 1496 - 9
Toxins-antitoxins: plasmid maintenance, programmed cell death, and cell cycle arrest; Hayes F; Antibiotic resistance, virulence, and other plasmids in bacteria use toxin-antitoxin gene pairs to ensure their persistence during host replication . The toxin-antitoxin system eliminates plasmid-free cells that emerge as a result of segregation or replication defects and contributes to intra- and interspecies plasmid dissemination . Chromosomal homologs of toxin-antitoxin genes are widely distributed in pathogenic and other bacteria and induce reversible cell cycle arrest or programmed cell death in response to starvation or other adverse conditions . The dissection of the interaction of the toxins with intracellular targets and the elucidation of the tertiary structures of toxin-antitoxin complexes have provided exciting insights into toxin-antitoxin behavior.

Plant Physiol, 2003 Sep, 133(1), 402 - 10
Persistence of unselected transgenic DNA during a plastid transformation and segregation approach to herbicide resistance; Ye GN et al.; The use of a nonlethal selection scheme, most often using the aadA gene that confers resistance to spectinomycin and streptomycin, has been considered critical for recovery of plastid transformation events . In this study, the plastid-lethal markers, glyphosate or phosphinothricin herbicides, were used to develop a selection scheme for plastids that circumvents the need for integration of an antibiotic resistance marker . The effect of selective agents on tobacco (Nicotiana tabacum) mesophyll chloroplasts was first examined by transmission electron microscopy . We found that at concentrations typically used for selection of nuclear transformants, herbicides caused rapid disintegration of plastid membranes, whereas antibiotics had no apparent effect . To overcome this apparent herbicide lethality to plastids, a "transformation segregation" scheme was developed that used two independent transformation vectors for a cotransformation approach and two different selective agents in a phased selection scheme . One transformation vector carried an antibiotic resistance (aadA) marker used for early nonlethal selection, and the other transformation vector carried the herbicide (CP4 or bar) resistance marker for use in a subsequent lethal selection phase . Because the two markers were carried on separate plasmids and were targeted to different locations on the plastid genome, we reasoned that segregation of the two markers in some transplastomic lines could occur . We report here a plastid cotransformation frequency of 50% to 64%, with a high frequency (20%) of these giving rise to transformation segregants containing exclusively the initially nonselected herbicide resistance marker . Our studies indicate a high degree of persistence of unselected transforming DNA, providing useful insights into plastid chromosome dynamics.

Plant J, 2003 Sep, 35(6), 811 - 21
Rapid and proven production of transplastomic tobacco plants by restoration of pigmentation and photosynthesis; Klaus SM et al.; Tobacco chloroplast transformation is typically achieved using dominant, selectable antibiotic resistance genes such as aadA, nptII and aphA-6 . An improvement would be the combination of such a marker with a visual screening system for the early and conclusive detection of plastid transformants . As such, we investigated the use of three photosynthesis-deficient plastid mutants, DeltapetA, Deltaycf3 and DeltarpoA, for the development of a phenotypic selection system . Mutant plants were used as an alternative to the wild-type as source tissue for transformation, re-introducing deleted plastid sequences and using the aphA-6 gene as a selection marker . The reconstitution of the deleted genes in transformed regenerants resulted in shoots with a visually distinct phenotype comparable to the wild-type . This transformation/selection system overcomes the common problems associated with plastid transformation, e.g . the recovery of spontaneous mutants or nuclear insertions . In addition to the benefits offered by phenotypic selection, phenotype reconstitution leads to restoration of photosynthesis, which we assume drives reconstituted plants rapidly towards homoplasmy . As such, repeated cycles of regeneration in the presence of an antibiotic selection agent are no longer required.

ORL Head Neck Nurs, 2003 Summer, 21(3), 14 - 7
The pediatric tympanic membrane: see it, describe it, treat it; McDivitt K; Acute otitis media (AOM) is one of the most common childhood conditions for which antibiotics are prescribed, often unnecessarily . Current medical literature focuses on the treatment of AOM, appropriate antibiotics, and antibiotic resistance . Placing more emphasis on examination and description of the tympanic membrane (TM) reduces the over-diagnosing of AOM and concerns associated with antibiotic use . A thorough examination of the ear must be performed in order to make an accurate diagnosis . This article presents a three-step technique to assist providers to perform pneumatic otoscopy, visualize the TM, describe their findings, and subsequently treat AOM.

Cleve Clin J Med, 2003 Aug, 70(8), 670, 672 - 4, 677-8 passim
Acne vulgaris: one treatment does not fit all; Longshore SJ et al.; With many treatments now available for acne vulgaris, the treatment must be tailored to the type and severity of the lesions . Most mild-to-moderate cases can be treated with a benzoyl peroxide product, a topical or oral antibiotic, a topical retinoid, or a combination of these medications . Antibiotic resistance is becoming a challenge for many once-reliable topical and oral antibiotics.

Appl Environ Microbiol, 2003 Sep, 69(9), 5039 - 44
Efficient allelic exchange and transposon mutagenesis in Mycobacterium avium by specialized transduction; Otero J et al.; Mycobacterium tuberculosis and Mycobacterium avium are pathogenic slow-growing mycobacteria that cause distinct human diseases . In contrast to recent advances in M . tuberculosis genetics and pathogenesis investigation, M . avium has remained genetically intractable and, consequently, its pathogenic strategies remain poorly understood . Here we report the successful development of efficient allelic exchange and transposon mutagenesis in an opaque clinical strain of M . avium by specialized transduction . Efforts to disrupt the leuD gene of M . avium by specialized transduction were successful but were complicated by inefficient isolation of recombinants secondary to high spontaneous antibiotic resistance . However, by using this leucine auxotroph as a genetic host and the Streptomyces coelicolor leuD gene as a selectable marker, we achieved efficient allelic exchange at the M . avium pcaA locus . A leuD-marked transposon delivered by specialized transduction mutagenized M . avium with efficiencies similar to M . tuberculosis . These results establish a system for random and directed mutagenesis of M . avium . In combination with the forthcoming M . avium genome sequence, these tools will allow the distinct physiologic and pathogenic properties of M . avium to be dissected in molecular detail.

Pharmacogenomics, 2003 Sep, 4(5), 657 - 65
Antibiotics and pharmacogenomics; Davison DB et al.; The widespread use of antibiotics impacts both the bacterial ecology and the host at multiple levels, both advantageously and deleteriously . Since serious bacterial infection can lead to death in the absence of antibiotic therapy, antibiotics remain a necessary weapon in the clinician's arsenal for maintaining good health . It is thus critical that the placement and usage of these crucial antibiotics be constantly improved, and that emerging antibiotic resistance is vigorously assessed . To realize both these ends, it may be valuable to turn to the discipline of pharmacogenomics and develop it for use in a fairly novel way, that is, as the means by which to determine and analyze the impact of antibiotic therapy on both the pathogen and host.

Antimicrob Agents Chemother, 2003 Sep, 47(9), 3030 - 3
Roles of TolC-dependent multidrug transporters of Escherichia coli in resistance to beta-lactams; Nishino K et al.; AcrAB exports some beta-lactam antibiotics in the periplasm out of cells via an outer-membrane channel, TolC . It has been reported that eight drug transporters in Escherichia coli cooperate with TolC . In this study, the roles of the drug exporters of E . coli in beta-lactam resistance were examined . We found that five of five resistance-nodulation-cell division-type drug exporters confer beta-lactam antibiotic resistance, while no other drug exporters confer any beta-lactam resistance even when they cooperate with TolC.

J Infect Dis, 2003 Sep 1, 188(5), 759 - 68 Epub 2003 Aug 15.
Phylogenetic origin and virulence genotype in relation to resistance to fluoroquinolones and/or extended-spectrum cephalosporins and cephamycins among Escherichia coli isolates from animals and humans; Johnson JR et al.; In Escherichia coli infection, the implications of fluoroquinolone (FQ) and extended-spectrum cephalosporin plus cephamycin (AmpC) resistance for phylogenetic origin and virulence potential are undefined, as is the influence of ecological context on these associations . Accordingly, 106 E . coli isolates exhibiting FQ and/or AmpC resistance and 98 susceptible isolates were compared with regard to phylogenetic background and virulence profiles, stratified by host group (104 predominantly extraintestinal human isolates and 100 predominantly intestinal cattle and swine isolates) . Although resistant isolates exhibited significant shifts in phylogenetic distribution and virulence profiles, human and animal isolates exhibited different phylogenetic shifts, and only among human isolates did resistance predict reduced virulence . Evidence for similar strains being resistant versus susceptible was scant . The O15:K52:H1 clonal group and the closely related "clonal group A" featured prominently among resistant and susceptible human isolates, respectively . Thus, in E . coli, antibiotic resistance predicts phylogenetic background and virulence potential in a complex, context-dependent fashion.

Trends Biochem Sci, 2003 Aug, 28(8), 411 - 8
RNA, the first macromolecular catalyst: the ribosome is a ribozyme; Steitz TA et al.; Recently, the atomic structures of the large ribosomal subunit from Haloarcula marismortui and its complexes with substrates have been determined . These have provided exciting new insights into the principles of RNA structure, the mechanism of the peptidyl-transferase reaction and early events in the evolution of this RNA-protein complex assembly that is essential in all cells . The structures of the large subunit bound to a variety of antibiotics explain the effects of antibiotic resistance mutations and provide promise for the development of new antibiotics.

Bull Entomol Res, 2003 Aug, 93(4), 323 - 33
Patterns of resistance to three cereal aphids among wheats in the genus Triticum (Poaceae); Lamb RJ et al.; Forty-one accessions of wild and cultivated wheats belonging to 19 Triticum species were tested in the field for resistance to three species of aphids, Rhopalosiphum padi Linnaeus, Sitobion avenae Fabricius and Schizaphis graminum Rondani . Antibiotic resistance was estimated by the increase in biomass of aphids over 21 days on adult plants . Overall resistance was estimated by the plant biomass lost due to aphid infestation . All three species of aphids survived and reproduced on all wheats, and reduced spike biomass compared to uninfested controls . The level of antibiosis varied among wheat species and among accessions, with accessions from three, five and one species showing antibiosis to R . padi, S . avenae and S . graminum, respectively . Overall resistance to the three aphid species was observed in five to seven accessions per aphid species . Resistance was usually specific to one aphid species . The frequency of accessions with antibiosis or overall resistance was associated with the ploidy level of the plant species . Except for overall resistance to R . padi, resistance was highest for diploid species and lowest for hexaploid species . No consistent relationship between resistance and level of domestication was detected . Accessions of the wild wheats, Triticum boeoticum Bois, Triticum tauschii (Coss.) Schmal . and Triticum araraticum Jakubz . exhibited high levels of resistance to aphids, as did Triticum monococcum L . which is derived from T . boeoticum . Nevertheless, individual susceptible or resistant accessions occurred at all levels within the evolutionary tree of wheat.

Can J Gastroenterol, 2003 Jun, 17 Suppl B, 33B - 35B
Is it time for quadruple therapy to be first line?
Laine L.
The most commonly used regimen for Helicobacter pylori therapy at present is twice-daily proton pump inhibitor (PPI)-based triple therapy . Bismuth-based therapy is the next most common treatment used by gastroenterologists . When a PPI is combined with bismuth-based triple therapy (quadruple therapy), eradication rates are increased as compared with the triple therapy alone . Three separate randomized trials from three continents that compare quadruple therapy and PPI-based triple therapy revealed remarkably similar results . Eradication rates with PPI-based triple therapy and quadruple therapy were not significantly different . The eradication rates with quadruple therapy were 3% to 6% higher than PPI triple therapy, indicating that quadruple therapy should be no less effective than PPI triple therapy . Furthermore, these two therapies had similar rates of compliance and adverse events . The major potential benefit of the quadruple therapy relates to antibiotic resistance . In patients with clarithromycin resistance, PPI-based triple therapy, but not quadruple therapy, had a significantly lower eradication rate . However, due to its ability to largely overcome metronidazole resistance, quadruple therapy had little if any decrement in eradication rates compared with PPI triple therapy inpatients with metronidazole-resistant H pylori . Therefore, quadruple therapy can be considered a first line therapy for H pylori.

J Am Chem Soc, 2003 Aug 13, 125(32), 9612 - 8
Structural aspects for evolution of beta-lactamases from penicillin-binding proteins; Meroueh SO et al.; Penicillin-binding proteins (PBPs), biosynthetic enzymes of bacterial cell wall assembly, and beta-lactamases, resistance enzymes to beta-lactam antibiotics, are related to each other from an evolutionary point of view . Massova and Mobashery (Antimicrob . Agents Chemother . 1998, 42, 1-17) have proposed that for beta-lactamases to have become effective at their function as antibiotic resistance enzymes, they would have had to undergo structure alterations such that they would not interact with the peptidoglycan, which is the substrate for PBPs . A cephalosporin analogue, 7beta-{N-Acetyl-L-alanyl-gamma-D-glutamyl-L-lysine}-3-acetoxymethyl-3-cephem-carboxylic acid (compound 6), was conceived and synthesized to test this notion . The X-ray structure of the complex of this cephalosporin bound to the active site of the deacylation-deficient Q120L/Y150E variant of the class C AmpC beta-lactamase from Escherichia coli was solved at 1.71 A resolution . This complex revealed that the surface for interaction with the strand of peptidoglycan that acylates the active site, which is present in PBPs, is absent in the -lactamase active site . Furthermore, insertion of a peptide in the beta-lactamase active site at a location where the second strand of peptidoglycan in some PBPs binds has effectively abolished the possibility for such interaction with the beta-lactamase . A 2.6 ns dynamics simulation was carried out for the complex, which revealed that the peptidoglycan surrogate (i.e., the active-site-bound ligand) undergoes substantial motion and is not stabilized for binding within the active site . These factors taken together disclose the set of structure modifications in the antibiotic resistance enzyme that prevent it from interacting with the peptidoglycan, en route to achieving catalytic proficiency for their intended function.

J Fam Pract . 2003 Aug;52(8):594, 597.
Low-dose doxycycline moderately effective for acne; Miller ST et al.; The authors propose that moderate acne may be treated with doxycycline in subantimicrobial doses (20-mg tablets taken twice daily) . This regimen was well-tolerated, moderately effective in reducing skin lesions, and did not have a detectable effect on the antibiotic resistance of skin flora . The cost of Periostat (the only form of doxycycline 20 mg available in the US) is about 55 US dollars per month, while generic doxycycline 100 mg is about 10 US dollars . This study has some significant flaws, but a trial of low-dose doxycycline in an adult with acne severe enough to warrant antibiotics would still seem a reasonable, albeit expensive, option

Saudi Med J, 2003 Jul, 24(7), 730 - 5
Morbidity and mortality data of cystic fibrosis patients; Banjar H; OBJECTIVE: To identify factors that contributed to morbidity and mortality of cystic fibrosis (CF) population in the Kingdom of Saudi Arabia (KSA) . METHODS: This retrospective chart review was carried out in King Faisal Specialist Hospital and Research Centre, Riyadh, KSA, during a 9 year period, November 1993 to November 2002, on confirmed CF patients, for demographic, clinical and mortality data . RESULTS: A total of 190 CF patients were diagnosed during the 9 years . One hundred and sixty-four (86%) patients are alive, 26 (14%) died . Ninety-nine (52%) were males and 91 (48%) were females . Age at diagnosis 2.8 3.5 years, and period of follow up 3 3 years . In 80% of patients, symptoms started <1 year of age . Sixty-five percent of patients were in the mild to moderate malnutrition stage (<90th percentile), and 63% are in the mild to moderate stunted growth (<90th percentile) . Factors that contributed to early mortality are: calculated weight/height (p-value 0.01), low albumin level at follow up (0.001), high hematocrit (HCT) (p-values=0.0002), low mean corpuscular volume (MCV) (p-0.0002), low mean corpuscular hemoglobin concentration (MCHC) (p-value 0.001), early development of antibiotic resistance (p-value=< 0.01) . CONCLUSION: High HCT, low MCV, low MCHC and low albumin are factors related to poor prognosis and early death in CF patients . Iron supplement should be given to these patients even in the presence of normal hemoglobin . Early nutritional rehabilitation is needed to improve survival of our CF patients . Cohort isolation should be encouraged in CF centres . Early treatment of chronic pseudo colonization should be adopted to improve survival.

Rev Med Chil, 2003 May, 131(5), 505 - 14
{Community acquired pneumococcal pneumonia in hospitalized adult patients}; Diaz A et al.; BACKGROUND: S pneumoniae is the most common cause of community-acquired pneumonia . AIM: To evaluate the clinical characteristics, antibiotic resistance, management and prognostic factors in pneumococcal pneumonia . METHODS: Prospective evaluation in 46 adults (age +/- sd: 68 +/- 17 years) hospitalized with pneumococcal pneumonia confirmed by sputum, blood or pleural fluid cultures . Clinical and radiographic variables, risk factors for antibiotic resistance, and hospital mortality rate were recorded . RESULTS: Heart disease (39%), COPD/asthma (25%), and diabetes mellitus (18%) were the most frequent underlying diseases . None of the patients had previously received pneumococcal vaccine . Only 17% of the patients had the classic triad of chills, fever and productive cough . At admission, interestingly, 17% presented with congestive heart failure . Resistance of pneumococci to penicillin, cefotaxime or erythromycin was 15%, 6% and 11%, respectively . Antibiotic use prior to admission was significantly associated with antibiotic resistance (OR = 6; CI 95% = 1.1-32; p < 0.05) . Fifty per cent of the patients were admitted to intermediate or intensive care units, 15% were mechanically ventilated, 20% developed septic shock, 20% developed acute renal failure and 13% died in the hospital . Clinical factors significantly associated with higher mortality were systolic hypotension (< or = 90 mmHg), ICU admission and BUN > 30 mg per dL . CONCLUSIONS: Our data suggest that pneumococcal pneumonia is still a severe infection with high mortality; hence, efforts should be made at prevention using pneumococcal immunization.

Ann Epidemiol, 2003 Jul, 13(6), 479 - 83
Antibiotic prescribing for cystitis: how well does it match published guidelines?
McEwen LN, Farjo R, Foxman B.
PURPOSE: Cystitis is one of the most common bacterial infections seen by physicians in outpatient settings . Published clinical guidelines by the Infectious Disease Society of America and other organizations have been established to enable effective treatment, while attempting to decrease cost and limit antibiotic resistance . METHODS: Insurance claims data for employees and dependents of a single Midwest corporation, with Preferred Provider Organization coverage, diagnosed with cystitis between 1996 and 1999 were matched to prescription drug claims for those who filled an antibiotic prescription within 3 days of diagnosis . RESULTS: For acute and recurrent cystitis physicians prescribed trimethoprim-sulfamethoxazole 37% and 18% respectively . The other most common antibiotics prescribed were the broad-spectrum flouroquinolones, and nitrofurantoin . The mean duration for these prescriptions was 10 days regardless of whether the infection was acute or recurrent . CONCLUSIONS: The first line recommended antibiotic, trimethoprim-sulfamethoxazole, was prescribed in 37% of acute infections, and for considerably longer than the suggested 3-day course of therapy . Steps should be taken to educate physicians and patients on the choice and dosage of antibiotics for cystitis to minimize emergence of antibiotic resistance.

Genetics, 2003 Jul, 164(3), 843 - 54
The evolution of mutator genes in bacterial populations: the roles of environmental change and timing; Tanaka MM et al.; Recent studies have found high frequencies of bacteria with increased genomic rates of mutation in both clinical and laboratory populations . These observations may seem surprising in light of earlier experimental and theoretical studies . Mutator genes (genes that elevate the genomic mutation rate) are likely to induce deleterious mutations and thus suffer an indirect selective disadvantage; at the same time, bacteria carrying them can increase in frequency only by generating beneficial mutations at other loci . When clones carrying mutator genes are rare, however, these beneficial mutations are far more likely to arise in members of the much larger nonmutator population . How then can mutators become prevalent? To address this question, we develop a model of the population dynamics of bacteria confronted with ever-changing environments . Using analytical and simulation procedures, we explore the process by which initially rare mutator alleles can rise in frequency . We demonstrate that subsequent to a shift in environmental conditions, there will be relatively long periods of time during which the mutator subpopulation can produce a beneficial mutation before the ancestral subpopulations are eliminated . If the beneficial mutation arises early enough, the overall frequency of mutators will climb to a point higher than when the process began . The probability of producing a subsequent beneficial mutation will then also increase . In this manner, mutators can increase in frequency over successive selective sweeps . We discuss the implications and predictions of these theoretical results in relation to antibiotic resistance and the evolution of mutation rates.

Mini Rev Med Chem, 2003 Sep, 3(6), 540 - 56
Plant-derived triterpenoids as potential antineoplastic agents; Setzer WN et al.; Man has relied on plants as a source of medicinal agents for centuries . Today, with the specter of antibiotic resistance, emerging infectious diseases, and cancers, phytochemicals continue to provide new structural leads for the chemotherapeutic industry . A number of triterpenoids have shown promise as antineoplastic agents . Members of the cycloartane, lupane, ursane, oleanane, friedelane (especially quinone methides), dammarane, cucurbitacin, and limonoid triterpenoids, have demonstrated anti-proliferative activity on various cancer cell lines . This review covers the recent developments regarding antineoplastic/cytotoxic triterpenoids, excluding saponins, from higher plants.

J Antimicrob Chemother, 2003 Aug, 52(2), 282 - 6 Epub 2003 Jul 15.
Antibiotic prescriptions in children; Resi D et al.; OBJECTIVES: The aim of this study was to evaluate antibiotic prescription for children in Emilia Romagna, a Northern Italian region with 414 880 people aged 1-14 years . METHODS: The regional Prescription Database of drugs reimbursed by the Italian National Health Service was used in this study . Antibiotic use was estimated as the proportion of children who received at least one prescription during the year 2000 (number of children treated per 100 inhabitants per year) . To evaluate the frequency of exposure for each child, all the prescriptions given within a period shorter than 12 days were considered as a single treatment . RESULTS: In the year surveyed, 511,270 antibiotic prescriptions in 219,257 children were identified . In all, 52.9% of children received at least one antibiotic; this percentage decreased with age, ranging from 70.4% in children 1-2 years old to 35.8% in children >11 years old . Fifty-two per cent of inhabitants under the age of 15 years were treated with systemic antibiotics at least once in the year . Cephalosporins were mostly prescribed in the youngest children, while macrolides were most frequently used in children over 6 years old . In all 3.9% of children were treated with topical antibiotics . CONCLUSIONS: This study has shown that paediatric antibiotic prescription rates can be derived from analysis of regional drug and resident databases . High antibiotic usage is shown in the paediatric population of Emilia Romagna, similar to that observed in other regions of Northern Italy . Broad-spectrum antibiotics are predominantly prescribed . Comparison with prescription rates from other countries' paediatric populations suggests there is extensive antibiotic overuse in Italy . This could be associated with selection for and dissemination of antibiotic resistance . Interventions are needed to reduce consumption.

Br J Clin Pharmacol, 2003 Jul, 56(1), 92 - 5
Antibiotic prescribing for children . Too much and too little? Retrospective observational study in primary care; Ekins-Daukes S et al.; AIMS: To investigate the extent of dose-related off-label antibiotic paediatric prescribing in primary care and to identify any potential clinical effects, particularly of lower than recommended dose prescribing . METHODS: Assessment of antibiotic prescribing in 168 396 children aged 0-16 years for the year 1999-2000 from data retrieved from 158 general practices using the national Scottish primary care computer system GPASS . The setting was general practices in Scotland . RESULTS: During the study period at least one course of antibiotics was prescribed to 23 911 children (14.2%) . A total of 4582 (19.2%) children were prescribed an antibiotic dose of less than that recommended in the Summary of Product Characteristics (SPC) . The number of children prescribed an antibiotic at less than recommended dose increased with age from 1154 (11.8%) aged 0-4 years to 1827 (30.0%) in the 12-16 years age group . For each antibiotic, prescribing lower than recommended dose occurred most frequently at those ages at which a dose increase was recommended in the SPC . Antibiotic prescribing at less than the recommended dose was not associated with an increased return rate for further antibiotic prescriptions during the following month, but in 5-11-year-olds was associated with a significant 48% increase in the total number of antibiotic courses prescribed during the study year {mean = 2.09 {95% confidence interval (CI) 1.79, 2.39}vs . 1.41 {95% CI 1.35, 1.47}} . Antibiotic prescribing at doses higher than recommended occurred less frequently (1.6%) and decreased steadily with age . CONCLUSIONS: Off-label prescribing of antibiotics at less than the recommended dose in children is common in primary care and occurs primarily as the result of a failure to increase antibiotic dosage with age in line with SPC recommendations . Adoption of a uniform approach to SPC age banding for antibiotic dose increments would reduce the frequency of dose-related off-label antibiotic prescribing in children and help minimize the potential for the development of antibiotic resistance.

Clin Microbiol Infect, 2003 May, 9(5), 349 - 59
Rapid detection of resistance in Mycobacterium tuberculosis: a review discussing molecular approaches; Garcia de Viedma D; The last few years have seen the development of several molecular designs to search for mutations encoding resistance to antituberculous drugs in Mycobacterium tuberculosis . Most of these are highly efficient for RIF-r detection and are well adapted to search for the most relevant INH-R mutations . In this review, these new molecular approaches are explained and are presented according to the molecular strategies on which they are based . In this sense, techniques based on DNA-sequencing, electrophoresis and hybridization are reviewed and the newer designs based on real-time PCR and microarrays are also included . Molecular methods are sure to transform standard approaches to the issue of resistance in the mycobacteriology laboratory . This will allow laboratories to speed up the performance of resistance assays and provide access to essential information for highly refined detection, follow-up and management of antibiotic resistance in M . tuberculosis.

Aliment Pharmacol Ther, 2003 Jul 1, 18(1), 101 - 7
High-dose rabeprazole-amoxicillin versus rabeprazole-amoxicillin-metronidazole as second-line treatment after failure of the Japanese standard regimen for Helicobacter pylori infection; Isomoto H et al.; BACKGROUND: There is currently no optimal second-line treatment after failure of Helicobacter pylori triple therapy . AIM: To determine effective salvage therapy after failure of lansoprazole-amoxicillin-clarithromycin . METHODS: After failure of lansoprazole-amoxicillin-clarithromycin 123 out-patients were randomized to receive either 2-week rabeprazole (20 mg b.d.) + amoxicillin (1000 mg b.d.) (RA group) or 1-week rabeprazole (10 mg b.d.) + amoxicillin (750 mg twice b.d.) + metronidazole (250 mg b.d.) (RAM group) . Eradication was assessed by the 13C-urea breath test . We also evaluated cytochrome p450 (CYP) 2C19 genotype status, determined by polymerase chain reaction - restriction fragment length polymorphism, and susceptibility to clarithromycin and metronidazole . RESULTS: On an intention-to-treat basis, H . pylori infection cure was achieved in 37 of 63 (59%) patients in the RA group and in 49 of 60 (82%) patients in the RAM group . Per protocol-based eradication rates in the RA and RAM groups were 66% (37/56) and 88% (49/56), respectively . In both analytic sets there were significant differences between the treatment groups (P < 0.01 in each) . Mild adverse events were observed in eight and five patients from the RA and RAM groups, respectively . Genetic predisposition of CYP2C19 and antibiotic resistance did not influence the treatment outcome either regimen . CONCLUSIONS: The rabeprazole + amoxicillin + metronidazole therapy yielded satisfactory results . In contrast, the cure rate in high-dose rabeprazole + amoxicillin was below an acceptable level.

Can J Gastroenterol, 2003 Jun, 17 Suppl B, 58B - 61B
Can the response to eradication therapy in Helicobacter pylori infection be predicted?
Clancy R, Borody T, Ren Z, Pang G.
The failure to eradicate Helicobacter pylori infection with antibiotic therapy has become a major clinical problem, not entirely accounted for by either poor compliance or antibiotic resistance . Recognition that failed eradication is one outcome of the host-parasite relationship focuses attention on impaired host protection as a determinant of nonresponse to antibiotics . A secreted interleukin (IL)-4 whole blood assay was developed to determine whether persistent infection was contributed to by impaired cytokine responses . The blood assay was shown to correlate well with mucosal organ cultures . Significantly lower levels of IL-4 were detected in the whole blood assays in 11 subjects with failed eradication compared with subjects with successful eradication (P<0.05) . This latter group underwent a Th1 to Th0 "switch", which appears to be important to successful eradication . Detection of subjects at risk for failing to eradicate infection with standard combination therapy, by virtue of low secreted IL-4 in whole blood cultures, may have clinical value.

Can J Gastroenterol, 2003 Jun, 17 Suppl B, 53B - 57B
What constitutes failure for Helicobacter pylori eradication therapy?
Malfertheiner P, Peitz U, Treiber G.
Apart from patients with peptic ulcer disease, the use of eradication therapy for Helicobacter pylori infection has been extended to patients with H pylori-positive dyspepsia and conditions at risk for gastric cancer . Standard treatments comprise a proton pump inhibitor plus two antibiotics for at least one week . The main factors leading to treatment failure are noncompliance and antibiotic resistance . Provided the patient is sufficiently informed about possible side effects, discontinuation of the newer triple therapies has become rare . The prevalence of antibiotic resistance varies considerably among different geographic regions, reflecting the habits of prescription of these antibiotics for other indications . Largely, it ranges from 1% to 15% for macrolides, and from 7% to 60% for nitroimidazoles . With nitroimidazole resistance, treatment failure occurs in only less than 50%; with macrolide resistance, by contrast, in more than 50% of the cases . Furthermore, bacterial and host-related factors (Cag A virulence factor, grade of inflammation) contribute to eradication success . In case of treatment failure, post-therapeutic resistance is frequent . Important principles for the choice of second-line treatment are: not to repeat an antibiotic with potential post-therapeutic resistance, and to ensure sufficient acid suppression.

J Indian Med Assoc, 2003 Jan, 101(1), 7, 9 - 10
Cross-sectional study of consumption, compliance and awareness about antibiotic utilisation amongst the urban community in Kolkata; Ray K et al.; A cross-sectional survey was conducted upon 500 respondents, comprising of 250 adults and 250 children who did consume antibiotics in the previous three months . Data were analysed to determine the patterns of utilisation, compliance and awareness regarding antibiotic medication amongst a selected urban population at Kolkata . Antibiotic consumption without prescription was evident amongst 41.2% of adults in comparison to that of 8.4% in children (P < 0.01) . Compliance to daily dosage was observed in 40.8% of adults in comparison to 82.8% in children (P < 0.01) . Awareness pattern regarding antibiotics were reported to have been more in the children group (16.4%) while compared to the adults (8%) . The knowledge regarding antibiotic resistance remained more or less similar in both the groups . The study concludes that high over the counter (OTC) sale and inadequate compliance to antibitotic medication needs further intervention approach towards information, education and communication (IEC) to all concerned.

Microbes Infect, 2003 Jul, 5(8), 705 - 13
Helicobacter in the developing world; Frenck RW Jr et al.; Helicobacter pylori in the developing world is associated with many unique challenges not encountered in an industrialized setting . The 20% prevalence of infection with H . pylori among adolescents in the United States pales in comparison to infection rates exceeding 90% by 5 years of age in parts of the developing world . While H . pylori within the developed world is associated with gastritis, which may lead to peptic ulcer and gastric carcinoma, the infection in the developing world appears to also be linked with chronic diarrhea, malnutrition and growth faltering as well as predisposition to other enteric infections, including typhoid fever and cholera . Once identified, treatment of H . pylori within the developing world presents increased difficulties due to the frequency of antibiotic resistance as well as the frequency of recurrence after successful treatment . Control, and possibly eradication, of H . pylori could likely be achieved through increased standards of living and improved public health, as it has in the industrialized world . However, these measures are distant objectives for most developing countries, making long-term control of the organism dependent on the development and administration of an effective vaccine.

Biotechniques, 2003 Jun, 34(6), 1250 - 8
Positive selection vectors for high-fidelity PCR cloning; Malo MS et al.; The power of PCR cloning of a target DNA fragment is limited by polymerase-induced mutations . While high-fidelity PCR products can be achieved by reducing the number of PCR cycles, the cloning of the very small amount of DNA thus amplified should give only a few recombinant clones (carrying an insert), which would be very difficult to screen from thousands of background false-positive clones generated by all the currently available vectors, including the positive selection vectors . False-positive clones are mostly generated by the recircularization of linearized vectors that have lost some bases at their ends due to digestion with contaminating exonuclease activities present in restriction enzymes, ligases, polymerases, and other reagents . To overcome this problem, two positive selection vectors, pRGR1Ap and pREM5Tc, have been developed, based on the principles of reporter gene reconstruction and regulatory element modulation, respectively . A PCR primer carrying a vector-specific sequence at its 5' end is used in PCR . When the resultant PCR products are ligated to the specific vector, an antibiotic resistance gene is expressed, thus donating positive selection capability to the harboring cells in a specific selection medium . These vectors cloned PCR fragments generated from less than a femtomole quantity of Escherichia coli genomic DNA after only three cycles of PCR amplification, thus greatly reducing the number of recombinant clones containing polymerase-induced mutations.

Health Bull (Edinb), 1999 Mar, 57(2), 128 - 33
Clinical record keeping in patients receiving antibiotics in hospital; Seaton RA et al.; BACKGROUND: Antibiotic prescribing is associated with the growth of antibiotic resistance in micro-organisms . Prescribing therefore needs to be restricted by well-designed antibiotic protocols . Audit of such protocols is only possible if clinical information about infection is properly recorded . OBJECTIVE: To assess the quality of clinical data recording in case notes of patients receiving antibiotics in hospital and to assess the effect of feedback of this information on clinical practice . DESIGN: Four point prevalence surveys . SETTING: Directorate of General Medicine, Dundee Teaching Hospitals NHS Trust . SUBJECTS: Inpatients receiving antibiotics . INTERVENTION: Feedback of results from the first two surveys to clinicians . MAIN OUTCOME MEASURES: Proportion of patients in the Medical Directorate with the recording of an antibiotic indication and the temperature in the case notes before, immediately after, and three months after, feedback of survey results . RESULTS: In each survey one third of patients were receiving antibiotics . The antibiotic indication was recorded in 64% of patients prior to feedback and in 86.5% after feedback (p = 0.004) . Three months later this had fallen to 74% . Temperature recording improved between 2nd and 3rd surveys (57% to 86.1%, p < 0.001) and declined to 61% in the 4th survey (p < 0.001) . Respiratory infection was the commonest recorded indication for an antibiotic; recording of this indication improved between the 2nd and 3rd surveys but declined by the 4th survey . CONCLUSIONS: Audit and feedback of information about clinical record keeping improves standards but improvements are not sustained . Clinical record keeping must be improved so compliance with antibiotic protocols can be properly audited . An "antibiotic sticker" in addition to immediate concurrent feedback may improve such deficiencies.

Genomics, 2003 Jul, 82(1), 68 - 77
Targeted modification of a human beta-globin locus BAC clone using GET Recombination and an I-Scei counterselection cassette; Jamsai D et al.; There is a need for better approaches to allow precise engineering of large genomic BAC DNA fragments, to facilitate the use of intact genomic loci for therapeutic and biotechnology applications . We report an efficient method to insert any modification in any genomic locus, using a human beta-globin locus BAC clone as a model system . The modifications can range from single base changes to large insertions or deletions and leave no operational sequences . A counterselection cassette, consisting of an inducible I-SceI gene, its recognition site, and an antibiotic resistance gene, is inserted into the targeted region using GET Recombination . A PCR fragment carrying the modification but no selectable marker replaces the counterselection cassette in a second round of GET Recombination . The unique I-SceI site in the counterselection cassette is cut by I-SceI endonuclease, strongly selecting against nonrecombinant clones and yielding up to 30% correct recombinants.

Commun Dis Intell, 2003, 27 Suppl, S39 - 41
Antibiotic prescribing for upper respiratory-tract infections in primary care; Patterson CA et al.; The use and overuse of antibiotics in humans is a major contributor to the selection of antibiotic resistance organisms . Recent evidence has shown that primary care prescribing selects for resistances of clinical importance . The National Prescribing Service runs both educational and audit activities . The latter provide some insight into general practice attitudes toward antibiotic prescribing.

Nat Rev Genet, 2003 Jun, 4(6), 432 - 41
Exploiting genomics, genetics and chemistry to combat antibiotic resistance; Hughes D; To address the worsening problem of antibiotic-resistant bacteria there is an urgent need to develop new antibiotics . Comparative genomics and molecular genetics are being applied to produce lists of essential new targets for compound screening programmes . Combinatorial chemistry and structural biology are being applied to rapidly explore and optimize the interactions between lead compounds and their biological targets . Several compounds that have been identified from target-based screens are now in development, but technical and economic constraints might result in a trickle, rather than a flood, of new antibiotics onto the market in the near future.

Aliment Pharmacol Ther, 2003 May 1, 17(9), 1137 - 43
Triple vs . quadruple therapy for treating Helicobacter pylori infection: a meta-analysis; Gene E et al.; BACKGROUND: Triple therapy (proton pump inhibitor, clarithromycin and amoxicillin or an imidazole) is the first-line treatment for Helicobacter pylori infection . However, the effectiveness of triple therapy is decreasing due to the increase in antibiotic resistance . Quadruple therapy (proton pump inhibitor, tetracycline, metronidazole and a bismuth salt) is a very effective regimen even in areas of high prevalence of antibiotic resistance, and may be an alternative first-line treatment . AIM: To compare triple vs . quadruple therapy for the first-line treatment of H . pylori infection . METHODS: An extensive literature search was performed to identify randomized trials comparing triple vs . quadruple therapy . Selected trials were included in a meta-analysis using Review Manager 4.1 . RESULTS: Four studies met the inclusion criteria . Eradication rates with quadruple therapy were slightly higher in both the intention-to-treat (81% vs . 78%; odds ratio, 0.83; 95% confidence interval, 0.61-1.14) and per protocol (88% vs . 85%; odds ratio, 0.81; 95% confidence interval, 0.55-1.20) analysis, although the differences were not statistically significant . Nor were there significant differences in compliance or adverse effects between the therapies . CONCLUSION: Triple and quadruple therapies seem to be roughly equivalent in terms of effectiveness, compliance and side-effects profile when administered as first-line treatment for H . pylori infection.

J Toxicol Environ Health B Crit Rev, 2003 May-Jun, 6(3), 211 - 25
Potential adverse health effects of genetically modified crops; Bakshi A; Genetically modified crops have the potential to eliminate hunger and starvation in millions of people, especially in developing countries because the genetic modification can produce large amounts of foods that are more nutritious . Large quantities are produced because genetically modified crops are more resistant to pests and drought . They also contain greater amounts of nutrients, such as proteins and vitamins . However, there are concerns about the safety of genetically modified crops . The concerns are that they may contain allergenic substances due to introduction of new genes into crops . Another concern is that genetic engineering often involves the use of antibiotic-resistance genes as "selectable markers" and this could lead to production of antibiotic-resistant bacterial strains that are resistant to available antibiotics . This would create a serious public health problem . The genetically modified crops might contain other toxic substances (such as enhanced amounts of heavy metals) and the crops might not be "substantially equivalent" in genome, proteome, and metabolome compared with unmodified crops . Another concern is that genetically modified crops may be less nutritious; for example, they might contain lower amounts of phytoestrogens, which protect against heart disease and cancer . The review of available literature indicates that the genetically modified crops available in the market that are intended for human consumption are generally safe; their consumption is not associated with serious health problems . However, because of potential for exposure of a large segment of human population to genetically modified foods, more research is needed to ensure that the genetically modified foods are safe for human consumption.

Folia Microbiol (Praha), 2003, 48(1), 83 - 9
Competition among Bradyrhizobium strains for nodulation of green gram (Vigna radiata): use of dark-nodule strain; Sindhu SS et al.; The competitiveness of dual-strain inoculum of Bradyrhizobium strains S24 and GR4 was demonstrated for nodulation of green gram (Vigna radiata) . Strain S24 formed pink nodules, GR4 produced visually distinguishable dark-brown nodules . When a mixture of these Bradyrhizobium strains was applied as inoculum, nodules of both pink and dark-brown types were formed on the same root . The strain GR4, which was less competitive than strain S24, was mutagenized with N-methyl-N'-nitro-N-nitrosoguanidine to obtain pigment-diverse mutants and six selected mutants were screened for symbiotic parameters . One mutant produced pink nodules and appreciably increased plant dry mass . The competitive ability of this mutant lacking brown pigment was compared with that of strain S24 by using antibiotic resistance markers; it showed increased nodulation competitiveness than its parent strain GR4 . The dark-brown nodule-phenotype could be useful in evaluating nodulation competitiveness of "cowpea miscellany" bradyrhizobia in soil where dark-brown nodule-forming strains are not indigenous.

Commun Dis Public Health, 2003 Apr, 6(1), 51 - 4
Survey of general practitioner satisfaction with a district communicable disease control service; Lamden K et al.; General practitioners (GPs) have an important role in disease surveillance and control, and are therefore a key target audience for a district communicable disease control (CDC) service . The feedback of surveillance information, and the provision of policies, guidance and specialist advice, are essential functions of a CDC service . This survey was undertaken to assess how well a CDC service was meeting the needs of GPs . Satisfaction with the service was assessed using a questionnaire survey of 166 GPs . The response rate was 42% . GPs reported a high level of overall satisfaction . The service was valued and it influenced GP practice . The service was valued more for responses to requests for information and assistance than for communicable disease control policies . The highest satisfaction was expressed for communication and for immunisation advice . Several areas of need were identified, including the need for surveillance information on antibiotic resistance and for sexually transmitted infections . The survey tool is recommended as a way of auditing standards for Good Public Health Practice, as identified by the Faculty of Public Health Medicine.

Dtsch Med Wochenschr, 2003 May 9, 128(19), 1038 - 41
{Nocardia cyriacigeorgici: First report of invasive human infection}; Fux C et al.; BACKGROUND: Diagnostic laboratories increasingly offer bacterial identification to the species level . The 17 nocardia species known to date differ in their clinical presentation, antibiotic resistance patterns and geographic distribution . The discovery of a new species with pathogenicity for humans calls for the characterization of its clinical and epidemiological properties . PATIENTS AND METHODS: Nocardia isolated from multifocal brain abscesses of an immunocompromised patient were further identified by the analysis of their cellular fatty acids and sequencing of the 16S ribosomal DNA . Quantitative antibiotic resistance testing was performed with E-tests . RESULTS: The 16S ribosomal DNA analysis showed a 99 % homology to Nocardia cyriacigeorgici . This is the first report of this species as an invasive human pathogen . N . cyriacigeorgici was found susceptible for meropenem, amikacin, ceftriaxon and cotrimoxazole . The combination of surgical drainage and antibiotic treatment for 13 months was curative . CONCLUSIONS: N . cyriacigeorgici has the potential to cause invasive infections at least in immunocompromised patients . Comparing clinical and in vitro characteristics with N . asteroides, the main causative agent of nocardial infections in Europe, we found no clinically relevant differences.

J Vet Diagn Invest, 2003 May, 15(3), 242 - 52
Isolation and association of Escherichia coli AIDA-I/STb, rather than EAST1 pathotype, with diarrhea in piglets and antibiotic sensitivity of isolates; Ngeleka M et al.; To identify emerging Escherichia coli that have the potential to cause diarrhea in pigs, the prevalence of E . coli pathotypes was determined among 170 and 120 isolates from diarrheic and nondiarrheic piglets, respectively . The isolates were tested for F4, F5, F6, F18, and F41 fimbriae, for E . coli attaching and effacing (EAE), porcine attaching and effacing-associated (Paa), and adhesin involved in diffuse adherence (AIDA-I) factors, for LT, STa, STb, and enteroaggregative heat-stable (EAST1) enterotoxins, and for Shiga toxins (Stxl, Stx2, and Stx2e), using DNA hybridization and polymerase chain reaction . All isolates were O-serotyped and tested for antibiotic resistance against 10 drugs . Seventeen different pathotypes, accounting for 40.0% of the isolates, were recovered from diarrheic piglets . The main pathotypes included EAST1 (13.5%), F4/LT/STb/EAST1 (6.5%), AIDA-I/STb/EAST1 (4.1%), F5/STa (2.9%), EAE/EAST1 (2.9%), and AIDA-I/F18 (2.3%) . Only 3 pathotypes, EAE (11.7%), EAST1 (10.8%), and EAE/EAST1 (3.3%), were recovered from nondiarrheic piglets . Paa factor was detected in 8.8% and 7.5% of isolates from diarrheic and nondiarrheic piglets, respectively, and always was associated with other virulence determinants . Overall, 22.9% of isolates from diarrheic piglets appeared to be enteropathogens: enterotoxigenic E . coli (11.7%), enteropathogenic E . coli (3.5%), and E . coli isolates (3.0%) for which none of the above adherence factors was detected . Pathotypes AIDA-I/STb/EAST1 and AIDA-I/STb were isolated only from diarrheic piglets and accounted for 4.7% of isolates . Strains of these pathotypes induced diarrhea when inoculated into newborn colostrum-deprived pigs, in contrast to an isolate positive only for EAST1, which did not induce diarrhea . Antibiotic sensitivity test showed that isolates of the AIDA-I/STb/EAST1 and AIDA-I/STb pathotypes were the only strains sensitive to enrofloxacin, gentamicin, neomycin, and trimethoprim-sulfamethoxazole . This study showed that at least 20.5% of isolates from diarrheic piglets appeared to be associated with AIDA-I/STb pathotype and that EAST1 pathotype is probably not an important marker for diarrhea in piglets.

Clin Geriatr Med, 2003 Feb, 19(1), 101 - 20
Community-acquired pneumonia in elderly patients; Niederman MS et al.; CAP in elderly patients carries a significant economic and clinical burden and will be more commonly encountered in the future as the US population ages . Diagnosis may be obscured by a nonclassic presentation in an elderly patient, and the clinician needs to be especially suspicious of pneumonia whenever the clinical status of an elderly patient deteriorates . The single most important clinical decision is the site of care; this determination is not always based on clinical factors but also on social factors . Severity assessment is key to stratifying appropriate therapy and to predicting outcome . Timely and appropriate empiric therapy enhances the likelihood of a good clinical outcome, although clinical resolution may be more delayed than in younger patients . Newly emerging patterns of antibiotic resistance have altered recent guidelines for CAP treatment; DRSP is now a consideration in elderly patients because an age older than 65 years is a well-described risk factor for infection with this organism . Prevention should always be implemented, with a focus on pneumococcal and influenza vaccination.

Plasmid, 2003 Mar, 49(2), 152 - 9
Analysis of pCU1 replication origins: dependence of oriS on the plasmid-encoded replication initiation protein RepA; Zoueva OP et al.; The broad-host-range replicon of the plasmid pCU1 has three origins of vegetative replication called oriB, oriS, and oriV . In the multi-origin replicon, individual origins can distinguish among replication factors provided by the host . It has been found that during replication in Escherichia coli polA(-) host, oriS was the only active origin of a mutant pCU1 derivative bearing a mutation in the gene encoding replication initiation protein RepA . To further investigate the capacity of oriS to function in an E . coli polA(-) host we constructed a number of clones of the basic replicon of pCU1 containing oriS as the only replication origin . An oriS construct created with pUC18 could transform the polA(-) strain when RepA was supplied in trans . When the oriS region (between nucleotides 290 and 832) was ligated to an antibiotic resistance Omega fragment, the construct could be recovered as a plasmid from polA(+) strain if functional RepA was provided in trans . Our results therefore indicate that the basic replicon of pCU1, containing oriS as the sole origin, does require RepA to initiate plasmid replication in E . coli

J Antibiot (Tokyo), 2003 Feb, 56(2), 135 - 42
Functional annotation of putative aminoglycoside antibiotic modifying proteins in Mycobacterium tuberculosis H37Rv; Draker KA et al.; The growing availability of sequences of bacterial genomes has revealed a number of open reading frames predicted by sequence alignment to encode antibiotic resistance proteins . The presence of these putative resistance genes within bacterial genomes raises important questions regarding potential reservoirs of resistance elements and their evolution . Here we examine four gene products encoding predicted aminoglycoside-aminocyclitol antibiotic modifying enzymes, two phosphotransferases and two acetyltransferases, derived from analysis of the genome sequence of Mycobacterium tuberculosis strain H37Rv with the goal of assigning biochemical function by purification of each protein and characterization of their ability to modify aminoglycoside antibiotics . Only one of these enzymes, the previously characterized aminoglycoside acetyltransferase AAC(2')-Ic, displayed compelling aminoglycoside modifying activity . While the putative phosphotransferase encoded by the Rv3225c gene did display low levels of aminoglycoside kinase activity, the predicted kinase encoded by the Rv3817 gene lacked any such activity . A potential aminoglycoside 6'-acetyltransferase, encoded by the Rv1347c gene, did not show antibiotic acylation activity but did demonstrate selective thioesterase activity with numerous acyl-CoAs . This activity, together with the genomic environment of the Rv1347c gene in a likely polyketide synthesis cluster, suggests a role for this protein in secondary metabolism and not in antibiotic modification . It was thus shown that only one of four putative aminoglycosides modifying enzymes derived from the whole genome sequencing of M . tuberculosis H37Rv showed sufficient predicted enzyme activity to be annotated as an aminoglycoside resistance element . This study demonstrates the necessity of biochemical annotation methods as a follow up to in silico sequence alignment-based methods of assigning gene product function.

Poult Sci, 2003 Apr, 82(4), 622 - 6
Monitoring and identifying antibiotic resistance mechanisms in bacteria; Roe MT et al.; Sub-therapeutic administration of antibiotics to animals is under intense scrutiny because they contribute to the dissemination of antibiotic-resistant bacteria into the food chain . Studies suggest that there is a link between the agricultural use of antibiotics and antibiotic-resistant human infections . Antibiotic-resistant organisms from animal and human wastes reenter the human and animal populations through a number of pathways including natural waters, irrigation water, drinking water, and vegetables and foods . Antibiotic usage in the United States for animal production (disease prevention and growth promotion) is estimated to be 18 million pounds annually . As much as 25 to 75% of the antibiotics administered to feedlot animals are excreted unaltered in feces . Because about 180 million dry tons of livestock and poultry waste is generated annually in the United States, it is not surprising that animal-derived antibiotic-resistant organisms are found contaminating groundwater, surface water, and food crops . It is extremely important to clearly understand the molecular mechanisms that could potentially cause lateral or horizontal gene transfer of antibiotic resistance genes among bacteria . Once the mechanisms and magnitude of resistance gene transfer are clearly understood and quantified, strategies can be instituted to reduce the potential for dissemination of these genes.

Genetics, 2003 Apr, 163(4), 1237 - 41
Experimental prediction of the natural evolution of antibiotic resistance; Barlow M et al.; The TEM family of beta-lactamases has evolved to confer resistance to most of the beta-lactam antibiotics, but not to cefepime . To determine whether the TEM beta-lactamases have the potential to evolve cefepime resistance, we evolved the ancestral TEM allele, TEM-1, in vitro and selected for cefepime resistance . After four rounds of mutagenesis and selection for increased cefepime resistance each of eight independent populations reached a level equivalent to clinical resistance . All eight evolved alleles increased the level of cefepime resistance by a factor of at least 32, and the best allele improved by a factor of 512 . Sequencing showed that alleles contained from two to six amino acid substitutions, many of which were shared among alleles, and that the best allele contained only three substitutions.

Acta Vet Hung, 2003, 51(1), 29 - 44
Spontaneous antibiotic resistance mutation associated pleiotropic changes in Escherichia coli O157:H7; Toth I et al.; Besides the well-known O157:H7 clone causing enterohaemorrhagic colitis and haemolytic uraemic syndrome in Europe, Japan and North America, the number of Escherichia coli isolates with non-motile (NM) phenotype has considerably increased . We supposed that spontaneous antibiotic resistance mutation could cause this phenotypic change . To model our hypothesis we isolated rifampicin--(Rif) and ampicillin--(Amp) resistant mutants from E . coli O157:H7 prototype strains 7785 and EDL933 . Among Rifr mutants we could isolate strains with no or reduced motility, while the Ampr mutants became hypermotile . The biochemical profile of the mutants had not changed but phage sensitivity and generation time of the mutants were altered . Among the representative strains we did not find polymorphism with Southern blot analysis and no polymorphism was found in the fliC gene of the mutants . The described characteristics have proven to be stable . In a mice virulence assay by intravenous infections the virulence of the derivatives was also found to be changed . In summary, we found that the antibiotic-resistant phenotype in E . coli O157:H7 was coexpressed with several other phenotypic changes including motility and virulence . It can be assumed that expression of the involved phenotypes may be under the influence of a common regulatory cascade . Further work is needed to identify the components and mechanism of this regulatory system.

J Ind Microbiol Biotechnol, 2003 Apr, 30(4), 195 - 204 Epub 2003 Apr 08.
Development of cloning vectors and transformation methods for Amycolatopsis; Dhingra G et al.; The genus Amycolatopsis is of industrial importance, as its species are known to produce commercial antibiotics . It belongs to the family Pseudonocardiaceae and has an eventful taxonomic history . Initially strains were identified as Streptomyces, then later as Nocardia . However, based on biochemical, morphological and molecular features, the genus Amycolatopsis, containing seventeen species, was created . The development of molecular genetic techniques for this group has been slow . The scarcity of molecular genetic tools including stable plasmids, antibiotic resistance markers, transposons, reporter genes, cloning vectors, and high efficiency transformation protocols has made progress slow, but efforts in the past decade have led to the development of cloning vectors and transformation methods for these organisms . Some of the cloning vectors have broad host range (pRL series) whereas others have limited host range (pMEA300 and pMEA100) . The cloning vector pMEA300 has been completely sequenced, while only the minimal replicon (pA- rep) has been sequenced from pRL plasmids . Direct transformation of mycelia and electroporation are the most widely applicable methods for transforming species of Amycolatopsis . Conjugational transfer from Escherichia coli has been reported only in the species A . japonicum, and gene disruption and replacements using homologous recombination are now possible in some strains.

WMJ, 2001, 100(8), 55 - 9
Feasibility survey for a study on selective antibiotic management of acute otitis media; Yen K et al.; OBJECTIVE: To assess the views of community physicians on management of acute otitis media (AOM) without antibiotics and their willingness to support research on this issue . METHODS: Community physicians who admit to a children's hospital were surveyed using a questionnaire containing questions on current issues in AOM management and their willingness to support research on management of AOM without antibiotics . RESULTS: Fifty-two percent of the surveys were returned . All respondents report concern about antibiotic resistant bacteria . Sixty-three percent treat otitis media with effusion with antibiotics and 68% give prophylactic antibiotics for recurrent otitis media . Thirty-five percent consider management of AOM without antibiotics as a possible alternative management practice . Forty-five percent of the respondents are willing to support research in this practice and 46% need more information . CONCLUSIONS: Surveyed physicians indicate concern about antibiotic resistance and a willingness to support further research on the initial management of acute otitis media without antibiotics.

Mol Biol Evol, 2003 Apr, 20(4), 653 - 9 Epub 2003 Apr 02.
Determining the limits of the evolutionary potential of an antibiotic resistance gene; Salipante SJ et al.; The AAC(6') enzymes inactivate aminoglycoside antibiotics by acetylating their substrates at the 6' position . Based on functional similarity and size similarity, the AAC(6') enzymes have been considered to be members of a single family . Our phylogenetic analysis shows that the AAC(6') enzymes instead belong to three unrelated families that we now designate as {A}, {B}, and {C} and that aminoglycoside acetylation at the 6' position has evolved independently at least three times . AAC(6')-Iaa is a typical member of the {A} family in that it acetylates tobramycin, kanamycin, and amikacin effectively but acetylates gentamicin ineffectively . The potential of the aac(6')-Iaa gene to increase resistance to tobramycin, kanamycin, or amikacin or to acquire resistance to gentamicin was assessed by in vitro evolution . Libraries of PCR mutagenized alleles were screened for increased resistance to tobramycin, kanamycin, and amikacin, but no isolates that conferred more resistance than the wild-type gene were recovered . The library sizes were sufficient to conclude with 99.9% confidence that no single amino acid substitution or combination of two amino acid substitutions in aac(6')-Iaa is capable of increasing resistance to the antibiotics used . It is therefore very unlikely that aac(6')-Iaa of S . typhimurium LT2 has the potential to evolve increased aminoglycoside resistance in nature . The practical implications of being able to determine the evolutionary limits for other antibiotic resistance genes are discussed.

Biotechnol Prog, 2003 Mar-Apr, 19(2), 612 - 23
Gene array-based identification of changes that contribute to ethanol tolerance in ethanologenic Escherichia coli: comparison of KO11 (parent) to LY01 (resistant mutant); Gonzalez R et al.; Escherichia coli KO11 (parent) and LY01 (mutant) have been engineered for the production of ethanol . Gene arrays were used to identify expression changes that occurred in the mutant, LY01, during directed evolution to improve ethanol tolerance (defined as extent of growth in the presence of added ethanol) . Expression levels for 205 (5%) of the ORFs were found to differ significantly (p < 0.10) between KO11 and LY01 under each of six different growth conditions (p < 0.000001) . Statistical evaluation of differentially expressed genes according to various classification schemes identified physiological areas of importance . A large fraction of differentially expressed ORFs were globally regulated, leading to the discovery of a nonfunctional fnr gene in strain LY01 . In agreement with a putative role for FNR in alcohol tolerance, increasing the copy number of fnr(+) in KO11(pGS196) decreased ethanol tolerance but had no effect on growth in the absence of ethanol . Other differences in gene expression provided additional clues that permitted experimentation . Tolerance appears to involve increased metabolism of glycine (higher expression of gcv genes) and increased production of betaine (higher expression of betIBA and betT encoding betaine synthesis from choline and choline uptake, respectively) . Addition of glycine (10 mM) increased ethanol tolerance in KO11 but had no effect in the absence of ethanol . Addition of betaine (10 mM) increased ethanol tolerance by over 2-fold in both LY01 and KO11 but had no effect on growth in the absence of ethanol . Both glycine and betaine can serve as protective osmolytes, and this may be the basis of their beneficial action . In addition, the marAB genes encoding multiple antibiotic resistance proteins were expressed at higher levels in LY01 as compared to KO11 . Interestingly, overexpression of marAB in KO11 made this strain more ethanol-sensitive . Overexpression of marAB in LY01 had no effect on ethanol tolerance . Increased expression of genes encoding serine uptake (sdaC) and serine deamination (sdaB) also appear beneficial for LY01 . Addition of serine increased the growth of LY01 in the presence and absence of ethanol but had no effect on KO11 . Changes in the expression of several genes concerned with the synthesis of the cell envelope components were also noted, which may contribute to increased ethanol tolerance.

Tex Med, 2003 Mar, 99(3), 66 - 9
Safety of foods derived from genetically modified plants; Thomas JA; Biopharmaceuticals have been available for clinical use for nearly three decades, but foods derived from agribiotechnology have been available for just under a decade . Controversy surrounding foods from genetically modified (GM) plants has focused primarily upon their allergenicity, with lesser concerns about antibiotic resistance genes . Concerns are related to possible environmental impacts on non-human species, including effects on non-target species (e.g., butterflies) and on the development of so-called "super weeds." Food allergies are no more prevalent in foods from GM plants than in conventional foods . Further, the use of antibiotics in the development of GM plants does not pose a significant risk to the human population . Foods from the current GM plant products have been shown not to pose any detrimental effects to humans, and, in fact, nutritionally enhanced products are being developed . GM foods are subjected globally to intense regulatory scrutiny, and extensive data have been provided consistently to regulatory agencies in the United States on a voluntary basis, with mandatory reporting of data soon to be in force . Existing environmental concerns appear to be unjustified on the basis of existing data and experience.

Ir Med J, 2003 Feb, 96(2), 43 - 4, 46
Antibiotic misuse in the community--a contributor to resistance?
Carey B, Cryan B.
The problem of antibiotic resistance is associated with the indiscriminate usage of antibiotics . Efforts have been directed at encouraging the rational use of these drugs to reduce the volume of antibiotic consumption and decrease resistance rates . There is evidence to suggest that the misuse of antibiotics by patients may also contribute to the problem . We describe a survey of a random selection of patients attending a General Practitioners' surgery over a six week period in an effort to estimate the level of non-compliance to antibiotic therapy in the community . The results suggest that there may be a significant level of antibiotic misuse prevalent in the local community . We discuss these results and present evidence in the literature suggesting how antibiotic misuse may affect resistance in the community . The factors affecting patient compliance to therapy are outlined along with suggested measures to improve compliance among patients.

J Bacteriol, 2003 Apr, 185(8), 2667 - 72
Global analysis of genes regulated by EvgA of the two-component regulatory system in Escherichia coli; Nishino K et al.; The response regulator EvgA controls expression of multiple genes conferring antibiotic resistance in Escherichia coli (K . Nishino and A . Yamaguchi, J . Bacteriol . 184:2319-2323, 2002) . To understand the whole picture of EvgA regulation, DNA macroarray analysis of the effect of EvgA overproduction was performed . EvgA activated genes related to acid resistance, osmotic adaptation, and drug resistance.

Biochem Soc Trans, 2003 Apr, 31(2), 299 - 306
Genes in food--why the furore?
Dixon B.
Although unprecedented and perhaps unique in its irrationality, the recent furore over genetically modified (GM) food holds extremely important lessons for scientists . Some sections of the media undoubtedly bear a heavy responsibility for giving the expression 'GM' threatening connotations that are quite unwarranted . However, influential contributions to the hysteria have come from a surprising range of other sources, including some scientists . The research community has failed in its responsibility to society in three ways . Firstly, plant scientists did not appreciate that certain techniques (such as the use of antibiotic resistance genes as markers during plant transformation) would inevitably provoke public consternation . As a result, they took no steps to address such concerns . Secondly, researchers overlooked, minimized or in some cases simply dismissed the significance of public fears that they were 'interfering with Nature' or 'playing God' . Thirdly, plant breeders apparently saw no need to take pro-active measures with regard to the media and public in placing potential environmental and nutritional benefits of GM crops on the agenda in a positive fashion . Partly because of this failure, GM food is now firmly established in the public mind as wholly objectionable . One measure of how far we have travelled down that road is that it hardly matters any more whether objections are based on alleged environmental risks of cultivating GM crops or alleged toxicological hazards of eating them . 'Genetically modified organism', like 'radioactivity', has become an odious, generic shibboleth . Given that millions of people throughout the world are already benefiting from pharmaceuticals made by GM organisms, this is bizarre.

Protein Eng, 2003 Jan, 16(1), 27 - 35
Evolutionary engineering of a beta-Lactamase activity on a D-Ala D-Ala transpeptidase fold; Peimbert M et al.; The beta-Lactamase hydrolytic activity has arisen several times from DD-transpeptidases . We have been able to replicate the evolutionary process of beta-Lactamase activity emergence on a PBP2X DD-transpeptidase . Some of the most interesting changes, like modifying the catalytic properties of an enzyme, may require several mutations in concert; therefore it is essential to explore efficiently sequence space by generating the right diversity . We designed a biased combinatorial library in which biochemical and structural information were incorporated by site directed mutagenesis on relevant residues and then subjected to random mutagenesis to allow for mutations in unforeseen positions . We isolated mutants from this library conferring 10-fold higher cefotaxime resistance levels than the background wild-type through mutations exclusively in the coding sequence . We demonstrate that only three substitutions in the DD-transpeptidase active site, two produced by the directed and one by the random mutagenesis, are sufficient to acquire this activity . The purified product of one mutant (MutE) had a 10(5)-fold increase in cefotaxime deacylation rate allowing it to hydrolyze beta-Lactams yet it has apparently conserved DD-peptidase activity . This work is the first to show a possible evolutionary intermediate between a beta-Lactamase and a DD-transpeptidase necessary for the development of antibiotic resistance.

Arch Inst Pasteur Madagascar, 2002, 68(1-2), 41 - 3
{Tuberculosis in children less than 11 years old: primary resistance and dominant genetic variants of Mycobacterium tuberculosis in Antananarivo}; Rasolofo Razanamparany V et al.; Tuberculosis during childhood is often due to Mycobacterium tuberculosis primo-infection . Tuberculosis is highly prevalent in Madagascar and most people are infected during childhood . Our objectives were to evaluate the primary resistance of M . tuberculosis and to determine the genotypes responsible for recent infection in the population . Thus we studied 142 isolated strains from 97 children (66 with pulmonary tuberculosis and 31 with extra-pulmonary tuberculosis) recruited in different health centers in Antananarivo from 1997 to 2000 . Excepting one strain resistant to isoniazide, all strains were susceptible to the four antibiotics (streptomycin, isoniazid, ryfampicin and ethambutol) . This result confirms the low rate of primary resistance reported during the two surveys in 1994-1995 and 1999-2000 . 67 strains of 1997-2000 were typed with the genetic marker IS6110, 44 has been assigned to 13 clusters containing each 2 to 8 similar strains . Some IS6110 clusters have already been reported in 1994-1995 . Some genotypes observed in 1994-1995 seemed to have disappeared in 1997-2000 . (As the rate of the frequency of some genetic variants according to the period are more likely due to a difference in strain virulence) . Since there is minimal antibiotic resistance versus M . tuberculosis in Madagascar, one can not explain the appearance or disappearance of certain variants because of drug resistance . Rather, this is due to the virulence of the various M . tuberculosis strains.

J Mol Biol, 2003 Mar 21, 327(2), 491 - 506
The crystal structure of aminoglycoside-3'-phosphotransferase-IIa, an enzyme responsible for antibiotic resistance; Nurizzo D et al.; A major factor in the emergence of antibiotic resistance is the existence of enzymes that chemically modify common antibiotics . The genes for these enzymes are commonly carried on mobile genetic elements, facilitating their spread . One such class of enzymes is the aminoglycoside phosphotransferase (APH) family, which uses ATP-mediated phosphate transfer to chemically modify and inactivate aminoglycoside antibiotics such as streptomycin and kanamycin . As part of a program to define the molecular basis for aminoglycoside recognition and inactivation by such enzymes, we have determined the high resolution (2.1A) crystal structure of aminoglycoside-3'-phosphotransferase-IIa (APH(3')-IIa) in complex with kanamycin . The structure was solved by molecular replacement using multiple models derived from the related aminoglycoside-3'-phosphotransferase-III enzyme (APH(3')-III), and refined to an R factor of 0.206 (R(free) 0.238) . The bound kanamycin molecule is very well defined and occupies a highly negatively charged cleft formed by the C-terminal domain of the enzyme . Adjacent to this is the binding site for ATP, which can be modeled on the basis of nucleotide complexes of APH(3')-III; only one change is apparent with a loop, residues 28-34, in a position where it could fold over an incoming nucleotide . The three rings of the kanamycin occupy distinct sub-pockets in which a highly acidic loop, residues 151-166, and the C-terminal residues 260-264 play important parts in recognition . The A ring, the site of phosphoryl transfer, is adjacent to the catalytic base Asp190 . These results give new information on the basis of aminoglycoside recognition, and on the relationship between this phosphotransferase family and the protein kinases.

FEMS Microbiol Lett, 2003 Feb 28, 219(2), 261 - 8
Production of unmarked mutations in mycobacteria using site-specific recombination; Malaga W et al.; Gene disruption experiments play an important role in the functional characterization of genes in mycobacteria and rely mostly on the use of one or two antibiotic resistance markers . We have developed a system for mycobacteria which features both the advantages of the use of antibiotic resistance markers for gene disruption experiments and the ability to efficiently rescue the marker leaving an unmarked mutation on the chromosome . This new genetic tool relies on the transposon gammadelta site-specific recombination system . A res-OmegaKm-res cassette was used to generate an insertional mutation by allelic exchange both in Mycobacterium smegmatis and Mycobacterium bovis BCG . Upon expression in the mutated strains of tnpR, the transposon gammadelta resolvase gene, res-OmegaKm-res, was excised efficiently leaving behind a single res sequence at the mutated locus . A plasmid was engineered allowing expression of tnpR from an easily curable mycobacterial vector . This system will be useful for simple construction of unmarked mutations or repeated use of the same antibiotic marker to generate multiple mutants.

Can Fam Physician, 2003 Feb, 49, 168 - 73
Saline nasal irrigation: Its role as an adjunct treatment; Papsin B et al.; OBJECTIVE: To review clinical evidence on the efficacy of saline nasal irrigation for treatment of sinonasal conditions and to explore its potential benefits . QUALITY OF EVIDENCE: Clinical trials, reviews, and treatment guidelines discussing nasal irrigation were obtained through a MEDLINE search from January 1980 to December 2001 . Most trials were small and some were not controlled; evidence, therefore, is level II, or fair . MAIN MESSAGE: Flushing the nasal cavity with saline solution promotes mucociliary clearance by moisturizing the nasal cavity and by removing encrusted material . The procedure has been used safely for both adults and children, and has no documented serious adverse effects . Patients treated with nasal irrigation rely less on other medications and make fewer visits to physicians . Treatment guidelines in both Canada and the United States now advocate use of nasal irrigation for all causes of rhinosinusitis and for postoperative cleaning of the nasal cavity . CONCLUSION: Nasal irrigation is a simple, inexpensive treatment that relieves the symptoms of a variety of sinus and nasal conditions, reduces use of medical resources, and could help minimize antibiotic resistance.

Eksp Klin Gastroenterol, 2002, (5), 21 - 3, 126
{Problem of antibiotic-resistant Helicobacter pylori and ways of overcoming it}; Bardakhch'ian EA et al.; Application of non-drug treatment methods along with the basic drug therapy can promote the overcoming of antibiotic resistance and contribute to the eradication of H . pylori.

Med Hypotheses, 2003 Apr, 60(4), 484 - 8
Bacterial multicellularity as a possible source of antibiotic resistance; Krasovec R et al.; Knowledge about survival of micro-organisms in stressful situations not only influences the evolutionary theory in a fundamental way, but bears an extraordinary importance in finding a global solution to a very concrete urgent problem of mankind, namely bacterial resistance to antibiotics . Recent in vitro experiments demonstrate that the adaptive mutation process involving transient hypermutators could be one of the most important mechanisms whereby bacterial cells achieve the antibiotic resistance . An effective response of the mutation rates to specific selective conditions and an increasing number of conclusive evidence that bacterial cells are indeed communicative and co-operative organisms lead us to a hypothesis that the emergence of the antibiotic resistant mutants through the so-called adaptive mutation is deeply connected with the multicellular organisation of bacterial cells .

Br J Health Psychol, 2001 Nov, 6(Part 4), 347 - 360
Salient beliefs and intentions to prescribe antibiotics for patients with a sore throat; Walker AE et al.; OBJECTIVES: General practitioners (GPs) in the UK continue to prescribe antibiotics for patients with sore throats despite evidence that they are ineffective and can contribute to the growth of antibiotic resistance in the population . This study uses the theory of planned behaviour (TPB) to investigate the strength of intention to prescribe antibiotics, and to identify the salient beliefs associated with this intention . DESIGN: Cross-sectional study testing hypotheses derived from the TPB . METHOD: A 66-item postal questionnaire was distributed to a random sample of GPs in one NHS region (N = 185) . The questionnaire included measures of intention to prescribe antibiotics, attitude, behavioural beliefs and evaluations, normative beliefs and evaluations, perceived behavioural control, control beliefs, and past prescribing . RESULTS: Two-thirds of the GPs returned complete questionnaires (N = 126, 68%) . The majority intended to prescribe antibiotics for less than half of their patients with sore throats (N = 69, 55%) . The variables specified in TPB predicted 48% of the variance in intention, with past behaviour adding a further 15% . Seven salient beliefs distinguished between doctors who intend to prescribe antibiotics and those who do not . CONCLUSIONS: Attitudes towards antibiotics and control beliefs are important predictors of intention to prescribe, as predicted by TPB . Interventions could target salient beliefs associated with motivation to prescribe.

Biol Reprod, 2003 Jun, 68(6), 2150 - 6 Epub 2003 Jan 22.
Human feeder layers for human embryonic stem cells; Amit M et al.; Human embryonic stem (hES) cells hold great promise for future use in various research areas, such as human developmental biology and cell-based therapies . Traditionally, these cells have been cultured on mouse embryonic fibroblast (MEF) feeder layers, which permit continuous growth in an undifferentiated stage . To use these unique cells in human therapy, an animal-free culture system must be used, which will prevent exposure to mouse retroviruses . Animal-free culture systems for hES cells enjoy three major advantages in the basic culture conditions: 1) . the ability to grow these cells under serum-free conditions, 2) . maintenance of the cells in an undifferentiated state on Matrigel matrix with 100% MEF-conditioned medium, and 3) . the use of either human embryonic fibroblasts or adult fallopian tube epithelial cells as feeder layers . In the present study, we describe an additional animal-free culture system for hES cells, based on a feeder layer derived from foreskin and a serum-free medium . In this culture condition, hES cells maintain all embryonic stem cell features (i.e., pluripotency, immortality, unlimited undifferentiated proliferation capability, and maintenance of normal karyotypes) after prolonged culture of 70 passages (>250 doublings) . The major advantage of foreskin feeders is their ability to be continuously cultured for more than 42 passages, thus enabling proper analysis for foreign agents, genetic modification such as antibiotic resistance, and reduction of the enormous workload involved in the continuous preparation of new feeder lines.

FEMS Microbiol Lett, 2003 Feb 14, 219(1), 75 - 9
A TPR-family membrane protein gene is required for light-activated heterotrophic growth of the cyanobacterium Synechocystis sp . PCC 6803; Kong R et al.; The unicellular cyanobacterium Synechocystis sp . PCC6803 can grow heterotrophically in complete darkness, given that a brief period of illumination is supplemented every day (light-activated heterotrophic growth, LAHG), or under very weak (<0.5 micromol m(-2) s(-1)) but continuous light . By random insertion of the genome with an antibiotic resistance cassette, mutants defective in LAHG were generated . In two identical mutants, sll0886, a tetratricopeptide repeat (TPR)-family membrane protein gene, was disrupted . Targeted insertion of sll0886 and three downstream genes showed that the phenotype was not due to a polar effect . The sll0886 mutant shows normal photoheterotrophic growth when the light intensity is at 2.5 micromol m(-2) s(-1) or above, but no growth at 0.5 micromol m(-2) s(-1) . Homologs to sll0886 are also present in cyanobacteria that are not known of LAHG . sll0886 and homologs may be involved in controlling different physiological processes that respond to light of low fluence.

Protein Sci, 2003 Mar, 12(3), 426 - 37
X-ray structure of the AAC(6')-Ii antibiotic resistance enzyme at 1.8 A resolution; examination of oligomeric arrangements in GNAT superfamily members; Burk DL et al.; The rise of antibiotic resistance as a public health concern has led to increased interest in studying the ways in which bacteria avoid the effects of antibiotics . Enzymatic inactivation by several families of enzymes has been observed to be the predominant mechanism of resistance to aminoglycoside antibiotics such as kanamycin and gentamicin . Despite the importance of acetyltransferases in bacterial resistance to aminoglycoside antibiotics, relatively little is known about their structure and mechanism . Here we report the three-dimensional atomic structure of the aminoglycoside acetyltransferase AAC(6')-Ii in complex with coenzyme A (CoA) . This structure unambiguously identifies the physiologically relevant AAC(6')-Ii dimer species, and reveals that the enzyme structure is similar in the AcCoA and CoA bound forms . AAC(6')-Ii is a member of the GCN5-related N-acetyltransferase (GNAT) superfamily of acetyltransferases, a diverse group of enzymes that possess a conserved structural motif, despite low sequence homology . AAC(6')-Ii is also a member of a subset of enzymes in the GNAT superfamily that form multimeric complexes . The dimer arrangements within the multimeric GNAT superfamily members are compared, revealing that AAC(6')-Ii forms a dimer assembly that is different from that observed in the other multimeric GNAT superfamily members . This different assembly may provide insight into the evolutionary processes governing dimer formation.

J Biol Chem, 2003 Apr 25, 278(17), 14769 - 75 Epub 2003 Feb 13.
Reduced affinity for Isoniazid in the S315T mutant of Mycobacterium tuberculosis KatG is a key factor in antibiotic resistance; Yu S et al.; Catalase-peroxidase (KatG) from Mycobacterium tuberculosis is responsible for the activation of the antitubercular drug isonicotinic acid hydrazide (INH) and is important for survival of M . tuberculosis in macrophages . Characterization of the structure and catalytic mechanism of KatG is being pursued to provide insights into drug (INH) resistance in M . tuberculosis . Site-directed mutagenesis was used to prepare the INH-resistant mutant KatG{S315T}, and the overexpressed enzyme was characterized and compared with wild-type KatG . KatG{S315T} exhibits a reduced tendency to form six-coordinate heme, because of coordination of water to iron during purification and storage, and also forms a highly unstable Compound III (oxyferrous enzyme) . Catalase activity and peroxidase activity measured using t-butylhydroperoxide and o-dianisidine were moderately reduced in the mutant compared with wild-type KatG . Stopped-flow spectrophotometric experiments revealed a rate of Compound I formation similar to wild-type KatG using peroxyacetic acid to initiate the catalytic cycle, but no Compound I was detected when bulkier peroxides (chloroperoxybenzoic acid, t-butylhydroperoxide) were used . The affinity of resting (ferric) KatG{S315T} for INH, measured using isothermal titration calorimetry, was greatly reduced compared with wild-type KatG, as were rates of reaction of Compound I with the drug . These observations reveal that although KatG{S315T} maintains reasonably good steady state catalytic rates, poor binding of the drug to the enzyme limits drug activation and brings about INH resistance.

FEBS Lett, 2003 Feb 11, 536(1-3), 97 - 100
Use of aminoglycoside adenyltransferase translational fusions to determine topology of thylakoid membrane proteins; Franklin JL et al.; We have developed a system to examine the topology of thylakoid membrane proteins using the bacterial aadA gene as a reporter . Translational fusions that place the aminoglycoside adenyltransferase domain in the stroma should provide high antibiotic resistance, while those that place it in the thylakoid lumen should give rise to low resistance . Genes encoding chimeric polypeptides consisting of AadA fused to varying lengths of the PsaA polypeptide, whose topology is known, were introduced into the chloroplast genome of Chlamydomonas reinhardtii . As expected, chimeras with an even number of alpha-helices in general resulted in higher resistance . This effect was not due to differences in expression or in catalytic activity . This system should prove useful in analysis of novel proteins predicted to be localized to the thylakoid membrane.

Health Econ, 2003 Feb, 12(2), 125 - 38
Deadweight loss of bacterial resistance due to overtreatment; Elbasha EH; Widespread use of antibiotics is considered the major driving force behind the development of antibiotic resistance . The benefits of exceeding the welfare-maximizing level of antibiotic use are below the costs of resistance created by this excess quantity of antibiotics used, thereby resulting in a welfare deadweight loss . This paper uses a simple economic model to examine the theoretical and empirical aspects of the welfare loss generated by resistance and analyzes its policy implications . The annual deadweight loss associated with outpatient prescriptions for amoxicillin in the United States is estimated at US dollars 225 million . Published in 2002 by John Wiley & Sons, Ltd.

Arch Microbiol, 2003 Jan-Feb, 179(2), 89 - 94 Epub 2002 Nov 15.
Development of a genetic system for Magnetospirillum gryphiswaldense; Schultheiss D et al.; Genetic analysis of bacterial magnetosome biomineralization has been hindered by the lack of an appropriate methodology for cultivation and genetic manipulation of most magnetotactic bacteria . In this report, a genetic system for Magnetospirillum gryphiswaldense is described . The system includes a plating technique that allows the screening of magnetic vs non-magnetic colonies, and a protocol for the transfer of foreign DNA by electroporation and high-frequency conjugation . Various broad-host-range vectors of the IncQ, IncP, and pBBR1 groups were found to be capable of replication in M . gryphiswaldense . Several antibiotic resistance markers that can be expressed in M . gryphiswaldense were identified . Tn 5 transposons delivered on a suicide plasmid showed transpositional insertion into random chromosomal sites.

Health Commun, 2003, 15(1), 79 - 99
Examining influences of pharmacists' communication with consumers about antibiotics; Coleman CL; A national study was conducted of community pharmacists to examine influences on their communication with consumers about antibiotics and antibiotic resistance . The objective was twofold: to explore the nature of barriers that might be changed to allow pharmacists greater participation in educational campaigns, and to discover which variables would best predict communication using a modification of the Theory of Reasoned Action model . Whereas most pharmacists agreed that their role in educating patients is important, they also noted several barriers prevent them from engaging in campaigns, such as time constraints, lack of educational materials, and fear of harming relations with physicians . Pharmacists' discussion about antibiotics in general was predicted primarily by attitudes about their role and efficacy, by their autonomy, and by enabling measures-such as pamphlets-that would better aid them in participating in a judicious antibiotic use campaign . Discussion about resistance was predicted by attitudes and enabling measures.

Wei Sheng Wu Xue Bao, 1998 Aug, 38(4), 256 - 60
{Characterization of the Streptomyces plasmid pSGL1}; Hong B et al.; The plasmid pSGL1(7.4 kb) which was isolated from antitumor antibiotic C-1027 producing strain Streptomyces globisporus C-1027 was characterized . It could be introduced into Streptomyces lividans by transformation . Following a series of deletion experiments, the minimal replication region was located within a 2.0 kb Sau 3AI fragment . pSGL1 is compatible with pIJ101 . pSGL1 and its derivatives have a copy number of 70-250 . Some of the derived plasmids such as pSGLN and pSGLS3 have suitable selectable antibiotic resistance marker and unique restriction endonuclease sites and may be potentially useful as cloning vectors . These plasmids may be used to design and construct efficient secretory expression vectors.

J Pediatr Gastroenterol Nutr, 2003 Feb, 36(2), 235 - 40
Knowledge, attitudes, and practice styles of North American pediatric gastroenterologists: Helicobacter pylori infection; Chang HY et al.; OBJECTIVE: Most Helicobacter pylori infections are acquired during childhood . The North American Society for Pediatric Gastroenterology, Hepatology and Nutrition (NASPGHAN) recently published practice guidelines for managing pediatric H . pylori infection . Before this publication, the authors conducted a survey to assess pediatric gastroenterologists' knowledge and practices regarding H . pylori . METHODS: One hundred nine of 514 NASPGHAN members completed an Internet-based questionnaire on H . pylori infection . RESULTS: Eighty-two percent of respondents performed outpatient testing for H . pylori . Of these, only 31% restricted testing to children aged >5 years . Most recommended testing for H . pylori in guideline-recommended conditions; some would not treat infected patients . Ninety-seven percent would test for H . pylori in a child with new duodenal ulcer (DU), 79% in a child with a history of DU, and 91% in a child with new gastric ulcer . However, only 86%, 60%, and 91%, respectively, would treat H . pylori infection in those conditions . A proton pump inhibitor (PPI)-based triple regimen was the first-choice therapy for 78% of respondents . Correct estimates of rates of resistance to amoxicillin, clarithromycin, metronidazole, and tetracycline were 10%, 17%, 43%, and 12%, respectively . Eighty-six percent believed there was insufficient research on H . pylori in children . CONCLUSIONS: North American pediatric gastroenterologists seem well informed about H . pylori infection in children despite the lack of published guidelines at the time of survey . Knowledge about antibiotic resistance rates was deficient . Most offered some outpatient testing for H . pylori and would test children with ulcer disease . However, some would not treat patients based on a positive result.

Exp Hematol, 2003 Jan, 31(1), 39 - 47
Transient expression of PU.1 commits multipotent progenitors to a myeloid fate whereas continued expression favors macrophage over granulocyte differentiation; McIvor Z et al.; OBJECTIVES: The Ets-family transcription factor PU.1 is expressed specifically in the hematopoietic system, in which it is absolutely required for the generation of B lymphocytes and macrophages . In contrast, overexpression of PU.1 blocks terminal differentiation of the erythroid lineage, in which it can act as an oncogene . In this study we used a multipotential progenitor cell line to examine the effects of PU.1 overexpression on myeloerythroid commitment within a single model system . MATERIALS AND METHODS: PU.1 cDNA was introduced transiently and stably into the multipotent, nonleukemic hemopoietic cell line FDCPmix . Transiently transfected cells were isolated by fluorescence-activated cell sorting within 18 hours of transfection . Stable transfectants were selected by antibiotic resistance over a number of weeks . The effects of short- and long-term overexpression of PU.1 on self-renewal, proliferation, and differentiation were investigated . RESULTS: A transient pulse of expression in multipotent progenitor cells eliminated the options of self-renewal and erythroid differentiation, resulting in commitment to the myeloid lineage . However, this transient pulse of expression did not affect the subsequent lineage choice of bipotent granulocyte/macrophage progenitors . In contrast, continuous expression of PU.1 resulted in a strong bias toward macrophage rather than granulocyte differentiation . CONCLUSIONS: These results demonstrate promyeloid effects of PU.1 at two distinct stages of hematopoiesis.

Cochrane Database Syst Rev . 2003;(1):CD004105.
Prophylactic antibiotic therapy for chronic bronchitis; Black P et al.; BACKGROUND: The use of prophylactic antibiotics to reduce the frequency and severity of acute exacerbations of chronic bronchitis is controversial . OBJECTIVES: To determine if prophylactic antibiotics reduce the frequency of exacerbations and/or days of disability in subjects with chronic bronchitis . SEARCH STRATEGY: We searched the Cochrane Airways Group Register of Clinical Trials and the bibliographies of relevant articles . SELECTION CRITERIA: Randomised controlled trials of prophylactic antibiotics in patients with chronic bronchitis and/or COPD were selected . DATA COLLECTION AND ANALYSIS: The eligibility of studies for inclusion was evaluated by three independent reviewers . MAIN RESULTS: Nine trials involving 1055 subjects were included in the analysis . All were performed before 1970 . Concealment of allocation was assessed as clearly adequate in only 3 studies . The likelihood of having a exacerbation at any time during the course of the study was decreased with treatment (Relative Risk 0.91, 95% Confidence Intervals (CI) 0.84, 0.99) . There was a small reduction in the number of exacerbations per patient per year with prophylactic antibiotics but this was not statistically significant ( Weighted Mean Difference (WMD) -0.15, 95%CI -0.34, 0.04 ) . There was a modest but significant reduction of 22% in the number of days of disability per patient per month treated ( WMD -0.95, 95%CI -1.89 to - 0.01 ) . A parallel reduction in the days of disability for each exacerbation (WMD -2.08, 95% CI -4.08 to -0.07) was seen . There was a small increase in adverse effects with antibiotics . REVIEWER'S CONCLUSIONS: Prophylactic antibiotics in chronic bronchitis / COPD have a small but statistically significant effect in reducing the days of illness due to exacerbations of chronic bronchitis . They do not have a place in routine treatment because of concerns about the development of antibiotic resistance and the possibility of adverse effects . The available data are over 30 years old, so the pattern of antibiotic sensitivity may have changed and there is a wider range of antibiotics in use.

Aliment Pharmacol Ther, 2003 Jan, 17(2), 259 - 64
Effect of different proton pump inhibitors, differences in CYP2C19 genotype and antibiotic resistance on the eradication rate of Helicobacter pylori infection by a 1-week regimen of proton pump inhibitor, amoxicillin and clarithromycin; Kawabata H et al.; AIM: To investigate the effect of different proton pump inhibitors, S-mephenytoin 4'-hydroxylase (CYP2C19) genotype and antibiotic susceptibility on the eradication rate of Helicobacter pylori . METHODS: One hundred and eighty-seven H . pylori-infected peptic ulcer patients were randomly treated with either rabeprazole (10 mg b.d.) or lansoprazole (30 mg b.d.) plus amoxicillin (750 mg b.d.) and clarithromycin (400 mg b.d.) for 1 week . The antibiotic susceptibility and CYP2C19 genotype (extensive or poor metabolizer) were investigated . RESULTS: The eradication rates in the rabeprazole-amoxicillin-clarithromycin (RAC) and lansoprazole-amoxicillin-clarithromycin (LAC) groups were 75% and 69%, respectively, on an intention-to-treat basis, and 80% and 75%, respectively, on a per protocol basis . The eradication rate for clarithromycin-resistant strains was significantly lower than that for clarithromycin-sensitive strains (24% vs . 86%, P < 0.05) . For clarithromycin-sensitive strains in the LAC group, there was a tendency for a lower eradication rate in extensive than poor metabolizers . The eradication rate in extensive metabolizers in the RAC group tended to be higher than that in extensive metabolizers in the LAC group (89% vs . 78%, P = 0.079726) . CONCLUSIONS: The success of the 1-week proton pump inhibitor-amoxicillin-clarithromycin regimen depends on the susceptibility of H . pylori to clarithromycin . Moreover, differences in CYP2C19 genotype influence the eradication rates of lansoprazole-based therapy, and the rabeprazole-based regimen has an advantage especially in extensive metabolizers.

LDI Issue Brief, 2002 May, 7(7), 1 - 4
Tensions in antibiotic prescribing; Metlay JP; Since 1999, the Agency for Healthcare Research and Quality has funded seven centers across the country to provide practical guidance to physicians and other health care professionals about the drugs they prescribe . These Centers for Education and Research on Therapeutics (CERTs) develop, translate and disseminate objective information on drugs to improve practice . The University of Pennsylvania's CERTs focuses on developing evidence for optimal treatment strategies for infectious diseases, and promoting the judicious use of antibiotics to combat the problem of antibiotic resistance . This Issue Brief explores one of the fundamental challenges physicians face in optimizing antibiotic use: the potential conflict between what is best for an individual patient, and what is best for society as a whole.

Jt Comm J Qual Saf, 2003 Jan, 29(1), 27 - 36
Deciding not to measure performance: the case of acute otitis media; Mangione-Smith R et al.; BACKGROUND: No Health Plan Employer Data and Information Set (HEDIS) performance measures evaluate health plans for possible overuse or inappropriate use of antibiotics . Acute otitis media (AOM), or infection of the middle ear, is one of the most common infections in children . The antibiotic resistance of the bacteria that cause AOM and the general overuse and inappropriate use of antibiotics for this condition are taking center stage as a major public health threat . An effort was undertaken to develop a new HEDIS performance measure that evaluates appropriate antibiotic use in children with AOM . THE MEASURE DEVELOPMENT PROCESS: The measure development process has three major phases: the initial development phase, the field-testing and analysis phase, and the measure refinement phase . With AOM, the measure development process could not proceed beyond the first phase for reasons that are be discussed in detail . CONCLUSIONS: Additional difficulties beyond feasibility issues may arise in developing a quality of care performance measure . The measure development process discussed in this article failed primarily because of issues related to relevance as well as the scientific soundness of the proposed measures . If the evidence base related to the diagnosis and management of a particular condition is sparse, no standard can be developed against which to measure performance, and the criterion of scientific soundness cannot be met . This is the case with AOM . Unfortunately, the evidence base will likely remain inadequate to support the development of such a measure in the future.

Genetics, 2002 Dec, 162(4), 1525 - 32
Plasmids spread very fast in heterogeneous bacterial communities; Dionisio F et al.; Conjugative plasmids can mediate gene transfer between bacterial taxa in diverse environments . The ability to donate the F-type conjugative plasmid R1 greatly varies among enteric bacteria due to the interaction of the system that represses sex-pili formations (products of finOP) of plasmids already harbored by a bacterial strain with those of the R1 plasmid . The presence of efficient donors in heterogeneous bacterial populations can accelerate plasmid transfer and can spread by several orders of magnitude . Such donors allow millions of other bacteria to acquire the plasmid in a matter of days whereas, in the absence of such strains, plasmid dissemination would take years . This "amplification effect" could have an impact on the evolution of bacterial pathogens that exist in heterogeneous bacterial communities because conjugative plasmids can carry virulence or antibiotic-resistance genes.

J Biomol NMR, 2002 Dec, 24(4), 351 - 6
Charged acrylamide copolymer gels as media for weak alignment; Meier S et al.; The use of mechanically strained acrylamide/acrylate copolymers is reported as a new alignment medium for biomacromolecules . Compared to uncharged, strained polyacrylamide gels, the negative charges of the acrylamide/acrylate copolymer strongly alter the alignment tensor and lead to pronounced electroosmotic swelling . The swelling itself can be used to achieve anisotropic, mechanical strain . The method is demonstrated for the alignment of TipAS, a 17 kDa antibiotic resistance protein, as well as for human ubiquitin, where alignment tensors with an A(ZZ,NH) of up to 60 Hz are achieved at a gel concentration of 2% (w/v) . The alignment can be modulated by the variation of pH, ionic strength, and gel concentration . The high mechanical stability of the swollen gels makes it possible to obtain alignment at polymer concentrations of less than 1% (w/v).

Toxicology, 2002 Dec 27, 181-182, 417 - 20
Current developments in food additive toxicology in the USA; Hattan DG et al.; A recently published proposal (Fed . Reg . 66 (2001) 4706) for mandatory submission of information on all plant-derived bioengineered foods fed to humans or animals will be reviewed . Under this proposal, information such as data on identity, level and function of the introduced substance(s); an estimate of dietary exposure; allergenic potential of the protein; data relevant to other safety issues that may be associated with the substance; selection of a comparable food; historic uses of comparable food; composition and characteristics of bioengineered food versus those of the comparable food should be provided . In addition, characterization of the parent plant; construction of the transformation vector and introduced genetic material along with number of insertion sites and genes; data on the genetic material and any newly inserted genes for antibiotic resistance should be submitted with the notification . The Interagency Coordinating Committee for Validation of Alternative Methods (ICCVAM) was identified by the U.S . Congress as the organization to review and validate new alternative toxicological test methods for 14 U.S . government agencies . Validated and accepted alternative toxicity tests will be incorporated into toxicity testing recommendations for regulatory agencies.

Lancet Infect Dis, 2003 Jan, 3(1), 13 - 21
The effect of drug resistance on the fitness of Mycobacterium tuberculosis; Cohen T et al.; Mycobacterium tuberculosis remains a leading infectious cause of morbidity and mortality . While antibiotic resistance is cited as a potential threat to efforts aimed at controlling the spread of this pathogen, it is not clear how drug resistance affects disease dynamics . The effect of mutational events that lead to antibiotic-resistant phenotypes may or may not have a predictable effect on the fitness of drug-resistant tuberculosis strains . Here, we review the literature on laboratory studies of the fitness of drug-resistant tuberculosis, we examine the evidence from cluster studies, and we consider the effect of drug resistance on disease dynamics in mathematical models . On the basis of these diverse lines of evidence, we conclude that the fitness estimates of drug-resistant M tuberculosis are quite heterogeneous and that this variation may preclude our ability to predict future trends of this pathogen.

Environ Int, 2003 Jan, 28(7), 587 - 95
Bacterial antibiotic resistance levels in Danish farmland as a result of treatment with pig manure slurry; Sengelov G et al.; Resistance to tetracycline, macrolides and streptomycin was measured for a period of 8 months in soil bacteria obtained from farmland treated with pig manure slurry . This was done by spread plating bacteria on selective media (Luria Bertani (LB) medium supplemented with antibiotics) . To account for seasonal variations in numbers of soil bacteria, ratios of resistant bacteria divided by total count on nonselective plates were calculated . Soil samples were collected from four different farms and from a control soil on a fifth farm . The control soil was not amended with animal manure . The occurrence of tetracycline-resistant bacteria was elevated after spread of pig manure slurry but declined throughout the sampling period to a level corresponding to the control soil . Higher load of pig manure slurry yielded higher occurrence of tetracycline resistance after spreading; however, the tetracycline resistance declined to normal occurrence defined by the tetracycline resistance occurrence in the control soil . Concentrations of tetracycline in soil and in pig manure slurry were measured using HPLC . No tetracycline exceeding the detection limit was found in soil samples . Manure slurry concentrations of tetracycline for three of the farms were 42, 81 and 698 microg/l, respectively . For streptomycin and macrolides, only minor variations in resistance levels were detected . Results obtained in this study thus indicate that tetracycline resistance levels in soil are temporarily influenced by the addition of pig manure slurry . The results indicate also that increased amount of pig manure slurry amendment may result in increased levels of tetracycline resistance in the soil.

Mol Biol (Mosk), 2002 Nov-Dec, 36(6), 1021 - 5
{The method of template construction for sequencing extended DNA fragments}; Ermolaeva MA et al.; A rapid and convenient method was proposed for constructing insertional mutants in the sequencing of extended DNA fragments . The gist is insertion of an antibiotic resistance gene in plasmid DNA digested with DNase I . DNase I provides for a uniform distribution of insertion sites along the plasmid, and the background is low owing to antibiotic-based selection . The method requires neither high quality nor large amounts of plasmid DNA (which is especially important with low-copied plasmids), yields the results independent of the plasmid nucleotide sequence, and allows highly accurate analysis and ordering of the insertional mutants.

Emerg Infect Dis, 2002 Dec, 8(12), 1460 - 7
Outpatient antibiotic use and prevalence of antibiotic-resistant pneumococci in France and Germany: a sociocultural perspective; Harbarth S et al.; The prevalence of penicillin-nonsusceptible pneumococci is sharply divided between France (43%) and Germany (7%) . These differences may be explained on different levels: antibiotic-prescribing practices for respiratory tract infections; patient-demand factors and health-belief differences; social determinants, including differing child-care practices; and differences in regulatory practices . Understanding these determinants is crucial for the success of possible interventions . Finally, we emphasize the overarching importance of a sociocultural approach to preventing antibiotic resistance in the community.

Eur J Pediatr, 2002 Dec, 161 Suppl 2, S135 - 9 Epub 2002 Oct 01.
Invasive pneumococcal disease in children: geographic and temporal variations in incidence and serotype distribution; Hausdorff WP; We examined published studies from the United States, Europe, and Latin America to better understand geographic and temporal variability in the epidemiology of invasive pneumococcal disease (IPD) in young children . A comparison of IPD incidence levels reported for children <5 years of age within the United States (CDC) surveillance system over the past 5 years revealed little variation over that time period . Within the Danish national surveillance system, a 3-4 fold increase in bacteremic IPD in that age group was reported over the past 15 years . Nonetheless, the United States IPD incidence remains 4-8 fold higher than that reported for Denmark or other Western European countries . A retrospective analysis at one North American hospital spanning 40 years indicated that the seven serogroups represented in the licensed pneumococcal conjugate vaccine (4, 6, 9, 14, 18, 19, 23) consistently comprised 80%-90% of all serogroups causing IPD in young children . More recent retrospective comparisons in Spain, the United Kingdom, France, Belgium, and Denmark suggest modest increases in vaccine serogroup coverage over the past 10-20 years . However, in European children vaccine serogroups consistently account for 10%-20% less of the overall IPD burden as compared to North American children . CONCLUSION: factors that likely underlie a significant proportion of these apparent temporal and geographic differences include variability in blood culturing rates, in antibiotic resistance levels, and in the precise age distributions of the populations studied.

Science, 2003 Jan 17, 299(5605), 411 - 4 Epub 2002 Dec 19.
Production of alpha 1,3-galactosyltransferase-deficient pigs; Phelps CJ et al.; The enzyme alpha1,3-galactosyltransferase (alpha1,3GT or GGTA1) synthesizes alpha1,3-galactose (alpha1,3Gal) epitopes (Galalpha1,3Galbeta1,4GlcNAc-R), which are the major xenoantigens causing hyperacute rejection in pig-to-human xenotransplantation . Complete removal of alpha1,3Gal from pig organs is the critical step toward the success of xenotransplantation . We reported earlier the targeted disruption of one allele of the alpha1,3GT gene in cloned pigs . A selection procedure based on a bacterial toxin was used to select for cells in which the second allele of the gene was knocked out . Sequencing analysis demonstrated that knockout of the second allele of the alpha1,3GT gene was caused by a T-to-G single point mutation at the second base of exon 9, which resulted in inactivation of the alpha1,3GT protein . Four healthy alpha1,3GT double-knockout female piglets were produced by three consecutive rounds of cloning . The piglets carrying a point mutation in the alpha1,3GT gene hold significant value, as they would allow production of alpha1,3Gal-deficient pigs free of antibiotic-resistance genes and thus have the potential to make a safer product for human use.

FEMS Microbiol Lett, 2002 Dec 17, 217(2), 191 - 7
Cell-detaching Escherichia coli (CDEC) strains from children with diarrhea: Identification of a protein with toxigenic activity; Abduch Fabrega VL et al.; Twenty-four strains of cell-detaching Escherichia coli (CDEC) isolated from stool specimens in different cities in Brazil were examined for virulence properties . Aerobactin production and multiple antibiotic resistance were observed in most of the isolates . In hybridization studies, the alphahly, pap, and cnf sequences, common properties of this category of E . coli, were found in a minority of isolates . Half of the CDEC isolates had enteroaggregative DNA sequences (pet, astA, aggA), six strains carried the shet1 gene, nine strains carried the daaC sequence, and one strain carried the stp gene . Thirteen strains induced fluid accumulation in the rabbit intestinal loop assay . Supernatant filtrate of one of those strains, which did not hybridize with any of the toxin probes tested, induced destructive lesions in the rabbit ileal loop and enterotoxic activity in the Ussing chamber . A 12-kDa protein purified by 60% ammonium sulfate precipitation of the supernatant filtrate demonstrated a toxigenic effect that was inhibited by the anti-12-kDa protein antiserum .

Arch Inst Pasteur Madagascar, 1999, 65(1-2), 124 - 6
{Evaluation of the use of antibiotics in the hospital pediatric milieu}; Rasamoelisoa JM et al.; Excessive and inappropriate use of antibiotics is a world-wide serious problem because it contributes to the development and the spreading of antibiotic resistance . The authors carried out a study for one year by 1998, in order to evaluate antibiotic prescriptions in three childhood diseases: respiratory, digestive and neurological diseases . Patients hospitalized at the unit paediatric of the General hospital of Befelatanana are 0 month to 15 years old . Three antibiotic families were frequently used: cotrimoxazole, penicillins derivatives and aminoglycosides . Results of this study noted that antibiotic precriptions were irrational . Some factors may give explanations to these facts: the insufficiency of biological labs, but also expensive costs of exams . It is desirable that all paediatricians can gather in order to draw up clinical protocols and to assess them by a multicentric survey . Further results may be used as reference for empirical or probabilist antibiotic prescriptions of which efficacy will be checked by an experienced lab.

J Health Econ, 2002 Nov, 21(6), 1071 - 83
Economic implications of antibiotic resistance in a global economy; Rudholm N; This paper concerns the economic implications of antibiotic resistance in a global economy . The global economy consists of several countries, where antibiotic consumption creates a stock of bacteria which is resistant to antibiotics . This stock affects the welfare in all countries because of the risk that resistant bacterial strains may be transmitted . The main purpose of the paper is to compare the socially optimal resource allocation with the allocation brought forward by the decentralized market economy . In addition, a dynamic Pigouvian tax designed to implement the globally optimal resource allocation is presented.

J Biomed Mater Res B Appl Biomater, 2003 Jan 15, 64(1), 1 - 5
The release of gentamicin from acrylic bone cements in a simulated prosthesis-related interfacial gap; Hendriks JG et al.; Gentamicin is added to polymethylmethacrylate bone cements in orthopedics as a measure against infection in total joint arthroplasties . Numerous studies have been published on gentamicin release from bone cements, but none have been able to estimate the local concentrations in the prosthesis-related interfacial gap, present after implantation . The aim of this study was to develop a method allowing determination of antibiotic release in such a gap . Two-hundred-micrometer-wide gaps with a volume of 6 microl and a surface area of 0.6 cm2 were created by inserting stainless-steel strips in gentamicin-loaded bone cement plugs prior to polymerization . After hardening, the gap surface was exposed to 6 microl or 10 ml of phosphate-buffered saline . Within 2 h, gentamicin concentrations in the gaps reached around 4000 microg/ml for 4 different CMW and Palamed cements and 2500 microg/ml for Palacos R . Concentrations measured in the larger volume were several hundred times lower than in the gaps . This simulated prosthesis-related interfacial gap model offers new insights in the clinical efficacy of antibiotic-loaded bone cements . It is demonstrated that concentrations up to 1000-fold the antibiotic resistance levels for most bacterial strains causing implant infection can be achieved in a realistic in vitro model .

Best Pract Res Clin Obstet Gynaecol, 2002 Dec, 16(6), 801 - 14
The role of serology, antibiotic susceptibility testing and serovar determination in genital chlamydial infections; Persson K; Systemic and local antibodies regularly develop in genital infections caused by Chlamydia trachomatis . Such antibodies cannot be used as a sign of current infection as they often persist for years after the infection has resolved . Chlamydial antibodies have, however, been extremely useful for demonstrating associations between C . trachomatis and clinical conditions such as ectopic pregnancy and tubal factor infertility . In particular, antibodies to the chlamydial heat shock protein 60 predict the presence of tubal scarring.C . trachomatis has been divided into 15 (or 18) serovars, with many genotypes within each serovar . Differences in pathogenicity between serovars have been reported but no general pattern has emerged . Genotyping is a powerful epidemiological tool but is not yet ready for routine clinical use.C . trachomatis infections can be successfully treated by tetracycline or macrolides . Some resistant strains have been reported, causing treatment failures, and the problem of emerging antibiotic resistance cannot be neglected.

Appl Microbiol Biotechnol, 2002 Dec, 60(4), 461 - 8 Epub 2002 Oct 18.
Escherichia coli cells penetrated by chrysotile fibers are transformed to antibiotic resistance by incorporation of exogenous plasmid DNA; Yoshida N et al.; A suspension of recipient Escherichia coli cells in stationary phase, chrysotile asbestos, and pUC18 donor DNA spread over the surface of a Luria-Bertani agar plate using a streak bar several times, resulted in intracellular uptake of the plasmid DNA by the E . coli cells . The transformation efficiency was highest with a duration of cell exposure to chrysotile of more than 60 s and an agar concentration of 2% . To improve chrysotile-mediated transformation efficiency, we systematically optimized various conditions and parameters . In comparison to chrysotile exposure without cations, exposure with cations produced up to 100-fold more transformants . Optimized conditions resulted in 10(6) transformants/ micro g pUC18 DNA . The drastic physical change due to 'quick drying on the surface of the agar plate' when cells were exposed to chrysotile, was essential for chrysotile-mediated transformation . We suggest that DNA uptake mediated by chrysotile asbestos is the result of a mechanical physical transformation of E . coli, since the E . coli cells are not chemically competent . Electron microscopy of cells exposed to chrysotile suggested penetration of the E . coli membrane by chrysotile fibers . It is suggested that E . coli transformation by the plasmid DNA was the result of penetration by chrysotile fibers to which plasmid DNA is bound or adsorbed.

J Theor Biol, 2003 Jan 7, 220(1), 67 - 74
The evolution of group-beneficial traits in the absence of between-group selection; Dugatkin LA et al.; One specific prediction emerging from trait-group models of natural selection is that when individuals possess traits that benefit other group members, natural selection will favor "cheating" (i.e . not possessing the group-beneficial trait) within groups . Cheating is selected within groups because it allows individuals to avoid bearing the relative costs typically associated with group-beneficial traits, but to still reap the benefits associated with the acts of other group members . Selection between groups favors traits that benefit other group members . The relative strength of within- and between-group selection then determines the equilibrium frequency of those who produce group-beneficial traits and those that do not . Here we demonstrate that individual-level selection, that is selection within groups can also produce an intermediate frequency of such group-beneficial traits by frequency-dependent selection . The models we develop are general in nature, but were inspired by the evolution of antibiotic resistance in bacteria . The theory developed here is distinct from prior work that relies on reciprocity or kinship per se to achieve cooperation and altruism among group members .

J Appl Microbiol, 1997 Feb, 82(2), 245 - 52
Metal tolerance and antibiotic resistance patterns of a bacterial population isolated from sea water; Sabry SA et al.; The total aerobic heterotrophic and metal-resistant bacterial communities were studied in marine water . The resistance patterns, expressed as MICs, for 81 bacterial isolates to eight heavy metals were surveyed by using the agar dilution method . A great proportion of the isolates were sensitive to cadmium (99%), mercury (91%), zinc (84%) and cobalt (83%) . On the other hand, 94%, 40%, 35% and 22% were resistant to lead, nickel, arsenate and copper, respectively . The majority of the tested strains (95.06%) were multiple metal-resistant, with pentametal resistance as the major pattern (25.9%) . The response of the isolates to 11 tested antibiotics was tested and ranged from complete resistance to total sensitivity and multiple antibiotic resistance was exhibited by 70.38% of the total isolated population . The highest incidence of metal-antibiotic double resistance existed between lead and all antibiotics (100%), copper and penicillin (95%) and nickel and ampicillin (83.3%).

Int J Antimicrob Agents, 2002, 20 Suppl 1, S1 - 12
Guidelines for the management of community-acquired pneumonia in Saudi Arabia: a model for the Middle East region; Memish ZA et al.; Worldwide, community-acquired pneumonia (CAP) is a common respiratory tract infection and is now a growing public health concern in Saudi Arabia . In an effort to simplify treatment regimens to aid the practitioner, empirical treatment guidelines for CAP have evolved across the international medical community, reducing the number of antibiotics used and improving outcomes . Saudi Arabia and the surrounding region have no such consensus guidelines and this document aims to redress this lack . The potential impacts of developing and implementing CAP treatment guidelines in Saudi Arabia, which are new to the Kingdom, will be examined . Widespread adoption of these SACAP guidelines could lead to nationwide reductions of antibiotic resistance and improvement of clinical outcomes . Ultimately, Kingdomwide uniformity of treatment algorithms provides a foundation for both database generation and valuable outcomes of research in the future.

Pediatr Infect Dis J, 2002 Nov, 21(11), 1023 - 8
Decreased number of antibiotic prescriptions in office-based settings from 1993 to 1999 in children less than five years of age; Halasa NB et al.; OBJECTIVE: Increasing rates of antibiotic resistance have stimulated efforts to decrease antibiotic use . To assess the success of these efforts, we analyzed antibiotic prescribing trends in children younger than 5 years old, the group with the highest use, from 1993 to 1999 . METHODS: Data from the National Ambulatory Medical Care Survey were analyzed to determine antibiotic prescribing patterns for office-based visits from 1993 to 1999 for children <5 years old . Data were stratified by US regions, patient's race and gender . Antibiotic prescription rates per 1,000 population were calculated with population data from the US Census Bureau as the denominator . Specific prescribing of penicillins, cephalosporins, macrolides and sulfas was also assessed . RESULTS: Overall antibiotic prescribing in the office-based setting peaked in 1995 at 1,191 antibiotic courses per 1,000 children, then declined to 698 per 1,000 in 1999, a decrease of 41% . Antibiotic prescribing was consistently higher in whites than blacks; however, declines in prescribing over time were observed in both groups . Although there was wide regional variation in antibiotic prescribing in the early 1990's, by the late 1990's prescribing rates were similar in all regions . Prescriptions for penicillins and cephalosporins combined comprised 77 and 70% of total prescriptions during 1993 to 1997 and 1998 to 1999, respectively . Macrolide prescriptions reached a nadir during 1993 to 1997, accounting for 9% of the total, but increased to 16% during 1998 to 1999 . CONCLUSION: Since 1995 the rates of antibiotic prescriptions in children <5 years of age have declined substantially . At the same time changes have occurred in the types of antibiotics prescribed . It appears that efforts to reduce antibiotic use have been successful . Whether this decrease in use will be accompanied by lower rates of antibiotic resistance will need to be determined.

J Virol, 2002 Dec, 76(24), 12925 - 33
RNA silencing of dengue virus type 2 replication in transformed C6/36 mosquito cells transcribing an inverted-repeat RNA derived from the virus genome; Adelman ZN et al.; Double-stranded RNA (dsRNA) initiates cellular posttranscriptional responses that are collectively called RNA silencing in a number of different organisms, including plants, nematodes, and fruit flies . In plants, RNA silencing has been associated with protection from virus infection . In this study, we demonstrate that dsRNA-mediated interference also can act as a viral defense mechanism in mosquito cells . C6/36 (Aedes albopictus) cells were stably transformed with a plasmid designed to transcribe an inverted-repeat RNA (irRNA) derived from the genome of dengue virus type 2 (DEN-2) capable of forming dsRNA . Clonal cell lines were selected with an antibiotic resistance marker and challenged with DEN-2 . The cell lines were classified as either susceptible or resistant to virus replication, based on the percentage of cells expressing DEN-2 envelope (E) antigen 7 days after challenge . Eight out of 18 (44%) cell lines designed to express irRNA were resistant to DEN-2 challenge, with more than 95% of the cells showing no DEN-2 antigen accumulation . One of the DEN-2-resistant cell lines, FB 9.1, was further characterized . DEN-2 genome RNA failed to accumulate in FB 9.1 cells after challenge . Northern blot hybridization detected transcripts containing transgene sequences of both sense and antisense polarity, suggesting that DEN-2-specific dsRNA was present in the cells . In addition, a class of small RNAs 21 to 25 nucleotides in length was detected that specifically hybridized to labeled sense or antisense DEN-2 RNA derived from the target region of the genome . These observations were consistent with RNA silencing as the mechanism of resistance to DEN-2 in transformed mosquito cells.

Clin Microbiol Infect, 2002 Sep, 8(9), 598 - 603
Prevalence of antibiotic resistance of Helicobacter pylori isolates in Estonia during 1995-2000 in comparison to the consumption of antibiotics used in treatment regimens; Loivukene K et al.; OBJECTIVE: To find a possible relation between the dynamics of antibiotic resistance of Helicobacter pylori isolates and the consumption of antibiotics during the last several years in Estonia . METHODS: Helicobacter pylori isolates were collected from the gastric mucosa of patients with peptic ulcer (153) and gastritis (68) and isolated on the Columbia Agar Base . From 1995 to 1997 the disk-diffusion method was used for testing of H . pylori susceptibility to metronidazole (115 isolates), erythromycin (119 isolates), tetracycline (119 isolates) and amoxicillin (119 isolates) . From 1998 to 2000 the susceptibility of H . pylori to metronidazole (106 isolates), amoxicillin (30 isolates), clarithromycin (106 isolates) and ciprofloxacin (30 isolates) was assessed by E tests . Data from the Estonian State Agency of Medicines were used to determine the antibiotic consumption rate . RESULTS: Up to the year 2000 all the investigated H . pylori isolates were susceptible to ciprofloxacin; the resistance to clarithromycin, tetracycline, amoxicillin and erythromycin was 3%, 1.7%, 0.7% and 2.5%, respectively . Forty-six percent of H . pylori isolates were resistant to metronidazole . During 1995-2000 the consumption of amoxicillin, erythromycin and ciprofloxacin increased and the consumption of tetracycline decreased . The increasing consumption of amoxicillin reached a level 5.7 times than that of the consistent use of metronidazole . The resistance to amoxicillin appeared to be very low and resistance to metronidazole was continuously high . The increase of clarithromycin consumption (from 0.002 to 1.119 defined daily doses/1000) during three years was associated with the appearance of the first clarithromycin-resistant isolates in 2000 . CONCLUSION: No relation was observed between the antibiotic consumption rate and the resistance pattern of H . pylori to metronidazole, amoxicillin, erythromycin, tetracycline and ciprofloxacin during recent years among the in population.

Antibiot Khimioter, 2002, 47(6), 7 - 11
{Effect of glucose on the antibiotic activity and antibiotic resistance of Streptomyces peucetius subsp . caesius ATCC 27952-2 and its mutants}; Dubitskaia LP et al.; The growth of anthracycline producer Streptomices peucetius subsp . caesius ATCC 27952-2 was inhibited by presence of glucose on complete media, containing alternative carbon sources . Amount of clones not producing antibiotic increased to 80.2 per cent along with elevation of glucose concentration in corn meal medium from 0.1 to 1.0 per cent . Mutants of S . peucetius subsp . caesius ATCC 27952-2 able to grow on complete media with 2 per cent of glucose (glr-mutants) were obtained . Glr-mutants had decreased antibiotic production in comparison with 27952-2 strain . 17 per cent of studied glr-mutants synthesized 1.6-3.1-fold quantities of anthracyclines in comparison with parental strain . Glr-mutants synthesized more biomass, although more slowly utilized glucose than strain 27952-2.

Wien Klin Wochenschr, 2002 Jun 28, 114(12), 448 - 53
The impact of primary antibiotic resistance on the efficacy of ranitidine bismuth citrate- vs . omeprazole-based one-week triple therapies in H . pylori eradication--a randomised controlled trial; Bago J et al.; AIMS: To compare ranitidine bismuth citrate with omeprazole as to their efficacy to eradicate H . pylori in two different treatment schedules both consisting of a combination of either of above with two antibiotics for 1 week, and to relate these treatment results to primary antibiotic resistance . METHODS: 256 H . pylori positive patients with non-ulcer dyspepsia were randomised to one of the following four treatment groups: omeprazole 20 mg + clarithromycin 500 mg + amoxycillin 1000 mg (OCA); ranitidine bismuth citrate 400 mg + clarithromycin 500 mg + amoxycillin 1000 mg (RBCCA); omeprazole 20 mg + clarithromycin 500 mg + metronidazole 500 mg (OCM); ranitidine bismuth citrate 400 mg + clarithromycin 500 mg + metronidazole 500 mg (RBCCM) . All drugs were given twice daily for one week . The patients were assessed for prevalence of H . pylori by CLO test, histology and culture on gastric biopsy samples obtained during upper gastrointestinal endoscopy before randomisation and 4-6 weeks after completion of therapy . Bacterial sensitivity to clarithromycin, metronidazole and amoxycillin was determined by E-test . RESULTS: On per-protocol analysis, overall eradication rates were 96% for RBCCA vs . 85% for OCA (p = 0.03), and 95% for RBCCM vs . 79% for OCM (p = 0.01) . Amongst the 196 patients (77% of the entire study group) in whom antibiotic sensitivity testing was technically feasible, primary resistance was found in 8% for clarithromycin, in 33% for metronidazole, and in 0% for amoxycillin . Eradication of clarithromycin sensitive/resistant strains was 89%/40% for OCA (p = 0.0042) and 98%/80% for RBCCA (p = 0.0428) . When strains were sensitive to both antibiotics, cure rates with OCM/RBCCM were 87%/96% respectively (p = 0.39), for strains resistant to clarithromycin only, eradication was achieved in 82% with OCM vs . 94% with RBCCM (p = 0.2), and in the case of metronidazole resistance in 85% with OCM vs . 94% with RBCCM (p = 0.09) . CONCLUSIONS: Ranitidine bismuth citrate in combination with clarithromycin and either metronidazole or amoxycillin produced higher eradication rates than omeprazole co-administered with the same antibiotics . This appeared especially prominent in the subgroups with clarithromycin resistance without, however, reaching statistical significance . Efficacy of neither eradication regimen was influenced by metronidazole sensitivity to a significant degree.

Anal Biochem, 2002 Oct 15, 309(2), 224 - 31
Development of an assay for beta-lactam hydrolysis using the pH-dependence of enhanced green fluorescent protein; Puckett LG et al.; An assay has been developed utilizing the pH-dependent fluorescence of enhanced green fluorescent protein (EGFP) . This photoprotein allows for the study of kinetic properties of hydrolytic enzymes based on the production of protons . As a model system, beta-lactamase, a well-characterized enzyme responsible for antibiotic resistance in many bacteria, was used . More specifically, EGFP and beta-lactamase were genetically fused using overlap extension PCR and incorporated into a bacterial expression vector . The vector was subsequently transformed into Escherichia coli, and the fusion protein was expressed and purified . beta-Lactamase catalyzes the hydrolysis of the beta-lactam ring of ampicillin . This causes a decrease in the local pH, which in turn changes the spectral properties of EGFP . This property was utilized to perform enzyme kinetic studies on the new fusion protein as well as on the beta-lactamase inhibitor, sulbactam . The assay can be used to evaluate substrates and inhibitors of beta-lactamase in a format that should be amenable to high-throughput screening.

Appl Environ Microbiol, 2002 Nov, 68(11), 5528 - 36
Construction of a shuttle vector for, and spheroplast transformation of, the hyperthermophilic archaeon Pyrococcus abyssi; Lucas S et al.; Our understanding of the genetics of species of the best-studied hyperthermophilic archaea, Pyrococcus spp., is presently limited by the lack of suitable genetic tools, such as a stable cloning vector and the ability to select individual transformants on plates . Here we describe the development of a reliable host-vector system for the hyperthermophilic archaeon Pyrococcus abyssi . Shuttle vectors were constructed based on the endogenous plasmid pGT5 from P . abyssi strain GE5 and the bacterial vector pLitmus38 . As no antibiotic resistance marker is currently available for Pyrococcus spp., we generated a selectable auxotrophic marker . Uracil auxotrophs resistant to 5-fluoorotic acid were isolated from P . abyssi strain GE9 (devoid of pGT5) . Genetic analysis of these mutants revealed mutations in the pyrE and/or pyrF genes, encoding key enzymes of the pyrimidine biosynthetic pathway . Two pyrE mutants exhibiting low reversion rates were retained for complementation experiments . For that purpose, the pyrE gene, encoding orotate phosphoribosyltransferase (OPRTase) of the thermoacidophilic crenarchaeote Sulfolobus acidocaldarius, was introduced into the pGT5-based vector, giving rise to pYS2 . With a polyethylene glycol-spheroplast method, we could reproducibly transform P . abyssi GE9 pyrE mutants to prototrophy, though with low frequency (10(2) to 10(3) transformants per micro g of pYS2 plasmid DNA) . Transformants did grow as well as the wild type on minimal medium without uracil and showed comparable OPRTase activity . Vector pYS2 proved to be very stable and was maintained at high copy number under selective conditions in both Escherichia coli and P . abyssi.

J Econ Entomol, 2002 Oct, 95(5), 1044 - 8
Antibiosis of the pith maize to Sesamia nonagrioides (Lepidoptera: Noctuidae); Ordas B et al.; Thirteen inbred lines of maize (Zea mays L.) with different levels of stem resistance to the stem borer Sesamia nonagrioides Lefevbre were evaluated in the field and the laboratory to determine the antibiotic resistance to this pest . Inbreds CM151, CO125, and EP39 had antibiotic pith as well as stem resistance, so the pith could play a role in stem resistance . Inbreds A509, F473, and PB130 did not have antibiotic pith but had stem resistance; therefore, other mechanisms could confer stem resistance . Finally, the inbred MS1334 had antibiotic pith and did not show stem resistance; thus, other factors could compensate the effect of the pith . Therefore, although pith antibiotic compound seems to play a role in the defense against S . nonagrioides attack, it is not the only possible mechanism of defense.

J Microbiol Methods, 2003 Jan, 52(1), 93 - 100
Direct random insertion mutagenesis of Helicobacter pylori; de Jonge R et al.; Random insertion mutagenesis is a widely used technique for the identification of bacterial virulence genes . Most strategies for random mutagenesis involve cloning in Escherichia coli for passage of plasmids or for phenotypic selection . This can result in biased selection due to restriction or instability of the cloned DNA, or toxicity of the encoded products . We therefore created two mutant libraries in the human pathogen Helicobacter pylori using a simple, direct mutagenesis technique, which does not require E . coli as intermediate . H . pylori total DNA was digested, circularized and digested again with a frequently cutting restriction enzyme, and the resulting fragments were ligated to a kanamycin antibiotic resistance cassette . Subsequently, the ligation mixture was transformed into the parental H . pylori strain 1061 . Insertion of the kanamycin cassette by double homologous recombination into the genome of H . pylori 1061 resulted in approximately 2500 kanamycin resistant colonies . Heterogeneity of kanamycin cassette insertion was confirmed by Southern blotting . The isolation of two independent H . pylori mutants defective in production of urease from this library underlines the potential of this mutagenesis strategy.

Arch Intern Med, 2002 Oct 28, 162(19), 2210 - 6
Antibiotic resistance: a survey of physician perceptions; Wester CW et al.; BACKGROUND: Antibiotic resistance is caused partly by excessive antibiotic prescribing, yet little is known about prescribers' views on this problem . METHODS: We surveyed 490 internal medicine physicians at 4 Chicago-area hospitals to assess their attitudes about the importance of antibiotic resistance, knowledge of its prevalence, self-reported experience with antibiotic resistance, beliefs about its causes, and attitudes about interventions designed to address the problem . RESULTS: The response rate was 87% (424 of 490 physicians) . Antibiotic resistance was perceived as a very important national problem by 87% of the respondents, but only 55% rated the problem as very important at their own hospitals . Nearly all physicians (97%) believed that widespread and inappropriate antibiotic use were important causes of resistance . Yet, only 60% favored restricting use of broad-spectrum antibiotics, although this percentage varied by hospital and physician group . CONCLUSIONS: Although most physicians view antibiotic resistance as a serious national problem, perceptions about its local importance, its causes, and possible solutions vary more widely . Disparities in physician knowledge, beliefs, and attitudes may compromise efforts to improve antibiotic prescribing and infection control practices.

Microbiology, 2002 Oct, 148(Pt 10), 3007 - 17
Specialized transduction: an efficient method for generating marked and unmarked targeted gene disruptions in Mycobacterium tuberculosis, M . bovis BCG and M . smegmatis; Bardarov S et al.; The authors have developed a simple and highly efficient system for generating allelic exchanges in both fast- and slow-growing mycobacteria . In this procedure a gene of interest, disrupted by a selectable marker, is cloned into a conditionally replicating (temperature-sensitive) shuttle phasmid to generate a specialized transducing mycobacteriophage . The temperature-sensitive mutations in the mycobacteriophage genome permit replication at the permissive temperature of 30 degrees C but prevent replication at the non-permissive temperature of 37 degrees C . Transduction at a non-permissive temperature results in highly efficient delivery of the recombination substrate to virtually all cells in the recipient population . The deletion mutations in the targeted genes are marked with antibiotic-resistance genes that are flanked by gammadelta-res (resolvase recognition target) sites . The transductants which have undergone a homologous recombination event can be conveniently selected on antibiotic-containing media . To demonstrate the utility of this genetic system seven different targeted gene disruptions were generated in three substrains of Mycobacterium bovis BCG, three strains of Mycobacterium tuberculosis, and Mycobacterium smegmatis . Mutants in the lysA, nadBC, panC, panCD, leuCD, Rv3291c and Rv0867c genes or operons were isolated as antibiotic-resistant (and in some cases auxotrophic) transductants . Using a plasmid encoding the gammadelta-resolvase (tnpR), the resistance genes could be removed, generating unmarked deletion mutations . It is concluded from the high frequency of allelic exchange events observed in this study that specialized transduction is a very efficient technique for genetic manipulation of mycobacteria and is a method of choice for constructing isogenic strains of M . tuberculosis, BCG or M . smegmatis which differ by defined mutations.

Curr Opin Crit Care, 2002 Oct, 8(5), 430 - 4
Antibiotic resistance in the intensive care unit; Baughman RP; The increase in antibiotic resistance over the past 10 years can be traced to several factors . This includes exogenous transmission of bacteria, usually by hospital personnel . The use of potent antibiotics also can select for resistant bacteria initially present in low quantities . Strategies to reduce antibiotic resistance can be tailored to specific outbreaks in a given ICU . General strategies for reducing antibiotic resistance, on the other hand, include varying the agents used in the ICU over time . Reduction of the duration of therapy may prove to be another method of reducing antibiotic resistance.

Curr Opin Pharmacol, 2002 Oct, 2(5), 487 - 92
Antibiotic resistance - is resistance detected by surveillance relevant to predicting resistance in the clinical setting?
Karlowsky JA, Sahm DF.
Local, regional, national and global surveillance initiatives have several important functions, which include identifying shifts in antibiotic resistance, detecting the emergence of new resistance mechanisms and monitoring the impact of changes made to empiric prescribing, infection control and public health guidelines . Although the need for surveillance is indubitable and its use in the treatment of individual patients important, it cannot unequivocally predict outcomes in patients with infections . Treatment regimens for individual patients with suspected or demonstrated infections should be developed following consideration of symptoms, laboratory findings and relevant medical history, and in the context of appropriate local and widespread antibiotic resistance trends.

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