Development Of High-level Resistance Research Articles

Aquatic Environments as Hotspots of Transferable Low-Level Quinolone Resistance and Their Potential Contribution to High-Level Quinolone Resistance.

The disposal of antibiotics in the aquatic environment favors the selection of bacteria exhibiting antibiotic resistance mechanisms. Quinolones are bactericidal antimicrobials extensively used in both human and animal medicine. Some of the quinolone-resistance mechanisms are encoded by different bacterial genes, whereas others are the result of mutations in the enzymes on which those antibiotics act. The worldwide occurrence of quinolone resistance genes in aquatic environments has been widely reported, particularly in areas impacted by urban discharges. The most commonly reported quinolone resistance gene, qnr, encodes for the Qnr proteins that protect DNA gyrase and topoisomerase IV from quinolone activity. It is important to note that low-level resistance usually constitutes the first step in the development of high-level resistance, because bacteria carrying these genes have an adaptive advantage compared to the highly susceptible bacterial population in environments with low concentrations of this antimicrobial group. In addition, these genes can act additively with chromosomal mutations in the sequences of the target proteins of quinolones leading to high-level quinolone resistance. The occurrence of qnr genes in aquatic environments is most probably caused by the release of bacteria carrying these genes through anthropogenic pollution and maintained by the selective activity of antimicrobial residues discharged into these environments. This increase in the levels of quinolone resistance has consequences both in clinical settings and the wider aquatic environment, where there is an increased exposure risk to the general population, representing a significant threat to the efficacy of quinolone-based human and animal therapies. In this review the potential role of aquatic environments as reservoirs of the qnr genes, their activity in reducing the susceptibility to various quinolones, and the possible ways these genes contribute to the acquisition and spread of high-level resistance to quinolones will be discussed.

Essential Oils and Their Formulations for the Control of Curculionidae Pests

Pesticides are widely used in producing food to control pests. However, it has been determined that synthetic pesticides present severe toxicity (residual), while they also result in environmental contamination and development of high-level resistance in some insect species. Due to this, some of these susbtances have been banned or restricted in many countries, which has reduced the number of agrochemicals that can be used for pest control, particularly in the case of crops exported to green markets such as Europe and Asia. Under this scenario, essential oils (EOs) are being increasingly studied as bioinsecticides because they are renewable, natural, biodegradable, non-persistent in the environment and safe to non-target organism and humans. It has been determined that EOs have repellent, ovicidal, larvicidal, and insecticidal effects against different types of pests, but they also have some drawbacks due to their high volatility and low aqueous solubility. This mini-review focusses on EOs used as bioinsecticides for the control of Curculionidae and on current stabilization techniques, such as nanoencapsulation, to prolog the biocidal effect of EOs against these pests.

The challenge of antibiotic selection in prosthetic joint infections due to Corynebacterium striatum: a case report

BackgroundCorynebacterium striatum is a gram-positive facultative anaerobe found in the environment and human flora that has historically been considered a contaminant. More recently, Corynebacterium striatum has been implicated in human infections, including respiratory infections, endocarditis, and bone and joint infections, particularly those involving hardware or implanted devices.Case presentationA 65-year-old man presented for washout of his left total knee arthroplasty following a revision 20 days prior. The patient underwent debridement of his left total knee and revision of the left total femur arthroplasty. Daptomycin was initiated empirically due to a previous rash from vancomycin. Operative tissue cultures grew Staphylococcus haemolyticus, Staphylococcus epidermidis and Corynebacterium striatum. Given concern for daptomycin resistance and the reliability of vancomycin susceptibility, daptomycin was discontinued and vancomycin initiated following a graded challenge. Within a few days, the patient developed a diffuse, blanching, erythematous, maculopapular rash and daptomycin was restarted. Over the next 72 h, his rash progressed and he met criteria for drug reaction with eosinophilia and systemic symptoms (DRESS) syndrome. Daptomycin was stopped and oral linezolid initiated; rash improved. C. striatum returned with susceptibility to gentamicin, linezolid, vancomycin and daptomycin. Due to concern for adverse effects on long-term linezolid, daptomycin was restarted and was tolerated for 20 days, at which point purulent drainage from incision increased. The patient underwent another arthroplasty revision and washout. Operative cultures from this surgery were again positive for C. striatum. Repeat C. striatum susceptibilities revealed resistance to daptomycin but retained susceptibility to linezolid. Daptomycin was again changed to linezolid. He completed six weeks of linezolid followed by linezolid 600 mg daily for suppression and ultimately opted for disarticulation.ConclusionsC. striatum has historically been regarded as a contaminant, particularly when grown in tissue culture in the setting of prosthetic joint infection. Based on the available literature and susceptibility patterns, the most appropriate first-line therapy is vancomycin or linezolid. Treatment with daptomycin should be avoided, even when isolates appear susceptible, due to the risk of development of high-level resistance (MIC > 256 µg/mL) and clinical failure.

Isoniazid Resistance and Dosage as Treatment for Patients with Tuberculosis.

The first-line TB antibiotic isoniazid (INH) serves as a central component of combined first-line anti-tuberculosis drug therapy. However, resistance to INH has hindered the functioning of this drug. Resistance is caused by several known and unknown mutations in genes/regions in Mycobacterium tuberculosis (M. tuberculosis), followed by selection of these mutants in the presence of the drug. INH resistance can be categorised as either "high-level" (minimum inhibitory concentration (MIC) of > 1µg/mL to INH) or "low-level" (MIC between 0.1-1.0 µg/L) resistance and is dependent on the specific mutation acquired. The level of resistance is relevant, as INH resistance is often considered to be the first step in development of Multi-Drug Resistant (MDR) and extremely resistant (XDR) TB. Isoniazid is a pro-drug in which first pass metabolism happens via N-acetyltransferase and is fast, intermediate or slow, depending on the genetics of the host. Thus, low-level INH resistance, particularly in the presence of fast metabolism, could allow additional mutations, development of high-level resistance and progression to multi-drug resistance. A structured search of bibliographic databases for peer-reviewed research literature was performed. Set parameters and specific inclusion criteria were used to filter the literature, based on our specified review questions. The quality and relevance of included papers was deduced using standard tools. The relevant content of cited papers was described, and an inferential qualitative content analysis methodology was utilised to analyse the inferences and findings of included studies using a conceptual framework. Seventy-eight papers were included in the review, of which a sub-set (36) of the papers describe how different genetic mutations result in low or high-level resistance to INH. These papers were also used to set up a diagram detailing how each mutation affects INH functionality in order to visualise the interactome of INH and M.tuberculosis A further twenty-eight out of the seventy-eight papers detail the methods for testing for INH resistance, current treatment regimens and factors that influence treatment outcome in order to better understand the role of INH within the current anti-tuberculosis treatment therapy and how its use can be optimised. The findings of this review suggest that low-level INH resistance, in the presence of fast-acetylation, is an underrated component of the global TB epidemic worldwide, and may be a significant problem in terms of treatment outcome and progression to antibiotic resistance. Thus, more research must be done to test whether personalised diagnostics and targeted high dose treatment with INH will reduce the incidence of isoniazid mono-resistant and multi-drug resistant (MDR) tuberculosis.

Current antimicrobial sensitivity pattern of typhoidal salmonellae in a referral diagnostic centre

Background: Infections caused by typhoidal salmonellae are an important public health concern in Pakistan. Inappropriate and injudicious use of fluoroquinolones has reduced their efficacy due to development of high level resistance. <br />Aim: To ascertain the current susceptibility pattern of typhoidal salmonellae thus guiding the physicians for better management of typhoid patients.<br />Materials and Methods: A study was conducted at our institution from January 2012 through December 2013 to investigate current susceptibility pattern of typhoidal salmonellae. <br />Results: Out of 200 isolates, 107 (53.5%) were identified as <em>Salmonella</em> Typhi and 93 (46.5%) as <em>Salmonella</em> Paratyphi A. Sensitivities of <em>Salmonella</em> Typhi were as follows: ampicillin (48.6%), chloramphenicol (45.8%), co-trimoxazole (40.1%), ciprofloxacin (11.2%). Sensitivities of <em>Salmonella</em> Paratyphi A were: ampicillin (80.6%), chloramphenicol (89.2%), co-trimoxazole (90.3%), and ciprofloxacin (16.1%). No resistance was detected against third generation cephalosporins. <br />Conclusions: Typhoidal salmonellae are still entirely susceptible to third generation cephalosporins in our setting. Marked rise in resistance to fluoroquinolones has reduced their empirical usage. Sensitivity of <em>Salmonella</em> Paratyphi A to conventional antityphoid drugs was encouraging.

Inheritance and relative dominance, expressed as toxicity response and delayed development, of phosphine resistance in immature stages of Rhyzopertha dominica (F.) (Coleoptera: Bostrichidae)

Fumigation of stored grain with phosphine (PH3) is used widely to control the lesser grain borer Rhyzopertha dominica. However, development of high level resistance to phosphine in this species threatens control. Effective resistance management relies on knowledge of the expression of resistance in relation to dosage at all life stages. Therefore, we determined the mode of inheritance of phosphine resistance and strength of the resistance phenotype at each developmental stage. We achieved this by comparing mortality and developmental delay between a strongly resistant strain (R-strain), a susceptible strain (S-strain) and their F1 progenies. Resistance was a maternally inherited, semi-dominant trait in the egg stage but was inherited as an autosomal, incompletely recessive trait in larvae and pupae. The rank order of developmental tolerance in both the sensitive and resistant strains was eggs > pupae > larvae. Comparison of published values for the response of adult R. dominica relative to our results from immature stages reveals that the adult stage of the S-strain is more sensitive to phosphine than are larvae. This situation is reversed in the R-strain as the adult stage is much more resistant to phosphine than even the most tolerant immature stage. Phosphine resistance factors at LC50 were eggs 400×, larvae 87× and pupae 181× with respect to reference susceptible strain (S-strain) adults indicating that tolerance conferred by a particular immature stage neither strongly nor reliably interacts with the genetic resistance element. Developmental delay relative to unfumigated control insects was observed in 93% of resistant pupae, 86% of resistant larvae and 41% of resistant eggs. Increased delay in development and the toxicity response to phosphine exposure were both incompletely recessive. We show that resistance to phosphine has pleiotropic effects and that the expression of these effects varies with genotype and throughout the life history of the insect.

Temporal Interplay between Efflux Pumps and Target Mutations in Development of Antibiotic Resistance in Escherichia coli

The emergence of resistance presents a debilitating change in the management of infectious diseases. Currently, the temporal relationship and interplay between various mechanisms of drug resistance are not well understood. A thorough understanding of the resistance development process is needed to facilitate rational design of countermeasure strategies. Using an in vitro hollow-fiber infection model that simulates human drug treatment, we examined the appearance of efflux pump (acrAB) overexpression and target topoisomerase gene (gyrA and parC) mutations over time in the emergence of quinolone resistance in Escherichia coli. Drug-resistant isolates recovered early (24 h) had 2- to 8-fold elevation in the MIC due to acrAB overexpression, but no point mutations were noted. In contrast, high-level (≥ 64× MIC) resistant isolates with target site mutations (gyrA S83L with or without parC E84K) were selected more readily after 120 h, and regression of acrAB overexpression was observed at 240 h. Using a similar dosing selection pressure, the emergence of levofloxacin resistance was delayed in a strain with acrAB deleted compared to the isogenic parent. The role of efflux pumps in bacterial resistance development may have been underappreciated. Our data revealed the interplay between two mechanisms of quinolone resistance and provided a new mechanistic framework in the development of high-level resistance. Early low-level levofloxacin resistance conferred by acrAB overexpression preceded and facilitated high-level resistance development mediated by target site mutation(s). If this interpretation is correct, then these findings represent a paradigm shift in the way quinolone resistance is thought to develop.

Role of Non-Nucleoside Reverse Transcriptase Inhibitors in Treating HIV-Infected Children

The first-generation non-nucleoside reverse transcriptase inhibitors (NNRTIs), efavirenz and nevirapine, fulfil key roles in antiretroviral therapy for HIV-infected paediatric patients, from lowering the incidence of mother-to-child transmission during pregnancy and birth to treatment throughout childhood and adolescence. Both agents have established efficacy, safety and tolerability profiles, and also offer advantages over other classes of therapy in terms of regimen simplicity and availability across different treatment settings. Although the role of NNRTIs in paediatric treatment strategies is largely determined by experience in adult patients, results of the recent phase II/III PENPACT-1 trial in infants and children aged between 30 days and 18 years have shown that there are no significant differences in 4-year virological, immunological or clinical outcomes between NNRTIs and protease inhibitors as first- and second-line agents. However, results from the IMPAACT P1060 study (cohort 2), conducted in resource-limited settings, showed that infants under 36 months unexposed to NNRTIs were significantly more likely to fail treatment when started on a nevirapine-based regimen than those on a lopinavir/ritonavir-based regimen. Unfortunately, the use of efavirenz and nevirapine in children can be limited by rapid development of high-level resistance to one or both agents, which may reduce the availability of viable treatment options, particularly in resource-limited settings. Several therapeutic strategies addressing this issue are currently under investigation, but a significant need for new NNRTI-based treatment options remains. The more recently approved NNRTI, etravirine, has demonstrated efficacy and safety benefits in HIV-1-infected, NNRTI-resistant adult patients, with a higher genetic barrier to the development of resistance relative to the first-generation NNRTIs. Another NNRTI, rilpivirine (TMC278), is approved for use in HIV-1-infected, treatment-naïve adult patients and has demonstrated an improved tolerability profile compared with efavirenz. Although available data on etravirine in children are currently limited, ongoing trials will provide important information on the potential for their use in novel paediatric treatment strategies. This review examines the role of efavirenz and nevirapine in paediatric antiretroviral therapy in children within different treatment settings. In addition, this review also outlines available clinical data on etravirine and rilpivirine in the context of how these antiretrovirals may address some of the limitations of efavirenz and nevirapine in paediatric patients.

Mechanisms of Phosphine Toxicity

Fumigation with phosphine gas is by far the most widely used treatment for the protection of stored grain against insect pests. The development of high-level resistance in insects now threatens its continued use. As there is no suitable chemical to replace phosphine, it is essential to understand the mechanisms of phosphine toxicity to increase the effectiveness of resistance management. Because phosphine is such a simple molecule (PH3), the chemistry of phosphorus is central to its toxicity. The elements above and below phosphorus in the periodic table are nitrogen (N) and arsenic (As), which also produce toxic hydrides, namely, NH3 and AsH3. The three hydrides cause related symptoms and similar changes to cellular and organismal physiology, including disruption of the sympathetic nervous system, suppressed energy metabolism and toxic changes to the redox state of the cell. We propose that these three effects are interdependent contributors to phosphine toxicity.

Developing strategies to manage highly phosphine resistant populations of flat grain beetles in large bulk storages in Australia

Development of high level resistance to phosphine fumigant in flat grain beetles ( Cryptolestes ferrugineus ) in large bulk storages in Australia poses a serious threat to the biosecurity of Australian grain. The level of resistance in this species is the highest ever detected in any stored grain insect pest in Australia with a resistance factor of 875. Laboratory studies showed that at 0.5 mg/L and at 1 mg/L of phosphine 30 and 24 days are required, respectively, to attain population extinction. These doses are currently being tested in field trials for their validation. Moreover, we have developed an action plan in collaboration with project scientists and the major Australian bulk handling companies aimed at eradicating infestations of phosphine resistant flat grain beetles and preventing their spread. The key components of this plan include the use of grain protectants and sulfuryl fluoride to eliminate phosphine-resistant populations, adoption of an intensive hygiene program and monitoring of insect populations through inspection, sampling and resistance testing. Keywords : Flat grain beetle, Cryptolestes ferrugineus , Phosphine, Resistance, Fumigation protocols

News About Influenza B Drug Resistance That Cannot Be Ignored

DURING THE LAST 2 YEARS, THE MEDICAL COMMUnity has been tensely following the emergence of resistance to oseltamivir in strains of influenza A. The clinical relevance and transmissibility of these resistant variants has been unknown, and planning for epidemic and pandemic influenza thus far has ignored the issue due to lack of evidence that it will be of medical consequence. Neuraminidase inhibitors are presently the only reliable antiviral option for treatment of influenza infection: the usefulness of the adamantanes (amantadine and rimantidine) has been virtually eliminated by the development of resistance. Until now, little spontaneous resistance to neuraminidase inhibitors has been documented, and no spontaneously resistant influenza viruses were identified in the world prior to the introduction of the drugs. This may now be changing. In this issue of JAMA, the report from Japan by Hatakeyama and colleagues documenting influenza B viruses with decreased sensitivity to oseltamivir and zanamivir increases concerns about the possibility and clinical significance of resistance. The article describes influenza B viruses partially resistant to neuraminidase inhibitors in individuals who had not been treated with antiviral medications and strongly suggests personto-person transmission of these resistant viruses, either within families or in the community. Japan has the highest use of neuraminidase inhibitors of any country in the world, with more than 90% of the prescriptions being for oseltamivir. While the mutations reported in the resistant isolates in the report are mainly those previously associated with selective pressure from oseltamivir, it is not certain whether the mutated viruses came from other treated individuals in the community or if they emerged spontaneously. The emergence of the variants having low-level resistance discussed in the study is also of concern because influenza B has lower baseline sensitivity to neuraminidase inhibitors compared with influenza A, and the presence of low-level resistance sets the stage for selective pressure for development of high-level resistance. In Australia, 2 influenza viruses with mutations conferring high-level resistance to neuraminidase inhibitors have been isolated from untreated patients. One of these viruses had a mutation in the neuraminidase that is known to confer high-level oseltamivir-specific resistance. Since there is little use of neuraminidase inhibitors in Australia, these viruses likely both arose due to spontaneous mutations. Part of the complacency about neuraminidase inhibitor– resistant influenza has been fueled by experiments in vitro and in animal models that have generally found neuraminidase inhibitor–resistant influenza viruses to be compromised in infectivity and transmissibility. This has led to the belief that significant transmission is unlikely to occur among humans. A 2003 model of the spread of resistant viruses in a pandemic concluded that community spread of such variants would be negligible because of decreases in biological fitness and transmissibility of drug-resistant viruses. However, a recent report predicted that, while use of antiviral drugs could significantly slow or stop transmission in the absence of drug resistance, the emergence of resistance could seriously diminish this benefit, depending on the cost of biological fitness: if there were a modest biological fitness cost and transmissibility were high, the effectiveness of antiviral use would plummet. Even if resistant strains emerge de novo at extremely low frequencies in individuals receiving antiviral drugs for treatment or prophylaxis, these strains may well make a significant contribution in an epidemic or pandemic setting. These concepts, together with the findings reported by Hatakeyama et al suggesting that partiallyresistant influenza B viruses are circulating, mean it is no longer possible to be confident that resistant strains will have little effect on epidemic or pandemic influenza. Resistance to oseltamivir has been predicted by in vitro and structural studies to be more likely to arise than resistance to zanamivir under the selective pressure of drug treatment. Zanamivir is more structurally similar to the natural substrate of neuraminidase and fits directly into the active site. For oseltamivir, a rearrangement of amino acids in the active site is necessary to accommodate the drug’s hydrophobic side chain; mutations that prevent this rearrangement might lead to resistance to oseltamivir but not to zanamivir. It has also

Comparative Toxicity of Fumigants and a Phosphine Synergist Using a Novel Containment Chamber for the Safe Generation of Concentrated Phosphine Gas

BackgroundWith the phasing out of ozone-depleting substances in accordance with the United Nations Montreal Protocol, phosphine remains as the only economically viable fumigant for widespread use. However the development of high-level resistance in several pest insects threatens the future usage of phosphine; yet research into phosphine resistance mechanisms has been limited due to the potential for human poisoning in enclosed laboratory environments.Principal FindingsHere we describe a custom-designed chamber for safely containing phosphine gas generated from aluminium phosphide tablets. In an improvement on previous generation systems, this chamber can be completely sealed to control the escape of phosphine. The device has been utilised in a screening program with C. elegans that has identified a phosphine synergist, and quantified the efficacy of a new fumigant against that of phosphine. The phosphine-induced mortality at 20°C has been determined with an LC50 of 732 ppm. This result was contrasted with the efficacy of a potential new botanical pesticide dimethyl disulphide, which for a 24 hour exposure at 20°C is 600 times more potent than phosphine (LC50 1.24 ppm). We also found that co-administration of the glutathione depletor diethyl maleate (DEM) with a sublethal dose of phosphine (70 ppm, <LC5), results in a doubling of mortality in C. elegans relative to DEM alone.ConclusionsThe prohibitive danger associated with the generation, containment, and use of phosphine in a laboratory environment has now been substantially reduced by the implementation of our novel gas generation chamber. We have also identified a novel phosphine synergist, the glutathione depletor DEM, suggesting an effective pathway to be targeted in future synergist research; as well as quantifying the efficacy of a potential alternative to phosphine, dimethyl disulphide.

Comment on: Quinolone resistance determinant qnrA3 in clinical isolates of Salmonella in 2000-2005 in Hong Kong

Sir, In their recent correspondence, Chu et al. noted the first observation of qnrA3 in salmonellae in Hong Kong, underlining the spread and diversity of plasmidic quinoloneresistance determinants worldwide. Such determinants now include qnrA1–5, qnrB and qnrS, some of which, at least, protect DNA gyrase, as well as the fluoroquinolone-acetylating variant of the aminoglycoside-modifying enzyme AAC(60)Ib, named AAC(60)Ib-cr. These proteins may influence the development of high-level resistance to at least some fluoroquinolones. In our recent article, we sought to explore the unexplained significant association between chromosomal fluoroquinolone resistance and plasmidic resistances to other antibiotic classes (such as ESBLs), which is seen in several bacterial genera. We were pleased to note that Chu et al. cited our article, but would like to clarify a misinterpretation of our hypothesis. Multiresistance plasmids can encode toxin–antitoxin systems to prevent their own loss from the cell, and some toxins target DNA gyrase. We asked whether the development of quinolone resistance by plasmid-bearing strains might be associated with such DNA gyrase-targeting toxins. We speculated on whether a cell might escape addiction to toxin–antitoxin systems by developing resistance to the toxin. Mutational events in DNA gyrase could play a role in cells escaping addiction to plasmids with gyrase-targeting toxins, and we suggested that these mutations might fortuitously confer a degree of cross-resistance to quinolones. Notably, whilst at least three qnrA variants likely originated from the chromosome of Shewanella algae, and although Qnr-type proteins inhibit/protect DNA gyrase, their precise cellular role(s) remains unknown. We would like to emphasize that we did not state, or wish to imply, that Qnr proteins could be part of a toxin–antitoxin pair as was attributed to our article by Chu et al. Rather, we postulated that gyrase ‘inhibitors/protectors’ such as Qnr might give some crossresistance to gyrase-targeting toxins in addition to the more obvious resistance to quinolones. By so doing these proteins might open a ‘window’ and influence the frequency at which cells could eject the toxin–antitoxin encoding replicons without suffering apoptosis, thereby providing the host organism with a toxin–antitoxin evasion mechanism. This might provide a competitive advantage in demanding psychrophilic aquatic environments, the natural environment of S. algae, by allowing enhanced ejection of ‘accessory’ replicons.

Identification and analysis of fitness of resistance mutations against the HCV protease inhibitor SCH 503034

HCV NS3 protease variants resistant to the protease inhibitor SCH 503034 were selected. Three mutations, T54A, V170A and A156S mutations conferred low to moderate levels of resistance (<20-fold). Longer exposure (>10 passages) or selection with higher levels of compound led to the selection of a more resistant variant, A156T (>100-fold). [Lin, C., Lin, K., Luong, Y.P., Rao, B.G., Wei, Y.Y., Brennan, D.L., Fulghum, J.R., Hsiao, H.M., Ma, S., Maxwell, J.P., Cottrell, K.M., Perni, R.B., Gates, C.A., Kwong, A.D., 2004. In vitro resistance studies of hepatitis C virus serine protease inhibitors, VX-950 and BILN 2061: structural analysis indicates different resistance mechanisms. J. Biol. Chem. 279(17), 17508–17514; Lu, L., Pilot-Matias, T.J., Stewart, K.D., Randolph, J.T., Pithawalla, R., He, W., Huang, P.P., Klein, L.L., Mo, H., Molla, A., 2004. Mutations conferring resistance to a potent hepatitis C virus serine protease inhibitor in vitro. Antimicrob. Agents Chemother. 48(6), 2260–2266.] Combination with IFN-α drastically reduced the number of emergent colonies. Resistant colonies showed no change in sensitivity to IFN-α. Although the A156T mutation conferred the highest level of resistance to SCH 503034, it significantly reduced the colony formation efficiency (CFE) of the mutant replicon RNA, and rendered replicon cells less fit than those bearing wild-type replicons. Replicon cells bearing mutation A156S were less fit than wild-type in co-culture growth competition assays but showed no impact on CFE. The V170A mutation, on the other hand, did not affect replicon fitness in either assay, which was consistent with its emergence as the dominant mutant after 12 months of continuous selection. The reduced fitness of the most resistant variant suggests that it may be rare in naïve patients and that development of high-level resistance may be slow. Combination therapy with IFN-α should also greatly reduce the potential emergence of resistance.

Seven years survey of susceptibility to marbofloxacin of bovine pathogenic strains from eight European countries

This study was conducted from 1994 to 2001 to determine the susceptibility of bovine pathogenic bacteria to marbofloxacin (a third generation fluoroquinolone used only in individual administration for animals). Strains originated in bovine diseases from eight European countries. They were isolated from gut infections ( Escherichia coli, salmonellae), mastitis ( E. coli, Staphylococcus aureus, Streptococcus uberis, S. agalactiae, S. dysgalactiae) and respiratory diseases ( Mannheimia haemolytica, Pasteurella multocida, Haemophilus somnus). There was no change in the MIC distributions for each species after the launch of marbofloxacin in 1997. In E. coli, a resistant population was present before the use of marbofloxacin having been induced by co- or cross-resistance to other antibiotics used previously. Over this period the only a significant change seen was an increase in MIC 90 of E. coli from the gut (1.275 μg/ml in 1994/1995 to 5.098 μg/ml in 2001). All the salmonellae were susceptible to marbofloxacin with a MIC 90 = 0.073 μg/ml in 2001 without development of high level resistance. The use of marbofloxacin seems not to have favoured a significant increase and spreading of resistant bacteria.