Important Gram-negative Bacteria Research Articles

Multiresistente Erreger

Infections caused by multidrug resistant (MDR) organisms are becoming more frequently in daily practice and are associated with an increase in duration of treatment and mortality. During the past decades, particular attention in the field of MDR pathogens was paid to methicillin-resistant staphylococcus aureus (MRSA). For the last years, MDR gram-negative organisms, with e.g., "extended-spectrum beta-lactamases" (ESBL), have been gaining a growing significance. Currently, treatment of infections with these organisms displays a greater challenge for the clinician compared to MRSA infections. This review illustrates the emergence of antibiotic resistance, provides information on the most important gram-negative and gram-positive bacteria, Clostridium difficile and measures to prevent their further spread.

Carbapenem susceptibilities and non-susceptibility concordance to different carbapenems amongst clinically important Gram-negative bacteria isolated from intensive care units in Taiwan: Results from the Surveillance of Multicentre Antimicrobial Resistance in Taiwan (SMART) in 2009

To investigate the in vitro susceptibilities to various carbapenems amongst clinical Gram-negative bacteria isolated from patients in intensive care units of ten major teaching hospitals in Taiwan in 2009, a survey was conducted to determine the minimum inhibitory concentrations (MICs) of ertapenem, imipenem, meropenem and doripenem against isolates of Enterobacteriaceae (n=594), Pseudomonas aeruginosa (n=185), Acinetobacter baumannii (n=192) and Burkholderia cepacia (n=23) using the agar dilution method. Susceptibilities were determined according to 2009, 2011 and 2012 MIC breakpoints recommended by the CLSI as well as 2012 MIC breakpoints recommended by EUCAST. Based on CLSI 2012 criteria, the ertapenem susceptible rate was 93%, 81%, 68% and 92% for Escherichia coli, Klebsiella pneumoniae, Enterobacter cloacae and Serratia marcescens, respectively. All Proteus mirabilis and Morganella morganii isolates were susceptible to ertapenem; however, 64% of P. mirabilis and all M. morganii isolates were non-susceptible to imipenem. Meropenem and doripenem had better activities than imipenem against ertapenem-non-susceptible Enterobacteriaceae isolates. E. coli, K. pneumoniae and E. cloacae with ertapenem MICs≥4mg/L were synchronously not susceptible to imipenem, meropenem and doripenem. Imipenem susceptibility was 65% and 29% for P. aeruginosa and A. baumannii, respectively. Additionally, P. aeruginosa and A. baumannii isolates with imipenem MICs≥8mg/L were also not susceptible to meropenem and doripenem. These data provide a better understanding of choosing appropriate carbapenem agents to treat infections caused by ertapenem-non-susceptible Enterobacteriaceae as well as P. aeruginosa and A. baumannii isolates with imipenem MICs≥4mg/L.

Yersinia enterocolitica and Photorhabdus asymbiotica β-lactamases BlaA are exported by the twin-arginine translocation pathway

In general, β-lactamases of medically important Gram-negative bacteria are Sec-dependently translocated into the periplasm. In contrast, β-lactamases of Mycobacteria spp. (BlaC, BlaS) and the Gram-negative environmental bacteria Stenotrophomonas maltophilia (L2) and Xanthomonas campestris (BlaXCC-1) have been reported to be secreted by the twin-arginine translocation (Tat) system. Yersinia enterocolitica carries 2 distinct β-lactamase genes (blaA and blaB) encoding BlaAYe and the AmpC-like β-lactamase BlaB, respectively. By using the software PRED-TAT for prediction and discrimination of Sec from Tat signal peptides, we identified a functional Tat signal sequence for Yersinia BlaAYe. The Tat-dependent translocation of BlaAYe could be clearly demonstrated by using a Y. enterocolitica tatC-mutant and cell fractionation. Moreover, we could demonstrate a unique unusual temperature-dependent activity profile of BlaAYe ranging from 15 to 60°C and a high ‘melting temperature’ (TM=44.3°) in comparison to the related Sec-dependent β-lactamase TEM-1 (20–50°C, TM=34.9°C). Strikingly, the blaA gene of Y. enterocolitica is present in diverse environmental Yersinia spp. and a blaA homolog gene could be identified in the closely related Photorhabdus asymbiotica (BlaAPa; 69% identity to BlaAYe). For BlaAPa of P. asymbiotica, we could also demonstrate Tat-dependent secretion. These results suggest that Yersinia BlaA-related β-lactamases may be the prototype of a large Tat-dependent β-lactamase family, which originated from environmental bacteria.

Crystal Structures of Pseudomonas aeruginosa GIM-1: Active-Site Plasticity in Metallo-β-Lactamases

Metallo-β-lactamases (MBLs) have rapidly disseminated worldwide among clinically important Gram-negative bacteria and have challenged the therapeutic use of β-lactam antibiotics, particularly carbapenems. The bla(GIM-1) gene, encoding one such enzyme, was first discovered in a Pseudomonas aeruginosa isolate from 2002 and has more recently been reported in Enterobacteriaceae. Here, we present crystal structures of GIM-1 in the apo-zinc (metal-free), mono-zinc (where Cys221 was found to be oxidized), and di-zinc forms, providing nine independently refined views of the enzyme. GIM-1 is distinguished from related MBLs in possessing a narrower active-site groove defined by aromatic side chains (Trp228 and Tyr233) at positions normally occupied by hydrophilic residues in other MBLs. Our structures reveal considerable flexibility in two loops (loop 1, residues 60 to 66; loop 2, residues 223 to 242) adjacent to the active site, with open and closed conformations defined by alternative hydrogen-bonding patterns involving Trp228. We suggest that this capacity for rearrangement permits GIM-1 to hydrolyze a wide range of β-lactams in spite of possessing a more constrained active site. Our results highlight the structural diversity within the MBL enzyme family.

Update of Peptides with Antibacterial Activity

For many years a battle has been going on between bacteria and humans, with bacteria trying to survive against the antibiotics used by humans. Bacteria are found to be dominant in this battle since they can develop resistance. In fact, in the last decade multi-, extended- and pan-drug resistant bacteria have been isolated. On the other hand, the number of new antibiotics approved by the FDA has dramatically decreased during the last 20 years. Therefore, there is a desperate need for developing new types of antibacterial agents, where antimicrobial peptides may play an important role. This review provides an update of the recently identified antimicrobial peptides. Three valid approaches for developing a future antibacterial agent, as are the mechanisms of action as well as the in vitro and in vivo assays have been described in depth. In comparison to the antibacterial agents available at present, the targets for most of the antimicrobial peptides are not well known. However several proposals having been introduced for many antimicrobial peptides of different mechanisms of action, there still lies some uncertainty about their utility. Hundreds of antimicrobial peptides have been tested in vitro against all types of bacteria, but in this review we will highlight only those which have been tested against the most important Gram-positive and Gram-negative bacteria. The last step to get a potential antibiotic includes studies with an in vivo model. Therefore only antimicrobial peptides with good activity are tested that have been described in this review.

Update of Peptides with Antibacterial Activity

For many years a battle has been going on between bacteria and humans, with bacteria trying to survive against the antibiotics used by humans. Bacteria are found to be dominant in this battle since they can develop resistance. In fact, in the last decade multi-, extended- and pan-drug resistant bacteria have been isolated. On the other hand, the number of new antibiotics approved by the FDA has dramatically decreased during the last 20 years. Therefore, there is a desperate need for developing new types of antibacterial agents, where antimicrobial peptides may play an important role. This review provides an update of the recently identified antimicrobial peptides. Three valid approaches for developing a future antibacterial agent, as are the mechanisms of action as well as the in vitro and in vivo assays have been described in depth. In comparison to the antibacterial agents available at present, the targets for most of the antimicrobial peptides are not well known. However several proposals having been introduced for many antimicrobial peptides of different mechanisms of action, there still lies some uncertainty about their utility. Hundreds of antimicrobial peptides have been tested in vitro against all types of bacteria, but in this review we will highlight only those which have been tested against the most important Grampositive and Gram-negative bacteria. The last step to get a potential antibiotic includes studies with an in vivo model. Therefore only antimicrobial peptides with good activity are tested that have been described in this review. Keywords: Antimicrobial peptides, mechanism of action, barrel model, toroidal model, aggregate model, carpet model, “in vitro” activity, “in vivo” activity, melittin, indolicidin, cecropin A, magainin II, pexiganan, buforin

Tigecycline prescribing in Salerno University Hospital: special forms and monitoring prevent inappropriate use

BackgroundAntibiotic resistance is an emerging and alarming problem in the European Union, considering the high clinical and socio-economic costs, and confirms the widespread inappropriate use of antibiotics. Tigecycline is a...

Essential Oil Composition and Antibacterial Activity of Monticalia greenmaniana (Asteraceae)

The essential oils from fresh aerial parts of Monticalia greenmaniana (Hieron) C. Jeffrey (Asteraceae) collected in March, were analyzed by GC/MS. Oil yields (w/v) of 0.1% (flowers), 0.07%, (stems) and 0.1% (leaves) were obtained by hydrodistillation. Thirteen, sixteen and eighteen components, respectively, were identified by comparison of their mass spectra with those in the Wiley GC-MS Library data base. The major components of the flower and stem oils were 1-nonane (38.8% flowers; 33.5% stems), alpha-pinene (29.0% flowers; 14.8% stems) and germacrene D (15.6% flowers; 18.6% stems). However, in the leaf oil, germacrene D was observed at 50.7%, followed by beta-cedrene at 8.4%. The leaf essential oil showed a broad spectrum of antibacterial activity against the important human pathogenic Gram-positive and Gram-negative bacteria Staphylococcus aureus (ATCC 25923), Enterococcus faecalis (ATCC 19433), Escherichia coli (ATCC 25922), Pseudomonas aeruginosa (ATCC 27853) and Klebsiella pneumoniae (ATCC 25955) with MIC values ranging from 75 to 6000 ppm.

The carbon source influences the efflux pump-mediated antimicrobial resistance in clinically important Gram-negative bacteria

Multidrug efflux pumps are proteins known to play an important role in resistance in bacteria. These proteins are located in the inner membrane (IM), together with many other proteins, including inducible permeases that participate in the uptake of non-phosphotransferase system (PTS) carbohydrates (i.e. carbohydrates uptaken by mechanisms other than the PTS). However, lipid bilayer space in the IM is limited. Therefore, we examined whether the overexpression of unrelated IM proteins is able to interfere with the efflux-mediated resistance mechanism, consequently increasing the susceptibility towards different antimicrobial compounds. We cultured bacteria under different conditions that increase the synthesis of unrelated IM proteins, either by using a non-PTS carbohydrate as the sole carbon source or by artificially overexpressing IM proteins, prior to determining the resistance to different antimicrobial compounds by disc diffusion assays. We observed that efflux-pump-mediated resistance is affected by the carbon source in all the strains tested, exhibiting increased susceptibility when a non-PTS carbohydrate was used as the sole carbon source. Moreover, when we artificially overexpressed an unrelated IM protein, we also observed decreased efflux-mediated resistance. These results strongly suggest that overexpression of IM proteins, by using a non-PTS carbohydrate as the sole carbon source, or by artificially introducing a high number of copies of an unrelated IM protein, competes with the antibiotic efflux systems, thereby decreasing the efflux-mediated resistance to different antimicrobial compounds. This sort of competition arises because of the limited available space in the bacterial IM, or by an unknown mechanism.

β-Carboline Alkaloid Constituents from a Thermoactinomyces SP. Strain Isolated from Livingston Island, Antarctica

ABSTRACTTwo natural β-carboline alkaloids were isolated from the culture filtrate of Thermoactinomyces sp. imbas-14 strain. The organism was isolated from penguin excrements collected on the Antarctic Livingston Island. Based on the effect of temperature on its growth rate, the strain is thermophilic with an optimal growth temperature at 50°C. The compounds were purified by solvent extraction, silica gel column chromatography and preparative TLC and HPLC consecutively. The structures of the two compounds were elucidated by using high-performance liquid chromatography, using double online detection with a diode array spectrophotometer and a mass spectrometer (HPLC-DAD-ESI-MS), and extensive one- and two-dimensional NMR experiments. Both substances—1-methyl-β-carboline (harmane)(1) and 1-acetyl-4-methyl-β-carboline (2), are natural products and are isolated for the first time from the culture filtrate of the antarctic strain Thermoactinomyces sp. imbas-14. These compounds showed activity against important Gram-positive (Bacillus mycoides, Bacillus subtilis) and Gram-negative bacteria (Escherichia coli), yeasts (Candida tropicalis) and filamentous fungi (Penicillium notatum).

Seasonal and Temperature-Associated Increases in Gram-Negative Bacterial Bloodstream Infections among Hospitalized Patients

BackgroundKnowledge of seasonal trends in hospital-associated infection incidence may improve surveillance and help guide the design and evaluation of infection prevention interventions. We estimated seasonal variation in the frequencies of inpatient bloodstream infections (BSIs) caused by common bacterial pathogens and examined associations of monthly BSI frequencies with ambient outdoor temperature, precipitation, and humidity levels.MethodsA database containing blood cultures from 132 U.S. hospitals collected between January 1999 and September 2006 was assembled. The database included monthly counts of inpatient blood cultures positive for several clinically important Gram-negative bacteria (Acinetobacter spp, Escherichia coli, Klebsiella pneumoniae, and Pseudomonas aeruginosa) and Gram-positive bacteria (Enterococcus spp and Staphylococcus aureus). Monthly mean temperature, total precipitation, and mean relative humidity in the postal ZIP codes of participating hospitals were obtained from national meteorological databases.ResultsA total of 211,697 inpatient BSIs were reported during 9,423 hospital-months. Adjusting for long-term trends, BSIs caused by each Gram-negative organism examined were more frequent in summer months compared with winter months, with increases ranging from 12.2% for E. coli (95% CI 9.2–15.4) to 51.8% for Acinetobacter (95% CI 41.1–63.2). Summer season was associated with 8.7% fewer Enterococcus BSIs (95% CI 11.0–5.8) and no significant change in S. aureus BSI frequency relative to winter. Independent of season, monthly humidity, monthly precipitation, and long-term trends, each 5.6°C (10°F) rise in mean monthly temperature corresponded to increases in Gram-negative bacterial BSI frequencies ranging between 3.5% for E. coli (95% CI 2.1–4.9) to 10.8% for Acinetobacter (95% CI 6.9–14.7). The same rise in mean monthly temperature corresponded to an increase of 2.2% in S. aureus BSI frequency (95% CI 1.3–3.2) but no significant change in Enterococcus BSI frequency.ConclusionsSummer season and higher mean monthly outdoor temperature are associated with substantially increased frequency of BSIs, particularly among clinically important Gram-negative bacteria.

Essential Oil Composition and Antibacteral Activity of Vismia Baccifera Fruits Collected from Mérida, Venezuela

The essential oil from the fruits of Vismia baccifera Triana & Planch. (Gutttiferae), collected in June 2009, was analyzed by GC/MS. A yield of 0.6% oil was obtained by hydrodistillation. Twenty-seven components were identified by comparison of their mass spectra with those in the Wiley GC-MS Library data base. The major components were trans-cadin-l1,4-diene (36.6%), cis-cadin-1,4-diene (18.8%) and beta-caryophyllene (11.9%). The essential oil showed a broad spectrum of antibacterial activity against the important human pathogenic Gram-positive and Gram-negative bacteria Staphylococcus aureus (ATCC 25923), Enterococcus faecalis (ATCC 29212), Escherichia coli (ATCC 25992), Pseudomonas aeruginosa (ATCC 27853) and Klebsiella pneumoniae (ATCC 23357) with MIC values ranging from 9 to 37 microg/mL.

In silico analysis of 16S rRNA gene sequencing based methods for identification of medically important aerobic Gram-negative bacteria

This study provides guidelines on the usefulness of full and 527 bp 16S rRNA gene sequencing and Microseq databases for identifying medically important aerobic Gram-negative bacteria. Overall, full and 527 bp 16S rRNA gene sequencing can identify 26.1 % and 32.6 %, respectively, of medically important aerobic Gram-negative bacteria confidently to the species level, whereas the full-MicroSeq and 500-MicroSeq databases can identify 15.2 % and 26.1 %, respectively, of medically important aerobic Gram-negative bacteria confidently to the species level. Among the major groups of aerobic Gram-negative bacteria, the methods and databases are least useful for identification of Aeromonas, Bordetella and Bartonella species. None of the Aeromonas species can be confidently or doubtfully identified, whereas only 0 % and 0-33.3 % of Bordetella species and 0-10 % and 0-10 % of Bartonella species can be confidently and doubtfully identified, respectively. On the other hand, these methods and databases are most useful for identification of members of the families Pasteurellaceae and Legionellaceae and Campylobacter species: 29.6-59.3 % and 7.4-18.5 % of members of Pasteurellaceae, 36-52 % and 12-24 % of members of Legionellaceae, and 26.7-60 % and 0-13.3 % of Campylobacter species can be confidently and doubtfully identified, respectively. Thirty-nine medically important aerobic Gram-negative bacteria that should be confidently identified by full 16S rRNA gene sequencing are not included in the full-MicroSeq database. Twenty-three medically important aerobic Gram-negative bacteria that should be confidently identified by 527 bp 16S rRNA gene sequencing are not included in the 500-MicroSeq database. Compared with results of our previous studies on anaerobic and Gram-positive bacteria, full and 527 bp 16S rRNA gene sequencing are able to confidently identify significantly more anaerobic Gram-positive and Gram-negative bacteria than aerobic Gram-positive and Gram-negative bacteria.

Bactericidal oncocin derivatives with superior serum stabilities

The proline-rich antimicrobial peptide oncocin is remarkably active in vitro against a number of important Gram-negative bacteria of concern to humans owing to their increasing resistance to antibiotics, i.e. Enterobacteriaceae (Escherichia coli, Klebsiella pneumoniae and Enterobacter cloacae) and non-fermenting species (Acinetobacter baumannii and Pseudomonas aeruginosa). Degradation of oncocin in mouse serum was investigated in this study. Several approaches to stabilise the main cleavage sites (C-terminal to Arg-15 and N-terminal to Arg-19) by substituting either or both arginine (Arg) residues with non-proteinogenic amino acids, i.e. α-amino-3-guanidino-propionic acid, homoarginine, nitro-arginine, N-methyl-arginine, β-homoarginine, d-arginine (d-Arg) or ornithine (Orn), were tested. These modifications were found to increase the half-life of oncocin in full mouse serum. For oncocin with two Orn residues in positions 15 and 19, the half-life in full serum increased from 25min to 3h. An increase of >8h was observed for oncocin with two d-Arg residues at these same positions. The antibacterial activities of these modified sequences were slightly better than the original oncocin sequence. Moreover, the three most stable analogues were found to be bactericidal against E. coli and were not toxic to HeLa cells or haemolytic to human erythrocytes.

Biosynthesis of silver nanoparticles by Streptomyces hygroscopicus and antimicrobial activity against medically important pathogenic microorganisms

Biological reduction of aqueous silver ions by extracellular components of Streptomyces hygroscopicus has facilitated the development of industrially viable greener methods for the synthesis of technologically important silver nanoparticles (AgNPs). The synthesized aqueous products showed the characteristic absorption spectra at 420 nm, which confirmed the presence of AgNPs. X-ray diffraction patterns displayed typical peaks of crystalline silver at ∼38°, ∼45° and ∼65°. The AgNPs were determined to be spherical (20–30 nm) with a purity of 70% as determined by FE-SEM, TEM, bio-AFM, XRD, and energy dispersive X-ray analysis. Furthermore, the biosynthesized AgNPs significantly inhibited the growth of medically important pathogenic gram-positive bacteria ( Bacillus subtilis and Enterococcus faecalis), gram-negative bacteria ( Escherichia coli and Salmonella typhimurium) and yeast ( Candida albicans). Thus, bioconversion of silver nanoparticles by S. hygroscopicus could be employed as a potential nanomedicine to eliminate pathogenic microorganisms.

Indirubin potentiates ciprofloxacin activity in the NorA efflux pump of Staphylococcus aureus

Staphylococcus aureus is a common multidrug-resistant organism in hospital-acquired infections, and the NorA efflux pump mechanism facilitates resistance to quinone compounds. In India, Wrightia tinctoria R. Br. leaves have traditionally been used to treat skin diseases and have been explored for antibacterial and efflux pump inhibition (EPI) compounds. In this study, indirubin isolated from the chloroform extract of W. tinctoria R. Br. leaves was tested for its antibacterial activity against clinically important Gram-positive and Gram-negative bacteria and the minimum inhibitory concentration (MIC) was determined using broth microdilution. The EPI properties of indirubin were investigated using Staphylococcus aureus SA1199B, and its synergistic effects were tested with ciprofloxacin. Indirubin showed antibacterial activity against both the type strain and drug-resistant S. aureus; the MIC was determined to be 12.5 mg/l for S. aureus and 25 mg/l for Staphylococcus epidermidis. However, it synergistically (fractional inhibitory concentration index = 0.45) potentiated the activity of ciprofloxacin, probably by inhibiting the NorA efflux pump. Indirubin exhibited EPI activity nearly comparable to that of reserpine by 4-fold reduction in ciprofloxacin MIC. Our results suggest that the natural compound indirubin could be used in future therapeutic applications as a potential EPI.

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Effects of methamidophos on the community structure, antagonism towards Rhizoctonia solani, and phlD diversity of soil Pseudomonas

A microcosm incubation study using an aquic brown soil from northeast China (a Cambisol in the UN Food and Agriculture Organization FAO Soil Taxonomy) was conducted to examine the effects of different concentrations (0, 50, 150, and 250 mg kg−1) of methamidophos (O,S-dimethyl phosphoramidothioato) on Pseudomonas, one of the most important gram-negative bacteria in soil. Amplified ribosomal DNA restriction analysis (ARDRA) was performed to study the Pseudomonas community structure, an in vitro assay was made to test the antagonistic activity of isolated Pseudomonas strains against soil-borne Rhizoctonia solani, a major member of the pathogens highly related to soil-borne plant diseases, and special primer amplification and sequencing were performed to investigate the diversity of phlD, an essential gene in the biosynthesis of 2, 4-diacetylphloroglucinol (2, 4-DAPG), which has biocontrol activity in phlD +isolates. With exposure to increasing methamidophos concentrations, the total number of soil Pseudomonas ARDRA patterns decreased significantly, but with less change in the same treatments over 1, 3, and 5 weeks of incubation. The number of isolated Pseudomonas strains with antagonistic activity against R. solani as well as the diversity and appearance frequency of the strains' phlD gene also decreased with increasing concentrations of methamidophos, especially at high methamidophos concentrations. Applying methamidophos could increase the risk of soil-borne plant diseases by decreasing the diversity of the soil Pseudomonas community and the amount of R. solani antagonists, particularly those with the phlD gene.

High Potency and Broad-Spectrum Antimicrobial Peptides Synthesized via Ring-Opening Polymerization of α-Aminoacid-N-carboxyanhydrides

Antimicrobial peptides (AMPs), particularly those effective against methicillin-resistant Staphylococcus aureus ( S. aureus ) and antibiotic-resistant Pseudomonas aeruginosa ( P. aeruginosa ), are important alternatives to antibiotics. Typical peptide synthesis methods involving solid-phase sequential synthesis are slow and costly, which are obstacles to their more widespread application. In this paper, we synthesize peptides via ring-opening polymerization of alpha-amino acid N-carboxyanhydrides (NCA) using a transition metal initiator. This method offers high potential for inexpensive synthesis of substantial quantities of AMPs. Lysine (K) was chosen as the hydrophilic amino acid and alanine (A), phenylalanine (F), and leucine (L) as the hydrophobic amino acids. We synthesized five series of AMPs (i.e., P(KA), P(KL), P(KF), P(KAL), and P(KFL)), varied the hydrophobic amino acid content from 0 to 100%, and determined minimal inhibitory concentrations (MICs) against clinically important Gram-negative and Gram-positive bacteria and fungi (i.e., Escherichia coli ( E. coli ), P. aeruginosa , Serratia marcescens ( S. marcescens ), and Candida albicans ( C. albicans ). We found that P(K(10)F(7.5)L(7.5)) and P(K(10)F(15)) show the broadest activity against all five pathogens and have the lowest MICs against these pathogens. For P(K(10)F(7.5)L(7.5)), the MICs against E. coli , P. aeruginosa , S. marcescens , S. aureus , and C. albicans are 31 microg/mL, 31 microg/mL, 250 microg/mL, 31 microg/mL, and 62.5 microg/mL, while for P(K(10)F(15)) the respective MICs are 31 microg/mL, 31 microg/mL, 250 microg/mL, 31 microg/mL, and 125 microg/mL. These are lower than the MICs of many naturally occurring AMPs. The membrane depolarization and SEM assays confirm that the mechanism of microbe killing by P(K(10)F(7.5)L(7.5)) copeptide includes membrane disruption, which is likely to inhibit rapid induction of AMP-resistance in pathogens.

In vitro activity of ceftobiprole against clinical isolates of Pseudomonas aeruginosa obtained from Canadian intensive care unit (ICU) patients as part of the CAN-ICU Study

Sir, In recent years, Pseudomonas aeruginosa isolates resistant to multiple classes of antimicrobial agents have become increasingly common. Several mechanisms may contribute to antimicrobial resistance among P. aeruginosa, including the production of a chromosomally encoded AmpC b-lactamase. Ceftobiprole (BAL9141), an investigational pyrrolidinone cephalosporin, is reported to have activity against a broad spectrum of clinically important Gram-negative bacteria including P. aeruginosa. Additionally, in vitro studies have demonstrated that ceftobiprole is hydrolysed very slowly by AmpC cephalosporinases. The purpose of this report was to describe the in vitro activity of ceftobiprole versus 419 clinical isolates of P. aeruginosa obtained from patients in an intensive care unit (ICU) setting. Cefepime was used as a comparator antimicrobial agent. From September 2005 to June 2006, inclusive, P. aeruginosa isolates were collected as part of the Canadian National Intensive Care Unit (CAN-ICU) Study. The CAN-ICU Study included 19 medical centres from all regions of Canada with active ICUs. Each centre submitted a maximum of 300 consecutive pathogens isolated from blood, urine, tissue/wound and respiratory specimens (one pathogen per cultured site per patient) of ICU patients. Centres were requested to only obtain ‘clinically important’ specimens from patients with a presumed infectious disease. Surveillance swabs, eye, ear, nose and throat swabs were excluded, as were anaerobic bacteria and fungi. Isolates were shipped to the reference laboratory (Health Sciences Centre, Winnipeg, MB, Canada) on Amies charcoal swabs, subcultured onto appropriate media and stocked in skimmed milk at 2808C until MIC testing was performed. The in vitro activities of ceftobiprole and cefepime were determined by microbroth dilution in accordance with the CLSI guidelines. MIC interpretive standards for cefepime were defined according to CLSI breakpoints. At present, susceptibility breakpoints for ceftobiprole do not exist. In total, 419 isolates of P. aeruginosa were collected as part of the CAN-ICU Study. The breakdown of these 419 isolates by specimen source was as follows: respiratory (69.0%), wound (12.4%), urine (10.7%) and blood (7.9%). The MIC distributions of ceftobiprole and cefepime for the isolates were very similar (Table 1). At an antimicrobial concentration of 8 mg/L, 74.7% and 78.7% of our isolates would be inhibited by ceftobiprole and cefepime, respectively. The MIC50 and MIC90 values of ceftobiprole were 4 and 16 mg/L. The corresponding MIC50 and MIC90 values of cefepime were 4 and 32 mg/L. The MIC50/ MIC90 values of ceftobiprole and cefepime were comparable, regardless of specimen source. Cross-resistance between ceftobiprole and cefepime was observed. Of 43 cefepime-resistant isolates (MIC 32 mg/L), only 3 (7.0%) had an MIC of ceftobiprole of 8 mg/L. Similarly, only 6 of 40 isolates (15%) with a ceftobiprole MIC of 32 mg/L were susceptible to cefepime. In agreement with our results, previous studies have reported similar in vitro activity between ceftobiprole and cefepime when evaluated against P. aeruginosa. A recent study by Pillar et al. documented a lower modal MIC of cefepime in comparison with ceftobiprole against P. aeruginosa isolates (1 mg/L versus 2 mg/L). However, the MIC50 and MIC90 values were identical (4 and 16 mg/L respectively) for both antimicrobials. Our data demonstrate a high degree of cross-resistance between ceftobiprole and cefepime. Hebeisen et al. have previously described the finding of cross-resistance between ceftazidime, cefepime and ceftobiprole. These investigators reported the activity of ceftobiprole against 17 ceftazidime-non-susceptible P. aeruginosa isolates (MIC50 of 16 mg/L and MIC90 of .64 mg/L for ceftazidime). The MIC50 and MIC90 values of ceftobiprole against these isolates were 16 and .64 mg/L, respectively. The corresponding MIC50 and MIC90 values of cefepime versus the ceftazidime-non-susceptible isolates were 32 and 32 mg/L. Cross-resistance has also been described in a study by Pillar et al., where the MIC50/MIC90 values of ceftobiprole against 491 ceftazidime-susceptible and 130 ceftazidime-non-susceptible P. aeruginosa isolates were 2/8 and 16/.32 mg/L, respectively. In summary, ceftobiprole and cefepime demonstrated comparable in vitro activity against P. aeruginosa clinical isolates from Canadian ICUs and cross-resistance between these agents was common. Whether ceftobiprole will be clinically useful in the treatment of serious infections caused by P. aeruginosa remains to be determined.