International Journal of Antimicrobial Agents

Title

2010-09-01

Evolving trends in Streptococcus pneumoniae resistance: implications for therapy of community-acquired bacterial pneumonia

Ronald N. Jones, Michael R. Jacobs, Helio S. Sader — 2010-06-18

Abstract: Pneumonia is a major infectious disease associated with significant morbidity, mortality and utilisation of healthcare resources. Streptococcus pneumoniae is the predominant pathogen in community-acquired pneumonia (CAP), accounting for 20–60% of bacterial cases. Emergence of multidrug-resistant S. pneumoniae has become a significant problem in the management of CAP. Although pneumococcal conjugate vaccine usage in children has led to significant decreases in morbidity and mortality due to S. pneumoniae in all age groups, disease management has been further complicated by the unexpected increase in resistant serotypes, such as 19A, in some regions. Until rapid and accurate diagnostic tests become available, initial treatment of CAP will remain empirical. Thus, selection of appropriate antimicrobial therapy for CAP must be based on prediction of the most likely pathogens and their local antimicrobial susceptibility patterns. This article reviews information on antimicrobial resistance patterns amongst S. pneumoniae and implications for managing CAP.

Guideline for phenotypic screening and confirmation of carbapenemases in Enterobacteriaceae

2010-07-05

Abstract: Adequate detection of carbapenemase-producing Enterobacteriaceae is crucial for infection control measures and appropriate choice of antimicrobial therapy. This guideline aims to improve the detection of carbapenemase-producing Enterobacteriaceae in the routine setting of clinical microbiology laboratories. Detection of carbapenemases in Enterobacteriaceae includes a screening step followed by a genotypic and optional phenotypic confirmatory step. For all Enterobacteriaceae, the meropenem screening breakpoint to detect carbapenemases is set at ≥0.5mg/L or a zone diameter of ≤23mm (10μg disk loading). For Escherichia coli, Klebsiella spp., Salmonella spp., Enterobacter spp. and Citrobacter spp., the imipenem screening breakpoint is set at ≥2mg/L or a zone diameter ≤21mm. Ertapenem is not advised as an indicator carbapenem as it has a lower specificity compared with imipenem and meropenem. On the first isolate from a patient with a positive carbapenemase screen test, a polymerase chain reaction (PCR)-based test should be performed to detect carbapenemase genes. However, if genotypic confirmation is not immediately available, phenotypic confirmation tests should be performed to avoid delayed reporting of carbapenemase-producers to the clinic. Recommended phenotypic confirmation tests are the modified Hodge test as well as carbapenemase inhibition tests with boronic acid for Ambler class A carbapenemases and with ethylene diamine tetra-acetic acid (EDTA) or dipicolinic acid for metallo-carbapenemases.

Ambroxol interferes with Pseudomonas aeruginosa quorum sensing

Qi Lu, Jialin Yu, Xiqiang Yang, Jiarong Wang, Lijia Wang, Yayin Lin, Lihua Lin — 2010-06-28

Abstract: The mucolytic agent ambroxol has been reported to interfere with the formation of Pseudomonas aeruginosa-derived biofilms in addition to reducing alginate production by undefined mechanisms. Since quorum sensing is a key regulator of virulence and biofilm formation, we examined the effects of ambroxol on P. aeruginosa PAO1 wild-type bacterial clearance rates, adhesion profiles and biofilm formation compared with the quorum sensing-deficient, double-mutant strains ΔlasR ΔrhlR and ΔlasI ΔrhlI. Data presented in this report demonstrated that ambroxol treatment reduced survival rates of the double-mutant strains compared with the wild-type strain in a dose-dependent manner even though the double-mutants had increased adhesion in the presence of ambroxol compared with the wild-type strain. The PAO1 wild-type strain produced a significantly thicker biofilm (21.64±0.57μm) compared with the biofilms produced by the ΔlasR ΔrhlR (7.36±0.2μm) and ΔlasI ΔrhlI (6.62±0.31μm) isolates. Ambroxol treatment reduced biofilm thickness, increased areal porosity, and decreased the average diffusion distance and textual entropy of wild-type and double-mutant strains. However, compared with the double-mutant strains, the changes observed for the wild-type strain were more clearly defined. Finally, ambroxol exhibited significant antagonistic quorum-sensing properties, suggesting that it could be adapted for use clinically in the treatment of cystic fibrosis and to reduce biofilm formation and in the colonisation of indwelling devices.

Dichotomous selection of high-level oxacillin resistance in Staphylococcus aureus by fluoroquinolones

Axel Dalhoff, Sabine Schubert — 2010-07-15

Abstract: The objective of this study was to determine whether exposure of Staphylococcus aureus to early (ciprofloxacin or levofloxacin) and a recent fluoroquinolone (moxifloxacin) has differential potential as a mutator or selector for meticillin resistance. The potential of fluoroquinolones to act as mutators or selectors was studied in 24 strains each of healthcare-associated meticillin-susceptible S. aureus (MSSA) and meticillin-resistant S. aureus (MRSA) as well as 6 strains of community-acquired MRSA. Mutator or selector potential was studied first by exposing isolates to 0.5× the fluoroquinolone minimal inhibitory concentration (MIC) and screening for either single-step fluoroquinolone resistance or high-level oxacillin resistance; second, by exposing the heteroresistant MRSA P8 parent strain as well as fluoroquinolone-resistant subpopulations derived from strain P8 to constant fluoroquinolone concentrations ranging from 0.015mg/L to 128mg/L; and third, by exposing the heteroresistant MRSA population of strain P8 to fluctuating concentrations of ciprofloxacin, levofloxacin and moxifloxacin simulating oral doses of 500mg twice a day, 500mg once daily (qd) and 400mg qd, respectively, compared with amoxicillin/clavulanic acid 500mg three times a day. Total viable counts and subpopulations resistant to 2×, 4× and 8× the fluoroquinolone MICs and to 32, 64 and 128mg/L oxacillin [high-level oxacillin (hl-OXA)-resistant] were quantitated. None of the fluoroquinolones acted as a mutator; ciprofloxacin and levofloxacin selected for hl-OXA resistance, whereas moxifloxacin selected towards hl-OXA resistance by one order of magnitude less frequently. The P8 parent and fluoroquinolone-resistant subpopulations were eliminated by ciprofloxacin or levofloxacin concentrations >10-fold higher than the MICs, whereas moxifloxacin eliminated all subpopulations by concentrations 2–3-fold the MIC. Finally, exposure of P8 to fluctuating amoxicillin/clavulanic acid, ciprofloxacin and levofloxacin concentrations, respectively, caused a rapid selection of fluoroquinolone and hl-OXA resistance. Moxifloxacin reduced total viable counts rapidly, thus preventing the emergence of resistant subpopulations. In conclusion, fluoroquinolones do not act as mutators towards hl-OXA resistance. However, ciprofloxacin and levofloxacin are potent selectors of hl-OXA resistance, whereas moxifloxacin is a poor selector. In contrast to ciprofloxacin and levofloxacin, moxifloxacin exerts a high bactericidal activity against staphylococci, thus minimising the probability for selection of resistance. Thus, fluoroquinolones exert a dichotomous MRSA-selective potential in heteroresistant MRSA.

In silico genetic correlations of multidrug efflux pump gene expression in Staphylococcus aureus

2010-07-05

Abstract: Regulatory mechanisms for chromosomal genes encoding multidrug resistance (MDR) efflux pumps (EPs) in Staphylococcus aureus are poorly defined. Microbiological, quantitative gene expression, mRNA half-life and genome data for 11 strains of S. aureus combined with bioinformatic analyses were used to identify correlates of increased MDR EP gene expression. The presence of qacA/B and/or increased expression of one to two MDR EP genes were identified in eight strains. Microbiological and gene expression data correlated in four instances, existing knowledge of the substrate specificity of NorC resulted in correlation in two others, and a transcriptional/translational disconnect is possible for the remaining two. In silico analyses and mRNA half-life determinations linked insertions of nucleotide repeats 3′ to the −10 motif of the norA promoter with increased promoter activity. Mutations in the 5′-untranslated and/or coding regions were identified that may affect transcription efficiency. Substitutions of residues in the helix-turn-helix (HTH) motif of NorG may augment its positive regulation of norB. The correlations proposed provide a guide for further experimentation leading to a better understanding of MDR EP gene expression in this important pathogen.

Antimicrobial activity of DW286 against Streptococcus pneumoniae

Hee Soo Park, Sung Ji Jung, Dong-Rack Choi, Jin-Hwan Kwak — 2010-06-07

Abstract: DW286 is a novel broad-spectrum fluoroquinolone with excellent antipneumococcal activity. The in vitro activity of DW286 was evaluated against quinolone-susceptible and -resistant Streptococcus pneumoniae and was compared with the activities of reference compounds. Among the tested agents, DW286 showed the most potent antibacterial activity against 94 quinolone-susceptible strains [minimum inhibitory concentration (MIC) 0.008–0.03mg/L]. Against 23 quinolone-resistant S. pneumoniae with known resistance mechanisms, DW286 also had the lowest MICs of all the tested quinolones [MIC at which 90% of isolates were inhibited (MIC90)=0.5mg/L], followed by ciprofloxacin, sparfloxacin, moxifloxacin and gemifloxacin. The in vivo activity of DW286 against penicillin-susceptible and -resistant S. pneumoniae was more effective than that of gemifloxacin.

Assessment of the activity of RND-type multidrug efflux pumps in Pseudomonas aeruginosa using tetraphenylphosphonium ions

Rimantas Daugelavičius, Andrius Buivydas, Ana Senčilo, Dennis H. Bamford — 2010-05-20

Abstract: Multidrug resistance (MDR) pumps are one of the major causes of antibiotic resistance in Pseudomonas aeruginosa. Thus, fast and reliable methods are needed to assay the efficiency of MDR pumps in these bacteria. In this study, it was demonstrated that a membrane voltage (Δψ) indicator tetraphenylphosphonium (TPP+) in combination with the efflux pump inhibitor phenylalanyl-arginyl-β-naphthylamide can be used to monitor the activity of resistance–nodulation–cell division (RND)-type efflux pumps in P. aeruginosa. By controlling the outer membrane permeability and Δψ, electrochemical measurements of RND pump activity in real time were performed. It was demonstrated that the composition of the medium, the presence of nutrients and the level of aeration affect the efficiency of the TPP+-extruding activity of P. aeruginosa, urging the standardisation of experimental conditions to obtain quantitative and comparative results.

Hypermutability in clinical isolates of Klebsiella pneumoniae is uncommon and is unrelated to ciprofloxacin resistance

S. Aathithan, G.L. French — 2010-06-14

Abstract: We investigated hypermutability in Klebsiella pneumoniae and its association with ciprofloxacin resistance and mutations in the quinolone resistance-determining region (QRDR). Sixty-four strains of K. pneumoniae isolated in London, UK, between 1995 and 2002 with widely differing ciprofloxacin minimum inhibitory concentrations (MICs) and known gyrA and parC sequences were tested for mutation frequencies by selection with rifampicin. Only three hypermutable (frequency ≥10−6) strains were identified, with ciprofloxacin MICs of 0.25μg/mL, 8μg/mL and 64μg/mL. There was no relationship between hypermutation and the ciprofloxacin MIC or QRDR mutations. Screening selected strains with streptomycin did not reveal any hypermutators, and screening with ciprofloxacin identified only two of the three hypermutators identified by rifampicin. Hypermutation in K. pneumoniae is uncommon and does not contribute to accumulation of QRDR mutations or directly to ciprofloxacin resistance.

Synergistic activities between carbapenems and other antimicrobial agents against Acinetobacter baumannii including multidrug-resistant and extensively drug-resistant isolates

Pattarachai Kiratisin, Anucha Apisarnthanarak, Srirumpa Kaewdaeng — 2010-06-14

Abstract: Treatment options for multidrug-resistant (MDR) and extensively drug-resistant (XDR) Acinetobacter baumannii have been seriously limited and may require combination antimicrobial therapy. In this study, we searched for synergistic activity between carbapenems (doripenem, imipenem and meropenem) and various non-traditional agents (cefoperazone/sulbactam, doxycycline, rifampicin, netilmicin and moxifloxacin) against 40 A. baumannii clinical isolates, including MDR and XDR isolates. The results showed that combination of each carbapenem with cefoperazone/sulbactam, based on the Etest method, demonstrated synergy more frequently (17.5–32.5%) than the other tested agents, which may suggest a role in combination therapy against highly resistant A. baumannii.

Prevalence of decreased susceptibility to triclosan in Salmonella enterica isolates from animals and humans and association with multiple drug resistance

Justin L. Copitch, Rebekah N. Whitehead, Mark A. Webber — 2010-06-14

Abstract: Previous laboratory studies have implicated triclosan as a possible selective force driving resistance to multiple antibiotics and have identified a number of triclosan resistance mechanisms in Salmonella enterica. The aim of this work was to determine the prevalence of decreased susceptibility to triclosan in a panel of human and animal isolates of S. enterica and to identify the mechanisms of triclosan resistance in these strains. Over 400 animal and human isolates of non-typhoidal Salmonella were screened for decreased susceptibility to triclosan and a panel of antibiotics. The prevalence of decreased susceptibility to triclosan was ca. 4%. Of the isolates with decreased triclosan susceptibility, 56% were multidrug-resistant (MDR) compared with 12% of triclosan-sensitive isolates. MDR and triclosan-resistant strains showed increased efflux activity compared with strains with reduced susceptibility to triclosan alone. No high-level triclosan resistance was seen in this panel of isolates. A reservoir of strains with low-level decreased triclosan susceptibility is present in animals and humans. These isolates are MDR as a result of generic mechanisms of antimicrobial resistance and do not carry specific mutations within fabI.

Presence of plasmid-mediated quinolone resistance in Klebsiella pneumoniae and Escherichia coli isolates possessing blaVIM-1 in Greece

I. Galani, M. Souli, N. Mitchell, Z. Chryssouli, H. Giamarellou — 2010-06-28

Abstract: Amongst nalidixic acid-resistant, ciprofloxacin-susceptible Escherichia coli and Klebsiella pneumoniae clinical isolates recovered over a 5-month period from inpatients and outpatients of Attikon University General Hospital (Athens, Greece), only one E. coli was positive for qnrB2 and one K. pneumoniae was positive for qnrA1. Both isolates were extended-spectrum β-lactamase-negative, metallo-β-lactamase-positive and carried the blaVIM-1 gene. Neither of the isolates had mutations in gyrA and parC or carried aac(6′)-Ib-cr or qepA. The K. pneumoniae isolate also harboured blaCMY-13 on the same transferable plasmid with qnrA1. This is the first report of a qnrA1-positive K. pneumoniae and qnrB2-positive E. coli harbouring a concurrent blaVIM-1 gene.

Emergence of clinical strains of Mycoplasma genitalium harbouring alterations in ParC associated with fluoroquinolone resistance

2010-06-28

Abstract: Surveillance for antimicrobial resistance in Mycoplasma genitalium clinical strains is extremely limited as culturing of strains from clinical specimens is still difficult. We therefore conducted a non-cultural assessment of fluoroquinolone resistance of M. genitalium clinical strains by analysing the quinolone-resistance determining regions (QRDRs) of the gyrA and parC genes. The QRDRs amplified from M. genitalium DNA taken from urine specimens of 28 men with non-gonococcal urethritis positive for M. genitalium by polymerase chain reaction were sequenced. An amino acid change (Phe-108→Iso) in GyrA was found in one specimen, and the same change was accompanied by an amino acid change (Lys-97→Arg) in ParC in another specimen. A single amino acid change (Ser-83→Asn, Asp-87→Tyr or Asp-87→Val) in ParC was also found in three other respective specimens without alterations in GyrA. No alterations in GyrA and ParC were found in the remaining 23 specimens. The alterations of Ser-83→Asn, Asp-87→Tyr and Asp-87→Val in ParC found in 3 (10.7%) of 28 specimens were analogous to those commonly observed in fluoroquinolone-resistant mutants of other Mycoplasma and Ureaplasma spp. M. genitalium harbouring mutations associated with fluoroquinolone resistance in the parC gene may have emerged clinically and the prevalence may be ca. 10% in Japan.

Characteristics of clinical isolates of Acinetobacter genomospecies 10 carrying two different metallo-β-lactamases

2010-07-05

Abstract: Acquired metallo-β-lactamase (MBL) production is an important mechanism of carbapenem resistance. To our knowledge, carriage of two different MBLs has not been described previously in Acinetobacter spp. We present the characteristics of five Acinetobacter isolates carrying two different MBL genes. The species of all five Acinetobacter isolates with two different MBL genes were Acinetobacter genomospecies 10, and blaIMP-1, blaVIM-2 and blaSIM-1 may coexist in this species. Minimum inhibitory concentrations (MICs) of imipenem for all five isolates with two different MBLs were ≥32μg/mL, whilst those for two segregants that lost both MBLs were 0.5μg/mL. The presence of MBL gene-carrying integrons with identical structures suggested the easily transferable nature of the elements, whilst instability of the MBL genes indicated potentially erroneous phenotypic and genetic characterisation.

The antimicrobial peptide Ci-MAM-A24 is highly active against multidrug-resistant and anaerobic bacteria pathogenic for humans

Henning Fedders, Rainer Podschun, Matthias Leippe — 2010-06-07

Abstract: Ci-MAM-A24, a synthetic antimicrobial peptide derived from a peptide precursor from immune cells of the marine invertebrate Ciona intestinalis, has been shown to be potently active against representatives of Gram-positive and Gram-negative bacteria by permeabilising their cytoplasmic membrane. In the present study, the activity of Ci-MAM-A24 against different bacterial pathogens frequently causing therapeutic problems was tested. In particular, the killing capacity of Ci-MAM-A24 against clinically important anaerobic bacteria as well as multiresistant aerobic strains such as meticillin-resistant Staphylococcus aureus (MRSA), vancomycin-resistant enterococci, extended-spectrum β-lactamase-producers and multiple-resistant Pseudomonas aeruginosa strains was monitored. Virtually all strains proved to be highly susceptible to Ci-MAM-A24 at low concentrations [minimum bactericidal concentration (MBC)<10μg/mL].

Antimicrobial susceptibility profiles of meticillin-susceptible and -resistant Staphylococcus aureus: focus on daptomycin minimum inhibitory concentrations at a tertiary care centre in Mumbai, India

Namita D'Souza, Anjali Shetty, Ajita Mehta, Camilla Rodrigues — 2010-07-01

Abstract: In this study, daptomycin minimum inhibitory concentrations (MICs) for Staphylococcus aureus isolates were determined using Etest strips and were correlated with staphylococcal cassette chromosome mec (SCCmec) types and Panton–Valentine leukocidin (PVL) gene positivity. In total, 60 meticillin-resistant S. aureus (MRSA) and 60 meticillin-susceptible S. aureus (MSSA) isolates from clinical samples were subjected to antimicrobial susceptibility testing by disk diffusion according to Clinical and Laboratory Standards Institute guidelines, polymerase chain reaction (PCR) detection of PVL and mecA genes, and SCCmec typing. Daptomycin MICs were determined using Etest strips. The mecA gene was present in all MRSA isolates and was absent in 59 MSSA isolates. The PVL gene was present in 41 MRSA isolates and 25 MSSA isolates. Amongst the MRSA isolates, 10 were SCCmec type III, 27 were SCCmec IV and 23 were SCCmec V. Moreover, 26 SCCmec IV and 15 SCCmec V isolates were PVL-positive. All SCCmec III isolates were multidrug-resistant and PVL-negative. Daptomycin MICs ranged from 0.047μg/mL to 1μg/mL for MRSA and from 0.19μg/mL to 1μg/mL for MSSA. All MRSA and MSSA isolates were susceptible to daptomycin. Although SCCmec III MRSA and PVL-positive MSSA were resistant to more antimicrobial classes than SCCmec IV and V MRSA and PVL-negative MSSA, there does not appear to be a significant correlation between SCCmec types, PVL positivity and daptomycin MICs. MICs were not as low as expected for a newly introduced drug.

Chicken cathelicidin-2-derived peptides with enhanced immunomodulatory and antibacterial activities against biological warfare agents

2010-07-15

Abstract: Host defence peptides (HDPs) are considered to be excellent candidates for the development of novel therapeutic agents. Recently, it was demonstrated that the peptide C1-15, an N-terminal segment of chicken HDP cathelicidin-2, exhibits potent antibacterial activity while lacking cytotoxicity towards eukaryotic cells. In the present study, we report that C1-15 is active against bacteria such as Bacillus anthracis and Yersinia pestis that may potentially be used by bioterrorists. Substitution of single and multiple phenylalanine (Phe) residues to tryptophan (Trp) in C1-15 resulted in variants with improved antibacterial activity against B. anthracis and Y. pestis as well as decreased salt sensitivity. In addition, these peptides exhibited enhanced neutralisation of lipopolysaccharide (LPS)-induced release of pro-inflammatory cytokines in human peripheral blood mononuclear cells (PBMCs). The antibacterial and LPS-neutralising activities of these C1-15-derived peptides are exerted at concentrations far below the concentrations that are toxic to human PBMCs. Taken together, we show that Phe→Trp substitutions in C1-15 variants enhances the antibacterial and LPS-neutralising activities against pathogenic bacteria, including those that may potentially be used as biological warfare agents.

Antiparasitic activity of alkaloids from plant species of Papua New Guinea and Australia

Liza S. Fernandez, Melissa L. Sykes, Katherine T. Andrews, Vicky M. Avery — 2010-06-28

Abstract: New drugs are needed to help overcome the increasing problem of drug resistance in parasites that cause diseases such as malaria and trypanosomiasis. In this study, alkaloid compounds isolated from extracts of the plants Flindersia amboinensis, Stephania zippeliana and Voacanga papuana from Papua New Guinea and Flindersia acuminata from Australia were examined for their antiparasitic activity against Plasmodium falciparum strains and Trypanosoma brucei brucei as well as their cytotoxicity against the mammalian cell lines HEK 293 and HeLa. The most active compound, dimethylisoborreverine (DMIB), showed submicromolar activity, with 50% inhibitory concentration (IC50) values between 20nM and 810nM both against drug-sensitive and drug-resistant P. falciparum strains, along with moderate selectivity against T. b. brucei and mammalian cells. Stage specificity studies revealed that P. falciparum trophozoite-stage parasites were more susceptible to DMIB than ring- or schizont-stage parasites. DMIB-treated trophozoites showed changes in food vacuole morphology, with an apparent reduction in haemozoin formation that does not appear to be inhibited via the direct binding of haem. These findings suggest a potential for indole alkaloids from Flindersia spp. as new antiparasitic agents.

Enhancement of the antibacterial properties of silver nanoparticles using β-cyclodextrin as a capping agent

Swarna Jaiswal, Brendan Duffy, Amit Kumar Jaiswal, Niall Stobie, Patrick McHale — 2010-06-28

Abstract: Silver nanoparticles (AgNPs) were synthesised by reducing silver salts using NaBH4 followed by capping with varying concentrations of β-cyclodextrin (β-CD) and were physically characterised. Antibacterial activity against Escherichia coli, Pseudomonas aeruginosa and Staphylococcus aureus was determined by a microtitre well method. The AgNPs were spherical under transmission electron microscopy, whilst dynamic light scattering showed average diameters of capped particles to be smaller (4–7nm) than their uncapped equivalents (17nm). Capped particles demonstrated superior photostability when exposed to intense ultraviolet radiation for 4h as well as significantly (P<0.05) higher (up to 3.5-fold) antibacterial activity. The influence of β-CD concentration was seen to delay bacterial growth, indicating that a Trojan horse mechanism may be occurring owing to bacterial carbohydrate affinity, thereby enhancing silver ion absorption.

Risk factors and treatment outcomes of community-onset bacteraemia caused by extended-spectrum β-lactamase-producing Escherichia coli

2010-06-28

Abstract: The purpose of this study was to identify risk factors for extended-spectrum β-lactamase (ESBL)-producing Escherichia coli amongst community-onset bacteraemia and to evaluate treatment outcomes. From the database of a nationwide surveillance programme for bacteraemia, data from patients with community-onset E. coli bacteraemia were analysed. Patients with ESBL-producing E. coli bacteraemia were compared with those with non-ESBL-producing bacteraemia. The overall proportion of ESBL-producers was 9.5% (82/865) amongst community-onset E. coli bacteraemia cases. Healthcare-associated infection, underlying liver disease and primary bacteraemia were significant independent factors associated with ESBL-producing E. coli bacteraemia (P<0.05). There was a trend toward mortality being higher in the ESBL group compared with the non-ESBL group (15.0% vs. 7.6%; P=0.096). ESBL production was found to be an independent factor associated with mortality after adjusting for confounding variables (odds ratio=2.99, 95% confidence interval 1.01–8.84; P=0.048), along with severe sepsis, higher Pitt bacteraemia score, primary bacteraemia, pneumonia and underlying liver disease (P<0.05). ESBL-producing E. coli is a significant cause of bacteraemia, even in patients with community-onset infections, predicting higher mortality, particularly in patients with primary bacteraemia, underlying liver disease or healthcare-associated infection.

A novel in vivo model to screen antimicrobial compounds

Khadijo Osman, Naveed Ahmed Khan — 2010-04-30

With an estimated annual mortality of more than 14 million people and a growing incidence of drug-resistant disease, there is an urgent need for new drugs to assist in the global control of infectious diseases . In the conventional approach to drug discovery, compounds are initially optimised for activity against microbial pathogens in vitro and subsequently assessed for in vivo activity in animal models during pre-clinical development. A range of assays are available to assess the efficacy of drugs in vitro, however one of the limitations of this approach is that compounds that are particularly active in vivo may be overlooked in the early discovery phase. Even with in vitro efficacy, potential antimicrobials or vaccine targets need to be tested in animals. To this end, although vertebrate model systems are seen as immediately more relevant, they are expensive, labour intensive, require legislative adherence and, more importantly, have ethical concerns. It is proposed here that the use of an invertebrate model at an early stage can offer several advantages in terms of speed, cost, technical convenience and ethical acceptance. Another attractive aspect of the locust model is that large numbers can be used in a single experiment; up to 50 individual locusts can be kept in a single cage, thus multiple experimental variables can be tested. Our recent studies have clearly shown that central nervous system infections due to Escherichia coli K1 and Acanthamoeba spp. can be modelled in the locust, thus alleviating the need to use newborn Sprague–Dawley rats and BALB/c mice, respectively, at an early stage . The aim of the present study was to test the usefulness of the newly developed locust model for the in vivo assessment of antibacterial compounds.

In vitro activity of tigecycline against carbapenemase-producing Acinetobacter baumannii

2010-06-18

Acinetobacter baumannii infections present a challenge to clinicians, with the incidence of multidrug-resistant (MDR) isolates increasing worldwide . Carbapenem resistance in this species is most often due to oxacillinases (OXA), which can be divided into four main subgroups, namely the intrinsic OXA-51-like and the acquired carbapenemases OXA-23-like, OXA-40-like and OXA-58-like .

Enemy within: strategies to kill ‘superbugs’ in hospitals

Naveed Ahmed Khan, Ruqaiyyah Siddiqui, Hany Elsheikha — 2010-06-14

Record numbers of patients are dying in hospitals and nursing homes after contracting superbugs such as meticillin-resistant Staphylococcus aureus (MRSA) and Clostridium difficile . MRSA and C. difficile were linked to more than 8000 deaths in England and Wales in 2006, up from 5300 the previous year, with the vast majority dying in hospitals. Overuse of antibiotics is considered as the basis in fuelling the spread of hospital infections . In a bid to halt this process, the Department of Health has invested £270 million to tackle infection control in the National Health Service (NHS) and doctors are being asked, repeatedly, to stop prescribing antibiotics for common conditions such as coughs, colds and sore throats . Whilst the introduction of ‘zero tolerance’ rules on hand hygiene to prevent the spread of superbugs as well as the use of high-concentration antibacterials in hospitals had some success, albeit at a slow rate, it is only a matter of time before bacteria will develop resistance against the chemotherapeutic artillery that we throw at them. More urgently, Gram-negative bacteria such as Acinetobacter, Pseudomonas and Burkholderia are now considered as the emerging superbugs, sometimes causing untreatable infections in particular patient groups such as cystic fibrosis patients, and have become a major threat in our fight against bacterial diseases. Thus, there is a continuous need to find additional strategies to confront this menace. A major factor that has hindered our success in preventing the spread of infections in hospitals and nursing homes is our inability to identify correctly the source of bacterial pathogens. To this end, several lines of evidence suggest that Acanthamoeba, a free-living protozoan, harbour diverse microbial organisms including viruses, yeast, protozoa and bacteria, some of which are potential human pathogens (reviewed in ). Acanthamoeba is one of the most ubiquitous protozoa with two stages in its life cycle, a vegetative trophozoite stage and a dormant cyst stage. Cysts are highly resistant to physical, chemical (antibiotics and biocides) and radiological conditions and can be airborne. Acanthamoeba has been shown to host a variety of bacterial pathogens, including Legionella, MRSA, Escherichia coli O157, Burkholderia, Clostridium, Coxiella, Salmonella, Pseudomonas, Vibrio, Helicobacter, Listeria, Chlamydophila, Rickettsia, Porphyromonas, Prevotella, Shigella, Pasteurella, Campylobacter and Mycobacterium, to name a few . The hardiness of cysts protects bacteria against harsh environments. The precise nature of this symbiosis is not clear but these findings do suggest that such interactions enable pathogenic bacteria to survive in a variety of hostile conditions such as the use of disinfectants in a hospital setting, which may lead to their transmission to susceptible hosts to establish infection. The ability of pathogenic bacteria such as MRSA or Legionella to survive and multiply inside Acanthamoeba suggests that amoeba act not only as a vector but also as a reservoir. In addition, cysts may provide resistance to intracellular bacteria for the recommended levels of antibiotics and biocides . The phenomenon of pathogenic bacteria acting as hyperparasites, i.e. a pathogen living inside another pathogen, requires an entirely new paradigm to fight infectious diseases. For the development of preventative and possibly therapeutic measures in such cases, we can no longer focus on a single aetiological agent, where the link between exposure and infection is clearly defined. On the contrary, the disease should be considered as having multiple and confounding factors, with different aetiologies specific to the different affected subpopulations, in order to enable us to understand fully the dynamics of disease outbreaks. To this end, the use of combinatorial approaches may hold promise in eradicating ‘superbugs’ in a clinical setting. Based on these observations, it is suggested that targeting amoebae cysts offers a potential infection control strategy. The proposition of employing antiamoebic approaches in eradicating ‘superbugs’ from the clinical setting should be considered in addition to the current measures, which are by no means futile and are proving successful in reducing the number of episodes of infections overall. Research is needed to demonstrate the efficacy of targeting the host that harbours ‘terror cells’ in our fight against infectious diseases.

Comparing antimicrobial susceptibility of meticillin-resistant Staphylococcus aureus (MRSA) to vancomycin using MicroScan® (Pos Combo 3.1J) and conventional methods

Hanae Nakashima, Mitsuaki Kameko, Hiroshi Takahashi, Hiroshi Saito — 2010-07-02

Vancomycin (VAN) minimum inhibitory concentrations (MICs) may vary depending on the method of measurement . For meticillin-resistant Staphylococcus aureus (MRSA), microbiologists from Europe and North America have reported that VAN MICs determined by Etest would reflect clinical efficacy more accurately than those determined by the VITEK® or Clinical and Laboratory Standards Institute (CLSI) reference broth microdilution (BMD) methods . However, in clinical laboratories in Japan bacterial identification and antimicrobial susceptibility testing are highly (ca. 81%) automated, with MicroScan® (Siemens Healthcare Diagnostics, Deerfield, IL) having 59% and VITEK®2 (bioMérieux, Durham, NC) having 16% of the market share . VAN MIC measurement using Etest has been conducted in research laboratories and university hospitals only.

Comparison of CLSI 2009, CLSI 2010 and EUCAST cephalosporin clinical breakpoints in recent clinical isolates of Escherichia coli, Klebsiella pneumoniae and Klebsiella oxytoca from the SMART Global Surveillance Study

2010-07-05

The Clinical and Laboratory Standards Institute (CLSI) recently published lower susceptibility breakpoints for third-generation cephalosporins. This was done in order to differentiate extended-spectrum β-lactamase (ESBL)-positive from ESBL-negative isolates, to alleviate the need for laboratories to perform ESBL screening and confirmation tests and to significantly reduce the reporting time for such testing in the clinical microbiology laboratory setting . Nevertheless, some investigators continue to report data suggesting that it is still important to know the ESBL status of an isolate to ensure proper therapy, or at least to be aware of local ESBL prevalence data so that empirical therapy can be as targeted as possible . Whilst ESBL-positive isolates are often resistant to one or more antibiotic classes, therapeutic options are more limited in areas with widespread ESBL occurrence such as Asia . The Study for Monitoring Antimicrobial Resistance Trends (SMART) is a longitudinal surveillance study that tracks susceptibility and epidemiological trends of pathogens causing intra-abdominal infections (IAIs) worldwide. All isolates of Escherichia coli, Klebsiella pneumoniae and Klebsiella oxytoca collected from 2008–2009 were tested for ESBL production using CLSI guidelines . This report compares the susceptibility rates of these isolates using the CLSI 2009 and the new CLSI 2010 interpretive criteria as well as those of the European Committee on Antimicrobial Susceptibility Testing (EUCAST) .