| 1. |
- Holmgren, Jan, 1944, et al.
(författare)
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Mucosal adjuvants and anti-infection and anti-immunopathology vaccines based on cholera toxin, cholera toxin B subunit and CpG DNA
- 2005
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Ingår i: Immunology Letters. - : Elsevier. - 0165-2478 .- 1879-0542. ; 97:2, s. 181-8
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Tidskriftsartikel (refereegranskat)abstract
- Mucosal immunisation may be used both to protect the mucosal surfaces against infections and as a means for immunological treatment of peripheral immunopathological disorders through the induction of systemic antigen-specific tolerance ('oral tolerance'). The development of mucosal vaccines, whether for prevention of infectious diseases or for oral tolerance immunotherapy, requires efficient antigen delivery and adjuvant systems that can help to present the appropriate vaccine or immunotherapy antigens to the mucosal immune system. The most potent (but also toxic) mucosal adjuvants are cholera toxin (CT) and the closely related Escherichia coli heat-labile enterotoxin (LT), and much effort and significant progress have been made recently to generate toxicologically acceptable derivatives of these toxins with retained adjuvant activity. Among these are the non-toxic, recombinantly produced cholera toxin B-subunit (CTB). CTB is a specific protective antigen component of a widely registered oral cholera vaccine as well as a promising vector for either giving rise to mucosal anti-infective immunity or for inducing peripheral anti-inflammatory tolerance to chemically or genetically linked foreign antigens administered mucosally. CT and CTB have also recently been used as combined vectors and adjuvants for markedly promoting ex vivo dendritic cell (DC) vaccination with different antigens and also steering the immune response to the in vivo-reinfused DCs towards either broad Th1 + Th2 + CTL immunity (CT) or Th2 or tolerance (CTB). Another type of mucosal adjuvants is represented by bacterial DNA or synthetic oligodeoxynucleotides containing CpG-motifs, which especially when linked to CTB have been found to effectively stimulate both innate and adaptive mucosal immune responses. The properties and clinical potential of these different classes of adjuvants are being discussed. © 2004 Elsevier B.V. All rights reserved.
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| 2. |
- Boulund, Fredrik, 1985, et al.
(författare)
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Computational discovery and functional validation of novel fluoroquinolone resistance genes in public metagenomic data sets
- 2017
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Ingår i: BMC Genomics. - : Springer Science and Business Media LLC. - 1471-2164. ; 18:1, s. Art 682-
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Tidskriftsartikel (refereegranskat)abstract
- Background: Fluoroquinolones are broad-spectrum antibiotics used to prevent and treat a wide range of bacterial infections. Plasmid-mediated qnr genes provide resistance to fluoroquinolones in many bacterial species and are increasingly encountered in clinical settings. Over the last decade, several families of qnr genes have been discovered and characterized, but their true prevalence and diversity still remain unclear. In particular, environmental and host-associated bacterial communities have been hypothesized to maintain a large and unknown collection of qnr genes that could be mobilized into pathogens. Results: In this study we used computational methods to screen genomes and metagenomes for novel qnr genes. In contrast to previous studies, we analyzed an almost 20-fold larger dataset comprising almost 13 terabases of sequence data. In total, 362,843 potential qnr gene fragments were identified, from which 611 putative qnr genes were reconstructed. These gene sequences included all previously described plasmid-mediated qnr gene families. Fifty-two of the 611 identified qnr genes were reconstructed from metagenomes, and 20 of these were previously undescribed. All of the novel qnr genes were assembled from metagenomes associated with aquatic environments. Nine of the novel genes were selected for validation, and six of the tested genes conferred consistently decreased susceptibility to ciprofloxacin when expressed in Escherichia coli. Conclusions: The results presented in this study provide additional evidence for the ubiquitous presence of qnr genes in environmental microbial communities, expand the number of known qnr gene variants and further elucidate the diversity of this class of resistance genes. This study also strengthens the hypothesis that environmental bacterial communities act as sources of previously uncharacterized qnr genes.
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| 3. |
- Flach, Carl-Fredrik, 1977, et al.
(författare)
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Does antifouling paint select for antibiotic resistance?
- 2017
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Ingår i: Science of the Total Environment. - : Elsevier BV. - 0048-9697 .- 1879-1026. ; 590, s. 461-468
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Tidskriftsartikel (refereegranskat)abstract
- There is concern that heavy metals and biocides contribute to the development of antibiotic resistance via co-selection. Most antifouling paints contain high amounts of such substances, which risks turning painted ship hulls into highly mobile refuges and breeding grounds for antibiotic-resistant bacteria. The objectives of this study were to start investigate if heavy-metal based antifouling paints can pose a risk for co-selection of antibiotic-resistant bacteria and, if so, identify the underlying genetic basis. Plastic panels with one side painted with copper and zinc-containing antifouling paint were submerged in a Swedish marina and biofilms from both sides of the panels were harvested after 2.5-4 weeks. DNA was isolated from the biofilms and subjected to metagenomic sequencing. Biofilm bacteria were cultured on marine agar supplemented with tetracycline, gentamicin, copper sulfate or zinc sulfate. Biofilm communities from painted surfaces displayed lower taxonomic diversity and enrichment of Gammaproteobacteria. Bacteria from these communities showed increased resistance to both heavy metals and tetracycline but not to gentamicin. Significantly higher abundance of metal and biocide resistance genes was observed, whereas mobile antibiotic resistance genes were not enriched in these communities. In contrast, we found an enrichment of chromosomal RND efflux system genes, including such with documented ability to confer decreased susceptibility to both antibiotics and biocides/heavy metals. This was paralleled by increased abundances of integron-associated integrase and ISCR transposase genes. The results show that the heavy metal-based antifouling paint exerts a strong selection pressure on marine bacterial communities and can co-select for certain antibiotic-resistant bacteria, likely by favoring species and strains carrying genes that provide cross-resistance. Although this does not indicate an immediate risk for promotion of mobile antibiotic resistance, the clear increase of genes involved in mobilizing DNA provides a foundation for increased opportunities for gene transfer in such communities, which might also involve yet unknown resistance mechanisms.
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| 4. |
- Flach, Carl-Fredrik, 1977, et al.
(författare)
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Functional verification of computationally predicted qnr genes
- 2013
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Ingår i: Annals of Clinical Microbiology and Antimicrobials. - : Springer Science and Business Media LLC. - 1476-0711. ; 12
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Tidskriftsartikel (refereegranskat)abstract
- Background The quinolone resistance (qnr) genes are widely distributed among bacteria. We recently developed and applied probabilistic models to identify tentative novel qnr genes in large public collections of DNA sequence data including fragmented metagenomes. Findings: By using inducible recombinant expressions systems the functionality of four identified qnr candidates were evaluated in Escherichia coli. Expression of several known qnr genes as well as two novel candidates provided fluoroquinolone resistance that increased with elevated inducer concentrations. The two novel, functionally verified qnr genes are termed Vfuqnr and assembled qnr 1. Co-expression of two qnr genes suggested non-synergistic action. Conclusion The combination of a computational model and recombinant expression systems provides opportunities to explore and identify novel antibiotic resistance genes in both genomic and metagenomic datasets.
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| 5. |
- Bengtsson-Palme, Johan, 1985, et al.
(författare)
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Elucidating selection processes for antibiotic resistance in sewage treatment plants using metagenomics
- 2016
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Ingår i: Science of the Total Environment. - : Elsevier BV. - 0048-9697 .- 1879-1026. ; 572, s. 697-712
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Tidskriftsartikel (refereegranskat)abstract
- Sewage treatment plants (STPs) have repeatedly been suggested as “hotspots” for the emergence and dissemination of antibiotic-resistant bacteria. A critical question still unanswered is if selection pressures within STPs, caused by residual antibiotics or other co-selective agents, are sufficient to specifically promote resistance. To address this, we employed shotgun metagenomic sequencing of samples from different steps of the treatment process in three Swedish STPs. In parallel, concentrations of selected antibiotics, biocides and metals were analyzed. We found that concentrations of tetracycline and ciprofloxacin in the influent were above predicted concentrations for resistance selection, however, there was no consistent enrichment of resistance genes to any particular class of antibiotics in the STPs, neither for biocide and metal resistance genes. The most substantial change of the bacterial communities compared to human feces occurred already in the sewage pipes, manifested by a strong shift from obligate to facultative anaerobes. Through the treatment process, resistance genes against antibiotics, biocides and metals were not reduced to the same extent as fecal bacteria. The OXA-48 gene was consistently enriched in surplus and digested sludge. We find this worrying as OXA-48, still rare in Swedish clinical isolates, provides resistance to carbapenems, one of our most critically important classes of antibiotics. Taken together, metagenomics analyses did not provide clear support for specific antibiotic resistance selection. However, stronger selective forces affecting gross taxonomic composition, and with that resistance gene abundances, limit interpretability. Comprehensive analyses of resistant/non-resistant strains within relevant species are therefore warranted.
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| 6. |
- Berglund, Fanny, 1988, et al.
(författare)
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Identification of 76 novel B1 metallo-beta-lactamases through large-scale screening of genomic and metagenomic data
- 2017
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Ingår i: Microbiome. - : Springer Science and Business Media LLC. - 2049-2618. ; 5:1, s. 134-134
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Tidskriftsartikel (refereegranskat)abstract
- Background: Metallo-beta-lactamases are bacterial enzymes that provide resistance to carbapenems, the most potent class of antibiotics. These enzymes are commonly encoded on mobile genetic elements, which, together with their broad substrate spectrum and lack of clinically useful inhibitors, make them a particularly problematic class of antibiotic resistance determinants. We hypothesized that there is a large and unexplored reservoir of unknown metallo-beta-lactamases, some of which may spread to pathogens, thereby threatening public health. The aim of this study was to identify novel metallo-beta-lactamases of class B1, the most clinically important subclass of these enzymes. Results: Based on a new computational method using an optimized hidden Markov model, we analyzed over 10,000 bacterial genomes and plasmids together with more than 5 terabases of metagenomic data to identify novel metallo-beta-lactamase genes. In total, 76 novel genes were predicted, forming 59 previously undescribed metallo-beta-lactamase gene families. The ability to hydrolyze imipenem in an Escherichia coli host was experimentally confirmed for 18 of the 21 tested genes. Two of the novel B1 metallo-beta-lactamase genes contained atypical zinc-binding motifs in their active sites, which were previously undescribed for metallo-beta-lactamases. Phylogenetic analysis showed that B1 metallo-beta-lactamases could be divided into five major groups based on their evolutionary origin. Our results also show that, except for one, all of the previously characterized mobile B1 beta-lactamases are likely to have originated from chromosomal genes present in Shewanella spp. and other Proteobacterial species. Conclusions: This study more than doubles the number of known B1 metallo-beta-lactamases. The findings have further elucidated the diversity and evolutionary history of this important class of antibiotic resistance genes and prepare us for some of the challenges that may be faced in clinics in the future.
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| 7. |
- Berglund, Fanny, et al.
(författare)
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The resistome and microbiome of wastewater treatment plant workers - The AWARE study.
- 2023
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Ingår i: Environment international. - 0160-4120 .- 1873-6750. ; 180
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Tidskriftsartikel (refereegranskat)abstract
- Urban wastewater treatment plants harbor a large collection of antibiotic resistant enteric bacteria. It is therefore reasonable to hypothesize that workers at such plants would possess a more diverse set of resistant enteric bacteria, compared to the general population. To address this hypothesis, we have compared the fecal microbiome and resistome of 87 workers at wastewater treatment plants (WWTPs) from Romania and the Netherlands to those of 87 control individuals, using shotgun metagenomics. Controlling for potential confounders, neither the total antibiotic resistance gene (ARG) abundance, nor the overall bacterial composition were significantly different between the two groups. If anything, the ARG richness was slightly lower in WWTP workers, and in a stratified analysis the total ARG abundance was significantly lower in Dutch workers compared to Dutch control participants. We identified country of residence, together with recent antibiotic intake in the Dutch population, as the largest contributing factors to the total abundance of ARGs. A striking side-finding was that sex was associated with carriage of disinfectant resistance genes, with women in both Romania and the Netherlands having significantly higher abundance compared to men. A follow up investigation including an additional 313 publicly available samples from healthy individuals from three additional countries showed that the difference was significant for three genes conferring resistance to chemicals commonly used in cosmetics and cleaning products. We therefore hypothesize that the use of cosmetics and, possibly, cleaning products leads to higher abundance of disinfectant resistance genes in the microbiome of the users. Altogether, this study shows that working at a WWTP does not lead to a higher abundance or diversity of ARGs and no large shifts in the overall gut microbial composition in comparison to participants not working at a WWTP. Instead, other factors such as country of residence, recent antibiotic intake and sex seem to play a larger role.
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| 8. |
- Bobis Camacho, Julián, 1997, et al.
(författare)
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Evaluation of culture conditions for sewage-based surveillance of antibiotic resistance in Klebsiella pneumoniae.
- 2024
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Ingår i: Journal of global antimicrobial resistance. - 2213-7173.
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Tidskriftsartikel (refereegranskat)abstract
- Recent studies have shown promise in predicting clinical antibiotic resistance rates from sewage data. Few have focused on Klebsiella pneumoniae, despite its virulence and importance as carrier of antibiotic resistance. Several media have been suggested for the isolation of K. pneumoniae from complex samples. However, comprehensive evaluations of culture protocols for isolation of K. pneumoniae from sewage are lacking.Here, influent samples from a major Swedish sewage treatment plant were used to evaluate ten culture conditions in parallel: cultivation on Brilliant green containing Inositol-Nitrate-Deoxycholate agar (BIND), Bruce agar, Klebsiella ChromoSelect Selective agar®, MacConkey-Inositol-Carbenicillin, or Simmons Citrate Agar with Inositol (SCAI) incubated at either 37˚C or 42˚C for 44 h. The culture conditions were compared based on colony counts of presumed K. pneumoniae and identification precision assessed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry.The sensitivity was lowest for BIND, whereas it was similar for the other media irrespective of incubation temperature. For four media, a better precision was observed after incubation at 42˚C compared to 37˚C, to a large extent explained by a lower frequency of captured Klebsiella oxytoca. SCAI incubated at 42˚C showed the highest precision (84.4%). By combining this protocol with subsequent antibiotic resistance screening of collected isolates, low resistance rates in sewage K. pneumoniae were revealed, potentially reflecting the local resistance landscape.When combined with downstream analyses, SCAI incubated at 42˚C could be a valuable culture protocol for sewage-based studies on various aspects of K. pneumoniae epidemiology including antibiotic resistance prevalence.
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| 9. |
- Böhm, Maria-Elisabeth, et al.
(författare)
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A novel, integron-regulated, class c β-lactamase
- 2020
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Ingår i: Antibiotics. - : MDPI AG. - 2079-6382. ; 9:3
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Tidskriftsartikel (refereegranskat)abstract
- © 2020 by the authors. Licensee MDPI, Basel, Switzerland. AmpC-type β-lactamases severely impair treatment of many bacterial infections, due to their broad spectrum (they hydrolyze virtually all β-lactams, except fourth-generation cephalosporins and carbapenems) and the increasing incidence of plasmid-mediated versions. The original chromosomal AmpCs are often tightly regulated, and their expression is induced in response to exposure to β-lactams. Regulation of mobile ampC expression is in many cases less controlled, giving rise to constitutively resistant strains with increased potential for development or acquisition of additional resistances. We present here the identification of two integron-encoded ampC genes, blaIDC-1 and blaIDC-2 (integron-derived cephalosporinase), with less than 85% amino acid sequence identity to any previously annotated AmpC. While their resistance pattern identifies them as class C β-lactamases, their low isoelectric point (pI) values make differentiation from other β-lactamases by isoelectric focusing impossible. To the best of our knowledge, this is the first evidence of an ampC gene cassette within a class 1 integron, providing a mobile context with profound potential for transfer and spread into clinics. It also allows bacteria to adapt expression levels, and thus reduce fitness costs, e.g., by cassette-reshuffling. Analyses of public metagenomes, including sewage metagenomes, show that the discovered ampCs are primarily found in Asian countries.
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| 10. |
- Böhm, Maria-Elisabeth, et al.
(författare)
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Discovery of a novel integron-borne aminoglycoside resistance gene present in clinical pathogens by screening environmental bacterial communities.
- 2020
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Ingår i: Microbiome. - : Springer Science and Business Media LLC. - 2049-2618. ; 8:1
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Tidskriftsartikel (refereegranskat)abstract
- New antibiotic resistance determinants are generally discovered too late, long after they have irreversibly emerged in pathogens and spread widely. Early discovery of resistance genes, before or soon after their transfer to pathogens could allow more effective measures to monitor and reduce spread, and facilitate genetics-based diagnostics.We modified a functional metagenomics approach followed by in silico filtering of known resistance genes to discover novel, mobilised resistance genes in class 1 integrons in wastewater-impacted environments. We identified an integron-borne gene cassette encoding a protein that conveys high-level resistance against aminoglycosides with a garosamine moiety when expressed in E. coli. The gene is named gar (garosamine-specific aminoglycoside resistance) after its specificity. It contains none of the functional domains of known aminoglycoside modifying enzymes, but bears characteristics of a kinase. By searching public databases, we found that the gene occurs in three sequenced, multi-resistant clinical isolates (two Pseudomonas aeruginosa and one Luteimonas sp.) from Italy and China, respectively, as well as in two food-borne Salmonella enterica isolates from the USA. In all cases, gar has escaped discovery until now.To the best of our knowledge, this is the first time a novel resistance gene, present in clinical isolates, has been discovered by exploring the environmental microbiome. The gar gene has spread horizontally to different species on at least three continents, further limiting treatment options for bacterial infections. Its specificity to garosamine-containing aminoglycosides may reduce the usefulness of the newest semisynthetic aminoglycoside plazomicin, which is designed to avoid common aminoglycoside resistance mechanisms. Since the gene appears to be not yet common in the clinics, the data presented here enables early surveillance and maybe even mitigation of its spread.
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