Factors associated with bacteremia due to multidrug-resistant Gram-negative bacilli in hematopoietic stem cell transplant recipients

The epidemiology of bacteremia developing during neutropenia has changed in the past decade, with the re-emergence of Gram-negative (GN) bacteria and the development of multidrug resistance (MDR) among GN bacteria. We conducted a case-control study in order to identify factors associated with bacteremia due to multidrug-resistant Gram-negative (MDRGN) isolates in hematopoietic stem cell transplant recipients. Ten patients with MDRGN bacteremia were compared with 44 patients with GN bacteremia without MDR. Bacteremia due to Burkholderia or Stenotrophomonas sp was excluded from analysis (3 cases), because the possibility of intrinsical resistance. Infection due to MDRGN bacteria occurred in 2.9% of 342 hematopoietic stem cell transplant recipients. Klebsiella pneumoniae was the most frequent MDRGN (4 isolates), followed by Pseudomonas aeruginosa (3 isolates). Among non-MDRGN, P. aeruginosa was the most frequent agent (34%), followed by Escherichia coli (30%). The development of GN bacteremia during the empirical treatment of febrile neutropenia (breakthrough bacteremia) was associated with MDR (P < 0.001, odds ratio = 32, 95% confidence interval = 5_190) by multivariate analysis. Bacteremia due to MDRGN bacteria was associated with a higher death rate by univariate analysis (40 vs 9%; P = 0.03). We were unable to identify risk factors on admission or at the time of the first fever, but the occurrence of breakthrough bacteremia was strongly associated with MDRGN bacteria. An immediate change in the antibiotic regimen in such circumstances may improve the prognosis of these patients.

Pharmacokinetic/pharmacodynamic target attainment of intravenous β-lactam regimens against Gram-negative bacteria isolated in a Brazilian teaching hospital

ABSTRACTINTRODUCTION: Monte Carlo simulations have been used for selecting optimal antibiotic regimens for treatment of bacterial infections. The aim of this study was to assess the pharmacokinetic and pharmacodynamic target attainment of intravenous β-lactam regimens commonly used to treat bloodstream infections (BSIs) caused by Gram-negative rod-shaped organisms in a Brazilian teaching hospital.METHODS: In total, 5,000 patients were included in the Monte Carlo simulations of distinct antimicrobial regimens to estimate the likelihood of achieving free drug concentrations above the minimum inhibitory concentration (MIC; fT > MIC) for the requisite periods to clear distinct target organisms. Microbiological data were obtained from blood culture isolates harvested in our hospital from 2008 to 2010.RESULTS: In total, 614 bacterial isolates, including Escherichia coli, Enterobacterspp., Klebsiella pneumoniae, Acinetobacter baumannii, and Pseudomonas aeruginosa, were analyzed Piperacillin/tazobactam failed to achieve a cumulative fraction of response (CFR) > 90% for any of the isolates. While standard dosing (short infusion) of β-lactams achieved target attainment for BSIs caused by E. coliand Enterobacterspp., pharmacodynamic target attainment against K. pneumoniaeisolates was only achieved with ceftazidime and meropenem (prolonged infusion). Lastly, only prolonged infusion of high-dose meropenem approached an ideal CFR against P. aeruginosa; however, no antimicrobial regimen achieved an ideal CFR against A. baumannii.CONCLUSIONS:These data reinforce the use of prolonged infusions of high-dose β-lactam antimicrobials as a reasonable strategy for the treatment of BSIs caused by multidrug resistant Gram-negative bacteria in Brazil.

Application of control measures for infections caused by multi-resistant gram-negative bacteria in intensive care unit patients

Multi-resistant gram-negative rods are important pathogens in intensive care units (ICU), cause high rates of mortality, and need infection control measures to avoid spread to another patients. This study was undertaken prospectively with all of the patients hospitalized at ICU, Anesthesiology of the Hospital São Paulo, using the ICU component of the National Nosocomial Infection Surveillance System (NNIS) methodology, between March 1, 1997 and June 30, 1998. Hospital infections occurring during the first three months after the establishment of prevention and control measures (3/1/97 to 5/31/97) were compared to those of the last three months (3/1/98 to 5/31/98). In this period, 933 NNIS patients were studied, with 139 during the first period and 211 in the second period. The overall rates of infection by multi-resistant microorganisms in the first and second periods were, respectively, urinary tract infection: 3.28/1000 patients/day; 2.5/1000 patients/day; pneumonia: 2.10/1000 patients/day; 5.0/1000 patients/day; bloodstream infection: 1.09/1000 patients/day; 2.5/1000 patients/day. A comparison between overall infection rates of both periods (Wilcoxon test) showed no statistical significance (p = 0.067). The use of intervention measures effectively decreased the hospital bloodstream infection rate (p < 0.001), which shows that control measures in ICU can contribute to preventing hospital infections.

Anticancer and antibacterial secondary metabolites from the endophytic fungus Penicillium sp. CAM64 against multi-drug resistant Gram-negative bacteria.

Background: The emergence of multiple-drug resistance bacteria has become a major threat and thus calls for an urgent need to search for new effective and safe anti-bacterial agents. Objectives: This study aims to evaluate the anticancer and antibacterial activities of secondary metabolites from Penicillium sp. , an endophytic fungus associated with leaves of Garcinia nobilis . Methods: The culture filtrate from the fermentation of Penicillium sp. was extracted and analyzed by liquid chromatography– mass spectrometry, and the major metabolites were isolated and identified by spectroscopic analyses and by comparison with published data. The antibacterial activity of the compounds was assessed by broth microdilution method while the anticancer activity was determined by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. Results: The fractionation of the crude extract afforded penialidin A-C (1-3), citromycetin (4), p-hydroxyphenylglyoxalaldoxime (5) and brefelfin A (6). All of the compounds tested here showed antibacterial activity (MIC = 0.50 – 128 μg/mL) against Gramnegative multi-drug resistance bacteria, Vibrio cholerae (causative agent of dreadful disease cholera) and Shigella flexneri (causative agent of shigellosis), as well as the significant anticancer activity (LC50 = 0.88 – 9.21 μg/mL) against HeLa cells. Conclusion: The results obtained indicate that compounds 1-6 showed good antibacterial and anticancer activities with no toxicity to human red blood cells and normal Vero cells.

The challenge of multidrug resistance: The treatment of gram-negative rod infections

Infections caused by multidrug-resistant gram-negative bacteria are an increasing problem worldwide. Treatment of these microorganisms is a challenge because resistance limits dramatically therapeutic options. In this review, we discuss data of in vitro susceptibility and clinical studies of possible agents for the management of these infections. Currently, published data are limited, and there are no randomized clinical trials involving the treatment of infections caused by multidrug-resistant gram-negative rods. For imipenem-resistant Acinetobacter spp., most studied options are polymyxins and sulbactam. No newer antimicrobials active against Pseudomonas aeruginosa are available or under investigation. Tigecycline presents a broad spectrum of activity in vitro but has been studied mainly as treatment of community-acquired infections, as has ertapenem. They are potential options against extended-spectrum P-lactamase-producing Enterobacteriaceae, and tigecycline may be useful in treating Acinetobacter infections.

Array based detection of antibiotic resistance genes in Gram negative bacteria isolated from retail poultry meat in the UK and Ireland

The use of antibiotics in birds and animals intended for human consumption within the European Union (EU) and elsewhere has been subject to regulation prohibiting the use of antimicrobials as growth promoters and the use of last resort antibiotics in an attempt to reduce the spread of multi-resistant Gram negative bacteria. Given the inexorable spread of antibiotic resistance there is an increasing need for improved monitoring of our food. Using selective media, Gram negative bacteria were isolated from retail chicken of UK-Intensively reared (n = 27), Irish-Intensively reared (n = 19) and UK-Free range (n = 30) origin and subjected to an oligonucleotide based array system for the detection of 47 clinically relevant antibiotic resistance genes (ARGs) and two integrase genes. High incidences of β-lactamase genes were noted in all sample types, acc (67%), cmy (80%), fox (55%) and tem (40%) while chloramphenicol resistant determinants were detected in bacteria from the UK poultry portions and were absent in bacteria from the Irish samples. Denaturing Gradient Gel Electrophoresis (DGGE) was used to qualitatively analyse the Gram negative population in the samples and showed the expected diversity based on band stabbing and DNA sequencing. The array system proved to be a quick method for the detection of antibiotic resistance gene (ARG) burden within a mixed Gram negative bacterial population.

Extended-spectrum cephalosporin-resistant Gram-negative organisms in livestock: an emerging problem for human health?

Escherichia coli, Salmonella spp. and Acinetobacter spp. are important human pathogens. Serious infections due to these organisms are usually treated with extended-spectrum cephalosporins (ESCs). However, in the past two decades we have faced a rapid increasing of infections and colonization caused by ESC-resistant (ESC-R) isolates due to production of extended-spectrum-β-lactamases (ESBLs), plasmid-mediated AmpCs (pAmpCs) and/or carbapenemase enzymes. This situation limits drastically our therapeutic armamentarium and puts under peril the human health. Animals are considered as potential reservoirs of multidrug-resistant (MDR) Gram-negative organisms. The massive and indiscriminate use of antibiotics in veterinary medicine has contributed to the selection of ESC-R E. coli, ESC-R Salmonella spp. and, to less extent, MDR Acinetobacter spp. among animals, food, and environment. This complex scenario is responsible for the expansion of these MDR organisms which may have life-threatening clinical significance. Nowadays, the prevalence of food-producing animals carrying ESC-R E. coli and ESC-R Salmonella (especially those producing CTX-M-type ESBLs and the CMY-2 pAmpC) has reached worryingly high values. More recently, the appearance of carbapenem-resistant isolates (i.e., VIM-1-producing Enterobacteriaceae and NDM-1 or OXA-23-producing Acinetobacter spp.) in livestock has even drawn greater concerns. In this review, we describe the aspects related to the spread of the above MDR organisms among pigs, cattle, and poultry, focusing on epidemiology, molecular mechanisms of resistance, impact of antibiotic use, and strategies to contain the overall problem. The link and the impact of ESC-R organisms of livestock origin for the human scenario are also discussed.

Genomics of pyrrolnitrin biosynthetic loci: evidence for conservation and whole-operon mobility within Gram-negative bacteria

Pyrrolnitrin (PRN) is a tryptophan-derived secondary metabolite produced by a narrow range of Gram-negative bacteria. The PRN biosynthesis by rhizobacteria presumably has a key role in their life strategies and in the biocontrol of plant diseases. The biosynthetic operon that encodes the pathway that converts tryptophan to PRN is composed of four genes, prnA through D, whose diversity, genomic context and spread over bacterial genomes are poorly understood. Therefore, we launched an endeavour aimed at retrieving, by in vitro and in silico means, diverse bacteria carrying the prnABCD biosynthetic loci in their genomes. Analysis of polymorphisms of the prnD gene sequences revealed a high level of conservation between Burkholderia, Pseudomonas and Serratia spp. derived sequences. Whole-operon- and prnD-based phylogeny resulted in tree topologies that are incongruent with the taxonomic status of the evaluated strains as predicted by 16S rRNA gene phylogeny. The genomic composition of c. 20 kb DNA fragments containg the PRN operon varied in different strains. Highly conserved and distinct transposase-encoding genes surrounding the PRN biosynthetic operons of Burkholderia pseudomallei strains were found. A prnABCD-deprived genomic region in B. pseudomallei strain K96243 contained the same gene composition as, and shared high homology with, the flanking regions of the PRN operon in B. pseudomallei strains 668, 1106a and 1710b. Our results strongly suggest that the PRN biosynthetic operon is mobile. The extent, frequency and promiscuity of this mobility remain to be understood.

The genetics of glycosylation in Gram-negative bacteria

In recent years there has been a dramatic increase in reports of glycosylation of proteins in various Gram-negative systems including Neisseria meningitidis, Neisseria gonorrhoeae, Campylobacter jejuni, Pseudomonas aeruginosa, Escherichia coli, Caulobacter crescentus, Aeromonas caviae and Helicobacter pylori. Although this growing list contains many important pathogens (reviewed by Benz and Schmidt [Mol. Microbiol. 45 (2002) 267-276]) and the glycosylations are found on proteins important in pathogenesis such as pili, adhesins and flagella the precise role(s) of the glycosylation of these proteins remains to be determined. Furthermore, the details of the glycosylation biosynthetic process have not been determined in any of these systems. The definition of the precise role of glycosylation and the mechanism of biosynthesis will be facilitated by a detailed understanding of the genes involved. (C) 2002 Federation of European Microbiological Societies. Published by Elsevier Science B.V. All rights reserved.

Antibacterial effect (in vitro) of Moringa oleifera and Annona muricata against Gram positive and Gram negative bacteria

Antibacterial effects of aqueous and ethanolic extracts of seeds of moringa (Moringa oleifera) and pods of soursop (Annona muricata) in the concentration of 1:5 and 1:10 in volumes 50, 100, 150 and 200 µL were examined against Staphylococcus aureus, Vibrio cholerae, Escherichia coli (isolated from the organism and the aquatic environment) and Salmonella Enteritidis. Antibacterial activity (inhibition halo > 13 mm) against S. aureus, V. cholerae and E. coli isolated from the whiteleg shrimp, Litopenaeus vannmaei, was detected in aqueous and ethanolic extracts of moringa. E. coli isolated from tilapiafish, Oreochromis niloticus, was sensitive to the ethanolic extract of moringa. The aqueous extracts of soursop showed an antibacterial effect against S. aureus and V. cholerae, but the antibacterial activity by the ethanol extracts of this plant was not demonstrated.

Crystal structure of the zinc-, cobalt-, and iron-containing adenylate kinase from Desulfovibrio gigas: a novel metal-containing adenylate kinase from Gram-negative bacteria

J Biol Inorg Chem (2011) 16:51–61 DOI 10.1007/s00775-010-0700-8

The interaction of Gram negative bacteria and S. mansoni in mice with experimental Schistosomiasis

Animals (122 mice) were infected each with eighty cercariae of S. mansoni and subsequently challenged intravenously eight weeks later with the following gram-negative organisms. S. typhi, E. coli, Klebsiella-enterobacter species, Proteus mirabilis and Pseudomonas aeruginosa. Enumeration of bacteria in the liver, spleen and blood and S. mansoni from the portal sistem was performed from one to four weeks later in infected animals. A significant difference between infection produced by S. typhi and other gram negative organisms was observed: S. typhi persisted longer in the spleen and liver and could be recovered from S. mansoni worms up to three weeks following bacterial infection. Other gram negative bacteria disappeared from S. mansoni worms after two weeks of initial challenge. Additional animals (51 mice) infected with S. mansoni were given S. typhi, E. coli or sterile saline. After two weeks, animals were sacrificed and the recovery rate of worms from the portal system, and the mesenteric and hepatic oogram were determined. in animals infected with E. coli a significant decrease in the number of worms was observed compared to the saline control group; thirty worms were recovered in the control group compared to two worms in e. coli infected animals. In addition, the patterns of oviposition was significantly different in these latter animals suggesting complete inhibition of this process. Following S. typhi infection the difference in recovery of worms and pattern of oviposition was minimal. These findings suggest a difference in the interaction of various gram negative bacteria and S. mansoni and are consistent with the clinical observation of prolonged salmonella bacteremia in patients with schistosomiasis.

Macrophage Migration Inhibitory Factor Deficiency Is Associated With Impaired Killing of Gram-Negative Bacteria by Macrophages and Increased Susceptibility to Klebsiella pneumoniae Sepsis.

The cytokine macrophage migration inhibitory factor (MIF) is an important component of the early proinflammatory response of the innate immune system. However, the antimicrobial defense mechanisms mediated by MIF remain fairly mysterious. In the present study, we examined whether MIF controls bacterial uptake and clearance by professional phagocytes, using wild-type and MIF-deficient macrophages. MIF deficiency did not affect bacterial phagocytosis, but it strongly impaired the killing of gram-negative bacteria by macrophages and host defenses against gram-negative bacterial infection, as shown by increased mortality in a Klebsiella pneumonia model. Consistent with MIF's regulatory role of Toll-like 4 expression in macrophages, MIF-deficient cells stimulated with lipopolysaccharide or Escherichia coli exhibited reduced nuclear factor κB activity and tumor necrosis factor (TNF) production. Addition of recombinant MIF or TNF corrected the killing defect of MIF-deficient macrophages. Together, these data show that MIF is a key mediator of host responses against gram-negative bacteria, acting in part via a modulation of bacterial killing by macrophages.

Regulatory roles of the GacS/GacA two-component system in plant-associated and other gram-negative bacteria.

The sensor kinase GacS and the response regulator GacA are members of a two-component system that is present in a wide variety of gram-negative bacteria and has been studied mainly in enteric bacteria and fluorescent pseudomonads. The GacS/GacA system controls the production of secondary metabolites and extracellular enzymes involved in pathogenicity to plants and animals, biocontrol of soilborne plant diseases, ecological fitness, or tolerance to stress. A current model proposes that GacS senses a still-unknown signal and activates, via a phosphorelay mechanism, the GacA transcription regulator, which in turn triggers the expression of target genes. The GacS protein belongs to the unorthodox sensor kinases, characterized by an autophosphorylation, a receiver, and an output domain. The periplasmic loop domain of GacS is poorly conserved in diverse bacteria. Thus, a common signal interacting with this domain would be unexpected. Based on a comparison with the transcriptional regulator NarL, a secondary structure can be predicted for the GacA sensor kinases. Certain genes whose expression is regulated by the GacS/GacA system are regulated in parallel by the small RNA binding protein RsmA (CsrA) at a posttranscriptional level. It is suggested that the GacS/GacA system operates a switch between primary and secondary metabolism, with a major involvement of posttranscriptional control mechanisms.

Sensing Gram-negative bacteria: a phylogenetic perspective.

Gram-negative bacteria represent a major group of pathogens that infect all eukaryotes from plants to mammals. Gram-negative microbe-associated molecular patterns include lipopolysaccharides and peptidoglycans, major immunostimulatory determinants across phyla. Recent advances have furthered our understanding of Gram-negative detection beyond the well-defined pattern recognition receptors such as TLR4. A B-type lectin receptor for LPS and Lysine-motif containing receptors for peptidoglycans were recently added to the plant arsenal. Caspases join the ranks of mammalian cytosolic immune detectors by binding LPS, and make TLR4 redundant for septic shock. Fascinating bacterial evasion mechanisms lure the host into tolerance or promote inter-bacterial competition. Our review aims to cover recent advances on bacterial messages and host decoding systems across phyla, and highlight evolutionarily recurrent strategies.