Polymyxin Derivatives that Sensitize Gram-Negative Bacteria to Other Antibiotics.

Polymyxins (polymyxin B (PMB) and polymyxin E (colistin)) are cyclic lipodecapeptide antibiotics, highly basic due to five free amino groups, and rapidly bactericidal against Gram-negative bacteria, such as the majority of Enterobacteriaceae as well as Acinetobacter baumannii and Pseudomonas aeruginosa. Their clinical use was abandoned in the 1960s because of nephrotoxicity and because better-tolerated drugs belonging to other antibiotic classes were introduced. Now, due to the global dissemination of extremely-drug resistant Gram-negative bacterial strains, polymyxins have resurged as the last-line drugs against those strains. Novel derivatives that are less toxic and/or more effective at tolerable doses are currently under preclinical development and their properties have recently been described in several extensive reviews. Other derivatives lack any direct bactericidal activity but damage the outermost permeability barrier, the outer membrane, of the target bacteria and make it more permeable to many other antibiotics. This review describes the properties of three thus far best-characterized "permeabilizer" derivatives, i.e., the classic permeabilizer polymyxin B nonapeptide (PMBN), NAB7061, and SPR741/NAB741, a compound that recently successfully passed the clinical phase 1. Also, a few other permeabilizer compounds are brought up.

New polymyxin derivatives that display improved efficacy in animal infection models as compared to polymyxin B and colistin.

Polymyxin B and colistin (polymyxin E) are bactericidal pentacationic lipopeptides that act specifically on Gram-negative bacteria, first by disrupting their outermost permeability barrier, the outer membrane (OM), and then damaging the cytoplasmic membrane. The discovery of both polymyxin B and colistin was published independently by three laboratories as early as in 1947. They were subsequently used in intravenous therapy. Unfortunately, they also exhibit significant and dose-limiting nephrotoxicity. Therefore, polymyxins were reserved as agents of last-line defense. The emergence of extremely multiresistant strains has now forced clinicians to reinstate polymyxins in the therapy of severe infections. However, the current dosage regimens lead to insufficient drug concentrations in serum and clinicians have been advised to use larger doses, which further increases the risk of nephrotoxicity. Very recently, the interest in developing better tolerated and more effective polymyxins has grown. This review focuses on describing four development programs that have yielded novel derivatives that are more effective than the old polymyxins in animal infection models. Compounds from three programs are superior to the old polymyxins in the rodent lung infection model with Acinetobacter baumannii and/or Pseudomonas aeruginosa. One of them is also more effective than polymyxin B in A. baumannii mouse thigh infection. The fourth program includes compounds that are approximately tenfold more effective in Escherichia coli murine pyelonephritis than polymyxin B.

Polymyxin derivatives NAB739 and NAB815 are more effective than polymyxin B in murine Escherichia coli pyelonephritis.

Objectives: Extremely multiresistant strains of Enterobacteriaceae, such as those of Escherichia coli and Klebsiella pneumoniae, are emerging and spreading at a worrisome speed. Polymyxins (polymyxin B, colistin) are used as last-line therapy against such strains, in spite of their notable nephrotoxicity that may even require discontinuation of the therapy. We have previously developed polymyxin derivatives NAB739 and NAB815 that are better tolerated in cynomolgus monkeys than polymyxin B and are, in contrast to polymyxin B, excreted in the cynomolgus urine to a very significant degree. Here we have compared the efficacy of these NAB compounds and polymyxin B in the therapy of murine pyelonephritis caused by E. coli. Methods: The challenge organism was a uropathogenic E. coli clinical isolate. Mice were inoculated via urethral catheterization with 5 × 108 cfu. All treatment groups consisted of 12 animals. On day 1 and day 2 post-infection, the mice were treated subcutaneously with NAB739, NAB815, polymyxin B or vehicle twice a day and on day 3 post-infection the animals were sacrificed. cfu in the kidney and bladder tissues and in the urine were determined. Results: NAB739 reduced the bacterial burden in the kidney, urine and bladder at doses approximately 10-fold lower than those of polymyxin B. In the kidneys, the half-maximal effective dose (ED50) was 9-fold lower for NAB739 than for polymyxin B (0.24 mg/kg versus 2.1 mg/kg, respectively). NAB815 was as effective as NAB739. Conclusions: NAB739 and NAB815 were unequivocally more effective than polymyxin B in the murine pyelonephritis model.

Potentiation of Antibiotic Activity by a Novel Cationic Peptide: Potency and Spectrum of Activity of SPR741.

Novel approaches for the treatment of multidrug-resistant Gram-negative bacterial infections are urgently required. One approach is to potentiate the efficacy of existing antibiotics whose spectrum of activity is limited by the permeability barrier presented by the Gram-negative outer membrane. Cationic peptides derived from polymyxin B have been used to permeabilize the outer membrane, granting antibiotics that would otherwise be excluded access to their targets. We assessed the in vitro efficacies of combinations of SPR741 with conventional antibiotics against Escherichia coli, Klebsiella pneumoniae, and Acinetobacter baumannii Of 35 antibiotics tested, the MICs of 8 of them were reduced 32- to 8,000-fold against E. coli and K. pneumoniae in the presence of SPR741. The eight antibiotics, azithromycin, clarithromycin, erythromycin, fusidic acid, mupirocin, retapamulin, rifampin, and telithromycin, had diverse targets and mechanisms of action. Against A. baumannii, similar potentiation was achieved with clarithromycin, erythromycin, fusidic acid, retapamulin, and rifampin. Susceptibility testing of the most effective antibiotic-SPR741 combinations was extended to 25 additional multidrug-resistant or clinical isolates of E. coli and K. pneumoniae and 17 additional A. baumannii isolates in order to rank the potentiated antibiotics. SPR741 was also able to potentiate antibiotics that are substrates of the AcrAB-TolC efflux pump in E. coli, effectively circumventing the contribution of this pump to intrinsic antibiotic resistance. These studies support the further development of SPR741 in combination with conventional antibiotics for the treatment of Gram-negative bacterial infections.

An outbreak of CTX-M-15 -producing Escherichia coli, Enterobacter cloacae, and Klebsiella in a children's hospital in Finland.

Four different extended-spectrum ß -lactamase (ESBL)-producing bacteria from a pediatric surgery ward were studied. The presence of TEM-, SHV-, and CTX-M-type ß -lactamases was analyzed and the relatedness of the isolates studied with a repetitive PCR system (DiversiLab) and pulsed-fi eld gel electrophoresis (PFGE). Molecular analysis showed that a clonal dissemination of CTX-M-15-producing Escherichia coli and Enterobacter cloacae had occurred.

Efficacy of a novel PCR- and microarray-based method in diagnosis of a prosthetic joint infection.

BACKGROUND AND PURPOSE: Polymerase chain reaction (PCR) methods enable detection and species identification of many pathogens. We assessed the efficacy of a new PCR and microarray-based platform for detection of bacteria in prosthetic joint infections (PJIs). METHODS: This prospective study involved 61 suspected PJIs in hip and knee prostheses and 20 negative controls. 142 samples were analyzed by Prove-it Bone and Joint assay. The laboratory staff conducting the Prove-it analysis were not aware of the results of microbiological culture and clinical findings. The results of the analysis were compared with diagnosis of PJIs defined according to the Musculoskeletal Infection Society (MSIS) criteria and with the results of microbiological culture. RESULTS: 38 of 61 suspected PJIs met the definition of PJI according to the MSIS criteria. Of the 38 patients, the PCR detected bacteria in 31 whereas bacterial culture was positive in 28 patients. 15 of the PJI patients were undergoing antimicrobial treatment as the samples for analysis were obtained. When antimicrobial treatment had lasted 4 days or more, PCR detected bacteria in 6 of the 9 patients, but positive cultures were noted in only 2 of the 9 patients. All PCR results for the controls were negative. Of the 61 suspected PJIs, there were false-positive PCR results in 6 cases. INTERPRETATION: The Prove-it assay was helpful in PJI diagnostics during ongoing antimicrobial treatment. Without preceding treatment with antimicrobials, PCR and microarray-based assay did not appear to give any additional information over culture.

A quantitative polymerase chain reaction assay for rapid detection of 9 pathogens directly from stools of travelers with diarrhea.

BACKGROUND & AIMS: Every year, 80 million tourists traveling to tropical and subtropical areas contract traveler's diarrhea (TD). Forty percent to 80% of cases are caused by bacteria, yet clinical diagnostic tests are available to identify only a few of the strains that cause TD. We aimed to develop a quantitative polymerase chain reaction (qPCR) assay to identify all major pathogens in stool samples. METHODS: We developed a low-cost, high-throughput, multiplex qPCR assay for simultaneous detection of 9 bacterial pathogens in stool samples: Salmonella, Yersinia, Campylobacter, and Vibrio cholerae, as well as Shigella or enteroinvasive Escherichia coli, enterohemorrhagic E coli, enterotoxigenic E coli (ETEC), enteroaggregative E coli (EAEC), and enteropathogenic E coli (EPEC). The assay was validated using positive (n = 245) and negative (n = 243) control strains, as well as preselected positive and negative stool samples. In addition, stool samples were collected from 96 returning travelers with TD. The findings were compared with those from routine diagnostic tests. RESULTS: The assay detected the bacterial strains with 100% sensitivity and specificity, compared with results from the reference tests. Of all stool samples collected from travelers with TD, EPEC was found in 47%, EAEC in 46%, ETEC in 22%, enterohemorrhagic E coli in 7%, Campylobacter in 6%, Shigella or enteroinvasive E coli in 2%, and Salmonella in 2%. Multiple pathogens were found in 37% of all samples. CONCLUSIONS: We developed a low-cost, high-throughput qPCR assay for use in routine diagnostic analysis and research. It detects the pathogenic bacteria most commonly associated with TD in stool samples with 100% sensitivity and specificity, compared with reference methods. The assay requires 4 hours, whereas current detection methods require 1 to 7 days. At least 1 TD pathogen was identified in stool samples from 76% of returning travelers, whereas conventional methods found a pathogen in only 17%. The most commonly detected bacteria were EPEC, EAEC, and ETEC.

Novel derivatives of polymyxins.

Polymyxin B and colistin (polymyxin E) are bactericidal pentacationic lipopeptides that act specifically on Gram-negative bacteria, first by disrupting their outermost permeability barrier, the outer membrane (OM), and then damaging the cytoplasmic membrane. Both were discovered in the mid-1950s and subsequently used in intravenous therapy, but soon largely abandoned because of nephrotoxicity. The emergence of extremely multiresistant strains has now forced clinicians to reinstate them in the therapy of severe infections caused by such strains. This article reviews recent attempts to develop novel derivatives of polymyxins that exhibit less toxicity and greater potency than the existing drugs. In addition, studies of novel des-fatty acyl-polymyxin derivatives that display activity against Pseudomonas aeruginosa are included. The review also covers recent studies of derivatives that lack potent bactericidal action, but which disrupt the OM, which increases bacterial permeability to other antibiotics, facilitating their entry into the cell.

Antimicrobial activity of the novel polymyxin derivative NAB739 tested against Gram-negative pathogens.

OBJECTIVES: In spite of reported nephrotoxicity, polymyxins have been reinstated as the last-line therapy to treat infections caused by Gram-negative bacterial strains that are resistant to other agents. NAB739 has a cyclic portion identical to that of polymyxin B, but its linear peptide portion consists of threonyl-d-serinyl instead of diaminobutyryl-threonyl-diaminobutyryl. Therefore, NAB739 lacks both of the positive charges present in the linear part of polymyxin B. Here, we compare the antibacterial activity of NAB739 with that of polymyxin B against a representative collection of contemporary Gram-negative bacteria. METHODS: NAB739 and polymyxin B MIC values were determined for 310 clinical isolates by the reference broth microdilution method according to CLSI document M07-A9 (2012). RESULTS: MIC(90)s of NAB739 for the subset consisting of polymyxin-susceptible (MIC, ≤ 2 mg/L) clinical isolates of Escherichia coli (n=51), Klebsiella pneumoniae (n=50), Acinetobacter spp. (n=49) and Pseudomonas aeruginosa (n=49) were 2, 2, 8 and 16 mg/L, respectively. For polymyxin-non-susceptible strains of E. coli (n=12), K. pneumoniae (n=11), Acinetobacter spp. (n=11) and P. aeruginosa (n=14) the NAB739 MIC(90) was ≥ 64 mg/L. CONCLUSIONS: The MIC(90) of NAB739 for polymyxin-susceptible strains of E. coli and K. pneumoniae was identical to and 2-fold higher than that of polymyxin B, respectively. For polymyxin-susceptible strains of Acinetobacter spp. and P. aeruginosa, the MIC(90) of NAB739 was 4-fold and 8-fold higher than that of polymyxin B, respectively. For polymyxin-non-susceptible strains of all these species, the MIC(90) values of NAB739 were high and 2- to 4-fold higher than those of polymyxin B.

Applicability of DiversiLab repetitive sequence-based PCR method in epidemiological typing of enterohemorrhagic Escherichia coli (EHEC).

Enterohemorrhagic Escherichia coli (EHEC) causes diarrhea, often with severe complications. Rapid and discriminatory typing of EHEC using advanced molecular methods is needed for determination of the genetic relatedness of clones responsible for foodborne outbreaks and for finding out the transmission sources of the outbreaks. This study evaluated the potential of DiversiLab, a semiautomated repetitive sequence-based polymerase chain reaction method for the genotyping of EHEC strains. A set of 52 EHEC strains belonging to 15 O:H serotypes was clustered into 10 DiversiLab groups. All of the O157 strains and one O55 strain were classified into the same group based on a 90% similarity threshold. The other serotypes were classified to their own DiversiLab group, with the exception of one R:H(-) strain that was grouped with O5:H(-) strains. In addition, O26 and O111 strains were grouped together but ultimately subdivided according to their O-serotypes based on a 95% similarity threshold. The O104 strain, which was associated with a major outbreak of hemolytic uremic syndrome in Germany in May 2011, was also classified independently. The DiversiLab performed well in identifying isolates, but the discriminatory power of the repetitive sequence-based polymerase chain reaction method was lower than that of pulsed-field gel electrophoresis. Analysis of 15 enteropathogenic E. coli (EPEC) strains revealed that some EPEC strains clustered together with EHEC strains. Therefore, the DiversiLab system cannot be used to discriminate between these pathogroups. In conclusion, DiversiLab is a rapid and easy system for the primary exclusion of unrelated EHEC strains based on their serotypes, but more discriminatory methods, such as pulsed-field gel electrophoresis, are needed for accurate typing of the EHEC strains.

Accurate and rapid identification of Candida spp. frequently associated with fungemia by using PCR and the microarray-based Prove-it Sepsis assay.

The rapid identification of microbes responsible for bloodstream infections (BSIs) allows more focused and effective therapies and outcomes. DNA sequence-based methods offer an opportunity for faster, accurate diagnosis and for effective therapy. As our objective of the study, the ability of the Prove-it Sepsis platform, already proven as a rapid PCR- and microarray-based assay for the majority of sepsis-causing bacteria, was extended to also rapidly identify clinically relevant yeasts in blood culture. The performance characteristics of this extended platform are described. We found that the extended diagnostic Prove-it Sepsis platform was found to be highly accurate when analyzing primary isolates, spiked blood cultures, nucleic acid extracts from a retrospective blood culture data set, and primary blood cultures. Comparison of the blood culture results from the Prove-it Sepsis platform with those from conventional culture-based methods or by gene sequencing demonstrated a sensitivity of 99% and a specificity of 98% for fungal targets (based on analysis of a total of 388 specimens). Total assay time was 3 h from DNA extraction to BSI diagnosis. These results extend the performance characteristics of the Prove-it platform for bacteria to the easy, rapid, and accurate detection and species identification of yeasts in positive blood cultures. Incorporation of this extended and rapid diagnostic platform into the tools for clinical patient management would allow possibly faster identification and more focused therapies for BSIs.

Carbapenemase-producing Enterobacteriaceae in Finland: the first years (2008-11).

OBJECTIVES: Carbapenemase-producing Enterobacteriaceae (CPE) are becoming a global problem; they are often resistant to nearly all available antibiotics. Here we report details on all Finnish CPE isolates found until the end of 2011: carbapenemase genes, travel history and multilocus sequence typing (MLST) data. METHODS: Enterobacteriaceae sent to the Antimicrobial Resistance Unit of the National Institute for Health and Welfare were tested for susceptibility to carbapenems, screened for carbapenemases by PCR and isolates with decreased susceptibility to carbapenems were tested for hydrolysis of imipenem. Carbapenemase-producing Escherichia coli and Klebsiella pneumoniae isolates were typed by MLST. RESULTS: In all, 26 CPE strains were found from 25 patients: 10 with OXA-48-like enzymes, 5 with KPC, 4 with VIM, 3 with NDM, 3 with IMI/NMC-A and 1 with GES-14. The species were K. pneumoniae (n = 16), E. coli (n = 6), Enterobacter cloacae (n = 3) and Raoultella planticola (n = 1). Of the 25 patients, 18 had a known travel history/hospital transfer from abroad. Local spread/transmission was suspected in 2011, but there were no hospital outbreaks. The K. pneumoniae multilocus sequence types ST258, ST182, ST147, ST244, ST14, ST13, ST383, ST101 and ST15, and the E. coli sequence types ST38 and ST90 were found. Many of these are global epidemic clones. CONCLUSIONS: CPE strains are increasingly found in Finland, but still at a very low prevalence.

Accurate and rapid identification of bacterial species from positive blood cultures with a DNA-based microarray platform: an observational study.

BACKGROUND: New DNA-based microarray platforms enable rapid detection and species identification of many pathogens, including bacteria. We assessed the sensitivity, specificity, and turnaround time of a new molecular sepsis assay. METHODS: 2107 positive blood-culture samples of 3318 blood samples from patients with clinically suspected sepsis were investigated for bacterial species by both conventional culture and Prove-it sepsis assay (Mobidiag, Helsinki, Finland) in two centres (UK and Finland). The assay is a novel PCR and microarray method that is based on amplification and detection of gyrB, parE, and mecA genes of 50 bacterial species. Operators of the test assay were not aware of culture results. We calculated sensitivity, specificity, and turnaround time according to Clinical and Laboratory Standards Institute recommendations. FINDINGS: 1807 of 2107 (86%) positive blood-culture samples included a pathogen covered by the assay. The assay had a clinical sensitivity of 94.7% (95% CI 93.6-95.7) and a specificity of 98.8% (98.1-99.2), and 100% for both measures for meticillin-resistant Staphylococcus aureus bacteraemia. The assay was a mean 18 h faster than was the conventional culture-based method, which takes an additional 1-2 working days. 34 of 3284 (1.0%) samples were excluded because of technical and operator errors. INTERPRETATION: Definitive identification of bacterial species with this microarray platform was highly sensitive, specific, and faster than was the gold-standard culture-based method. This assay could enable fast and earlier evidence-based management for clinical sepsis.

Comparative activity of tigecycline and tetracycline on Gram-negative and Gram-positive bacteria revealed by a multicentre study in four North European countries.

BACKGROUND: This study involves a multicentre surveillance of tigecycline and tetracycline activity against Gram-negative and Gram-positive bacteria from primary care centres (PCCs), general hospital wards (GHWs) and intensive care units (ICUs) in Denmark (n = 9), Finland (n = 10), Norway (n = 7) and Sweden (n = 19). METHODS: The hospitals were each asked to test 30 consecutive Gram-positive and 30 Gram-negative clinical isolates. Supportive information accompanying each isolate included the study centre, ward level (PCC, GHW, or ICU), patient identification and source of the isolate. Minimum inhibitory concentrations (MICs) for tetracycline and tigecycline were determined with the Etest. RESULTS: The isolates collected comprised 1610 Gram-negative and 1767 Gram-positive clinical isolates. The study showed low rates of non-susceptibility (intermediate (I) and resistant (R)) to tigecycline: <1% in Escherichia coli, though other Enterobacteriaceae showed higher rates (Enterobacter cloacae (7%), Klebsiella pneumoniae (9%) and Serratia spp. (23%)). The overall non-susceptibility rate for tigecycline in Enterobacteriaceae with species-related breakpoints for tigecycline was 6% (4% excluding Serratia spp.). The activity of tigecycline against Haemophilus influenzae and Acinetobacter spp. was high with a MIC(50) of 0.25 mg/l and MIC(90) of 1 mg/l. The prevalence of non-susceptibility to tigecycline among Gram-positive bacteria was <1%. The corresponding figure for tetracycline was 14%. The activity of tigecycline against Streptococcus pneumoniae was high with MIC(50) and MIC(90) of 0.125 mg/l. CONCLUSION: Tigecycline showed good overall in vitro activity against Gram-positive and Gram-negative isolates, including both tetracycline-susceptible and resistant isolates. Most non-susceptibility to tigecycline among Enterobacteriaceae other than E. coli was I (6%), rather than R (<1%). This indicates a problem setting interpretive species-related tigecycline breakpoints for Enterobacteriaceae other than E. coli.

Shiga toxin-producing Escherichia coli O100:H⁻: stx2e in drinking water contaminated by waste water in Finland.

In November 2007, 450 m(3) of treated wastewater leaked into the drinking water distribution system contaminating the drinking water of over 10,000 inhabitants of Nokia, Southern Finland. Nearly 1,000 people visited the health centre because of gastroenteritis during the following 5 weeks. A wide range of enteric pathogens was found in the patients. The authors used the 16-plex PCR to investigate whether the five major diarrheagenic Escherichia coli pathotypes (EPEC, ETEC, STEC, EIEC or EAEC) were present in the contaminated drinking water and in the patients' stool samples. The contaminated drinking water was positive for genes characteristic of various E. coli pathotypes: pic, invE, hlyA, ent, escV, eae, aggR, stx(2) , estIa and astA. These genes, except stx(2), hlyA and invE, were also detected in the stool samples of the patients linked to this outbreak. A sorbitol positive, streptomycin resistant STEC strain was isolated from the drinking water, and belonged to the serotype O100:H(-), produced Stx2 toxin (titre 1:8 by reversed-passive latex agglutination method), and carried the genes stx(2e), estIa and irp2.

Isolation and characterization of Streptococcus mitis from blood of child with osteomyelitis.

OBJECTIVES: Osteomyelitis is an inflammatory process accompanied by bone destruction that is caused by bacterial infection, with most child cases showing a haematogenous origin and metaphysis of the long bones. The aim of the present study was to characterize streptococcal strains isolated from the blood of a child diagnosed with osteomyelitis in a long bone and investigate the biological properties related to virulence of strains associated with osteomyelitis. METHODS: Blood isolate species were determined based on the 16S rRNA sequence. Next, the blood isolates were analysed for phagocytosis susceptibility by polymorphonuclear leukocytes, platelet aggregation, inhibitory effects on osteoblastic cells, and their properties of adhesion with cells, and compared to the reference strain Streptococcus mitis ATCC49456. RESULTS: The blood isolates were found to be a single clone (named SA1101), which was determined to be S. mitis. The phagocytosis susceptibility of SA1101 was significantly lower than that of ATCC49456, while its platelet aggregation rate was higher. Furthermore, SA1101 showed an inhibitory effect toward the growth of osteoblastic cells and had greater properties of adhesion to those cells as compared to ATCC49456. CONCLUSIONS: These results suggest that S. mitis SA1101 is a possible etiological agent and caused osteomyelitis in the present case.

A novel polymyxin derivative that lacks the fatty acid tail and carries only three positive charges has strong synergism with agents excluded by the intact outer membrane.

Polymyxins are cationic lipopeptides (five cationic charges) and the last resort for the treatment of serious Gram-negative infections caused by multiresistant strains. NAB741 has a cyclic peptide portion identical to that of polymyxin B but carries in the linear peptide portion a threonyl-D-serinyl residue (no cationic charges) instead of the diaminobutyryl-threonyl-diaminobutyryl residue (two cationic charges). At the N terminus of the peptide, NAB741 carries an acetyl group instead of a mixture of methyl octanoyl and methyl heptanoyl residues. NAB741 sensitized Escherichia coli, Klebsiella pneumoniae, Enterobacter cloacae, and Acinetobacter baumannii to antibiotics against which the intact outer membrane is an effective permeability barrier. When tested by using Etest strips on plates containing increasing concentrations of NAB741, the fractional inhibition concentration index (FICI) of the combination of NAB741 with rifampin ranged from

Novel polymyxin derivatives are effective in treating experimental Escherichia coli peritoneal infection in mice.

OBJECTIVES: Novel synthetic polymyxin derivatives including NAB737 and NAB739 are as effective as polymyxin B in vitro against the common opportunistic pathogen Escherichia coli. Another derivative, NAB7061, lacks direct antibacterial action but sensitizes E. coli to several other antibacterial agents including macrolides. The renal handling of NAB739 and NAB7061 in rats differs from that of polymyxin B. Furthermore, the affinities of NAB739 and NAB7061 for isolated rat kidney brush border membrane are significantly lower than that of polymyxin B. Here we investigate the in vivo antibacterial effect of these compounds. METHODS: The polymyxin derivatives were evaluated in an experimental murine peritonitis model. Immunocompetent mice were infected intraperitoneally with E. coli IH3080 and were subcutaneously treated with NAB737, NAB739 or NAB7061. RESULTS: A >4.0 log(10) reduction in bacterial load compared with saline control was achieved 6 h after initiation of treatment with 1 mg/kg of NAB739 twice or 4 mg/kg of NAB737 twice. Combination therapy with NAB7061 (5 mg/kg) twice and erythromycin (10 mg/kg) resulted during the same time course in a >2.0 log(10) reduction in bacterial load compared with saline control. Neither NAB7061 nor erythromycin was effective as monotherapy. Together with the ability to reduce bacterial load, the NAB compounds also improved the clinical status of the mice. CONCLUSIONS: We found that the three novel synthetic polymyxin B derivatives had a potent in vivo bactericidal effect against E. coli.

Susceptibility of carbapenemase-producing strains of Klebsiella pneumoniae and Escherichia coli to the direct antibacterial activity of NAB739 and to the synergistic activity of NAB7061 with rifampicin and clarithromycin.

OBJECTIVES: To determine the susceptibility of carbapenemase-producing strains of Klebsiella pneumoniae and Escherichia coli to the direct antibacterial activity of NAB739 and to the synergistic activity of NAB7061 with rifampicin and clarithromycin. NAB739 and NAB7061 are novel polymyxin derivatives that lack the cationic charges in the linear peptide portion of polymyxin B and have pharmacokinetic properties different from those of polymyxin B. METHODS: MIC determinations were performed by the agar dilution method using CLSI guidelines. Polymyxin B was used as a comparison. Synergism studies measured fractional inhibitory concentration indices (FICIs) by using increasing concentrations of the compounds in Mueller-Hinton agar and Etests. RESULTS: The MICs of NAB739 for all nine polymyxin-susceptible, carbapenemase-producing strains were identical or very close to those determined for E. coli ATCC 25922, for K. pneumoniae ATCC 13883, as well as for 18 clinical carbapenem-susceptible isolates. At a concentration of 4 mg/L, NAB7061 decreased the MIC of rifampicin and clarithromycin for all carbapenemase strains by factors ranging from 6 to 500. The polymyxin-resistant strain K. pneumoniae CL5762B was sensitized by a factor of 24 to rifampicin (FICI, 0.167) and by a factor of 12 to clarithromycin (FICI, 0.208). CONCLUSIONS: Polymyxin-susceptible, carbapenemase-producing strains are as susceptible to NAB739 as are the carbapenem-susceptible clinical isolates. In addition, NAB7061 has notable synergism with rifampicin and clarithromycin against all the carbapenemase-producing strains tested, including the polymyxin-resistant K. pneumoniae strain.

Pneumococcal serotype distribution in invasive paediatric disease in Southern Finland before the introduction of vaccine.

Vaccination with pneumococcal conjugate vaccines (PCV) has been started in many countries, resulting in a marked decrease in the incidence of invasive pneumococcal disease (IPD). After the decline, an increase in the incidence of IPD caused by non-vaccine serotypes has been observed. Finland has just decided to incorporate PCV into the national immunization programme, starting 1 September 2010. We describe the patients treated for IPD in one hospital in Finland before the use of PCV. Details of patients treated for IPD at the Children's Hospital, University of Helsinki between 2002 and 2007, were sought using the hospital discharge register and laboratory data. In total 118 cases of IPD were diagnosed in 113 children between 1 January 2002 and 31 December 2007. The yearly number of cases of IPD increased from 10 in 2002 to 26 in 2007. There were 5 deaths, all in patients with meningitis (5/19, 26%), and the total mortality was 4% (5/113). Seventy-five (77%) of the 97 pneumococcal isolates for which serotype was available were covered by the 7-valent PCV; 77% were also in the 10-valent PCV and 91% in the 13-valent PCV. Considering the serotype distribution of the pneumococcal isolates, a marked benefit is expected from including PCV in the vaccination programme.