187 research outputs found
Significant variability exists in the cytotoxicity of global methicillin-resistant Staphylococcus aureus lineages.
Staphylococcus aureus is a major human pathogen where the emergence of antibiotic resistant lineages, such as methicillin-resistant S. aureus (MRSA), is a major health concern. While some MRSA lineages are restricted to the healthcare setting, the epidemiology of MRSA is changing globally, with the rise of specific lineages causing disease in healthy people in the community. In the past two decades, community-associated MRSA (CA-MRSA) has emerged as a clinically important and virulent pathogen associated with serious skin and soft-tissue infections (SSTI). These infections are primarily cytotoxin driven, leading to the suggestion that hypervirulent lineages/multi-locus sequence types (STs) exist. To examine this, we compared the cytotoxicity of 475 MRSA isolates representing five major MRSA STs (ST22, ST93, ST8, ST239 and ST36) by employing a monocyte-macrophage THP-1 cell line as a surrogate for measuring gross cytotoxicity. We demonstrate that while certain MRSA STs contain highly toxic isolates, there is such variability within lineages to suggest that this aspect of virulence should not be inferred from the genotype of any given isolate. Furthermore, by interrogating the accessory gene regulator (Agr) sequences in this collection we identified several Agr mutations that were associated with reduced cytotoxicity. Interestingly, the majority of isolates that were attenuated in cytotoxin production contained no mutations in the agr locus, indicating a role of other undefined genes in S. aureus toxin regulation
Different bacterial gene expression patterns and attenuated host immune responses are associated with the evolution of low-level vancomycin resistance during persistent methicillin-resistant Staphylococcus aureus bacteraemia
BACKGROUND: Low-level vancomycin resistance in Staphylococcus aureus (vancomycin-intermediate S. aureus (VISA) and hetero-VISA [hVISA]) emerges during persistent infection and failed vancomycin therapy. Up-regulation of genes associated with the "cell wall stimulon" and mutations in the vraSR operon have both been implicated in the development of resistance, however the molecular mechanisms of resistance are not completely understood. To further elucidate the mechanisms leading to resistance transcriptome comparisons were performed using multiple clinical pairs of vancomycin-susceptible S. aureus (VSSA) and hVISA/VISA (n = 5), and three VSSA control pairs from hospitalized patients with persistent bacteraemia that did not develop hVISA/VISA. Based on the transcriptome results multiple genes were sequenced and innate immune system stimulation was assessed in the VSSA and hVISA/VISA pairs. RESULTS: Here we show that up-regulation of vraS and the "cell wall stimulon" is not essential for acquisition of low-level vancomycin resistance and that different transcriptional responses occur, even between closely related hVISA/VISA strains. DNA sequencing of vraSR, saeSR, mgrA, rot, and merR regulatory genes and upstream regions did not reveal any differences between VSSA and hVISA/VISA despite transcriptional changes suggesting mutations in these loci may be linked to resistance in these strains. Enhanced capsule production and reduced protein A expression in hVISA/VISA were confirmed by independent bioassays and fully supported the transcriptome data. None of these changes were observed in the three control pairs that remained vancomycin-susceptible during persistent bacteremia. In a macrophage model of infection the changes in cell surface structures in hVISA/VISA strains were associated with significantly reduced NF-kappaB activation resulting in reduced TNF-alpha and IL-1beta expression. CONCLUSION: We conclude that there are multiple pathways to low-level vancomycin resistance in S. aureus, even among closely related clinical strains, and these can result in an attenuated host immune response. The persistent infections associated with hVISA/VISA strains may be a consequence of changes in host pathogen interactions in addition to the reduced antibiotic susceptibility
Low incidence of recurrent Buruli ulcers in treated Australian patients living in an endemic region
We examined recurrent Buruli ulcer cases following treatment and assumed cure in a large cohort of Australian patients living in an endemic area. We report that while the recurrence rate was low (2.81 cases/year/1000 population), it remained similar to the estimated risk of primary infection within the general population of the endemic area (0.85-4.04 cases/year/1,000 population). The majority of recurrent lesions occurred in different regions of the body and were separated by a median time interval of 44 months. Clinical, treatment and epidemiological factors combined with whole genome sequencing of primary and recurrent isolates suggests that in most recurrent cases a re-infection was more likely as opposed to a relapse of the initial infection. Additionally, all cases occurring more than 12 months after commencement of treatment were likely re-infections. Our study provides important prognostic information for patients and their health care providers concerning the nature and risks associated with recurrent cases of Buruli ulcer in Australia
Mycobacterium ulcerans and Other Mycolactone-Producing Mycobacteria Should Be Considered a Single Species
Polyclonal but not monoclonal circulating memory CD4+ T cells attenuate the severity of Staphylococcus aureus bacteremia
Staphylococcus aureus bacteremia causes significant morbidity and mortality. Treatment of staphylococcal infections is hindered by widespread antibiotic resistance, and attempts to develop an S. aureus vaccine have failed. Improved S. aureus treatment and infection prevention options require a deeper understanding of the correlates of protective immunity. CD4+ T cells have been identified as key orchestrators in the defense against S. aureus, but uncertainties persist regarding the subset, polarity, and breadth of the memory CD4+ T-cell pool required for protection. Here, using a mouse model of systemic S. aureus infection, we discovered that the breadth of bacterium-specific memory CD4+ T-cell pool is a critical factor for protective immunity against invasive S. aureus infections. Seeding mice with a monoclonal bacterium-specific circulating memory CD4+ T-cell population failed to protect against systemic S. aureus infection; however, the introduction of a polyclonal and polyfunctional memory CD4+ T-cell pool significantly reduced the bacterial burden. Our findings support the development of a multi-epitope T-cell-based S. aureus vaccine, as a strategy to mitigate the severity of S. aureus bacteremia
Mycobacterium ulcerans in Mosquitoes Captured during Outbreak of Buruli Ulcer, Southeastern Australia
Mosquitoes positive for M. ulcerans were linked to
outbreaks of Buruli ulcer in humans
Single Nucleotide Polymorphism Typing of Mycobacterium ulcerans Reveals Focal Transmission of Buruli Ulcer in a Highly Endemic Region of Ghana
Buruli ulcer (BU) is an emerging necrotizing disease of the skin and subcutaneous tissue caused by Mycobacterium ulcerans. While proximity to stagnant or slow flowing water bodies is a risk factor for acquiring BU, the epidemiology and mode of M. ulcerans transmission is poorly understood. Here we have used high-throughput DNA sequencing and comparisons of the genomes of seven M. ulcerans isolates that appeared monomorphic by existing typing methods. We identified a limited number of single nucleotide polymorphisms (SNPs) and developed a real-time PCR SNP typing method based on these differences. We then investigated clinical isolates of M. ulcerans on which we had detailed information concerning patient location and time of diagnosis. Within the Densu river basin of Ghana we observed dominance of one clonal complex and local clustering of some of the variants belonging to this complex. These results reveal focal transmission and demonstrate, that micro-epidemiological analyses by SNP typing has great potential to help us understand how M. ulcerans is transmitted
Insulinresistenz im postprandialen Lipidstoffwechsel bei Metabolischem Syndrom : Untersuchung der Glucosetoleranz nach oraler Fettaufnahme
Das Metabolische Syndrom als Risikocluster von Stoffwechselstörungen spielt bei der Entstehung von atherosklerotischen
GefĂ€Ăerkrankungen eine zentrale Rolle. Störungen des Glukose- und Triglyzeridstoffwechsels werden dabei hĂ€ufig gemeinsam
beobachtet. Freie FettsĂ€uren und Insulin werden als Mediatoren im IntermediĂ€rstoffwechsel sowohl fĂŒr physiologische SteuerungsvorgĂ€nge
als auch fĂŒr pathologische VerĂ€nderungen verantwortlich gemacht. Eine Insulinresistenz kann zur Entwicklung einer HyperinsulinĂ€mie
fĂŒhren, die durch direkte Einwirkung auf die Arterienwand und durch BegĂŒnstigung von pathologischen ZustĂ€nden, wie Hypertonie und
DyslipoproteinĂ€mie, die Atherosklerose unterstĂŒtzt. Alternative Vorstellungen gehen von einer Störung des Triglyzeridstoffwechsels mit
Erhöhung freier FettsĂ€uren aus, die ĂŒber eine Reduktion der Glukoseaufnahme der Muskulatur und Beeinflussung von Insulinsekretion und
-wirkung zur Insulinresistenz und HyperinsulinĂ€mie fĂŒhren könnte.
Mit der hier vorgestellten Dissertationsarbeit sollte ein Beitrag dazu geleistet werden, einen klinisch anwendbaren Test zur Diagnostik des
Triglyzerid- und Glukosemetabolismus zu finden, der die Beobachtung der postprandialen Stoffwechselparameter ermöglicht. Das
Augenmerk wurde auf die KonzentrationsverlÀufe von Glucose, Insulin und freien FettsÀuren gerichtet. Auch wurde der Einfluà klinischer
MeĂgröĂen der Adipositas auf die Stoffwechselparameter beleuchtet. Weiterhin sollte untersucht werden, ob die VerĂ€nderungen der
postprandialen Glukosetoleranz zwischen gesunden Probanden und Patienten mit angiographisch gesicherter KHK RĂŒckschlĂŒsse auf
pathologische ZustÀnde bei koronarer Herzerkrankung und Metabolischem Syndrom zulassen. Hierzu wurde das entwickelte Testverfahren
mit je einer Gruppe gesunder Probanden und Patienten, die eine angiographisch gesicherte KHK aufwiesen, durchgefĂŒhrt. Es wurde die
Kombination aus einer Belastung des Probanden mit oral aufgenommenem Fett via naturalis und die eher experimentelle Beobachtung
des Kohlenhydratstoffwechsels mittels intravenösem Glukosetoleranztest gewÀhlt. Zur Anwendung kam ein standardisierter intravenöser
Glukosetoleranztest nĂŒchtern (Test 1) und 4 Stunden nach oraler Aufnahme (Test 2) einer definierten Triglyzeridemulsion.
Die orale Fettgabe fĂŒhrte bei Test 2 zu einem signifikanten Anstieg der Plasmakonzentration der freien FettsĂ€uren gegenĂŒber Test 1 ohne
Fettgabe. Nach Injektion des Glukosebolus und Anstieg des Plasma-Insulins wurde die FettsÀure-Konzentration bei den Probanden
wĂ€hrend Test 2 signifikant höher unterdrĂŒckt als bei Test 1. Die KHK-Patienten erreichten hier keine Zunahme der UnterdrĂŒckung der
FettsĂ€urespiegel. Das Verhalten der Insulin- und Glukosekonzentrationen zeigte eine erhöhte InsulinausschĂŒttung nach vorheriger oraler
Fettzufuhr (Test 2) in den ersten 30 min des Glukosetoleranztests gegenĂŒber Test 1. Dieser Effekt war in der folgenden Stunde bei den
Gesunden nicht mehr nachzuweisen, wÀhrend die KHK-Patienten auch in der SpÀtphase eine erhöhte Insulinkonzentration bei Test 2
gegenĂŒber Test 1 zeigten. Die Glukosekonzentrationen blieben weitgehend unverĂ€ndert in beiden Gruppen. ZusammenhĂ€nge der
Adipositas mit dem Verlauf der Stoffwechseltests wurden durch die Korrelation von KörpermaĂzahlen mit verschiedenen
Testablaufparametern untersucht. Als Adipositas-MeĂwerte wurden der Body-mass-index, die Waist-to-Hip-Ratio und die Ergebnisse der
Bioimpedanzanalyse, welche durch elektrische Hautwiderstandsmessung die Bestimmung des relativen Körperwassergehalts, der
Magermasse und des Körperfettanteil ermöglicht, in die Korrelationsrechung eingeschlossen. Hervorzuheben ist eine konstante positive
Korrelation der Waist-to-Hip-Ratio mit der glucosestimulierten Insulinkonzentration bei beiden Tests und beiden Gruppen. Diese
Ergebnisse deuten auf einen Zusammenhang der Menge des intraabdominalen Fettgewebes mit der Insulinkinetik hin. Die
Insulinkonzentration könnte dabei ĂŒber eine Interaktion einer erhöhten freien FettsĂ€urekonzentration im Portalblut bei zunehmender
StoffwechselaktivitĂ€t des intraabdominalen Fettgewebes beeinfluĂt werden. Die DurchfĂŒhrung der Untersuchung belegte die klinische
Anwendbarkeit eines Belastungstestverfahrens zur Beobachtung der Interaktionen des Glukose- und Triglyzerid-stoffwechsels. Die
Beobachtung einer Zunahme der glukosestimulierten Insulinkonzentration durch orale Fettgabe deutet darauf hin, daà freie FettsÀuren eine
Abnahme der Glukosetoleranz bewirken könnten, die durch eine erhöhte InsulinausschĂŒttung kompensiert wurde. Die Glukosehomöostase
wurde dadurch aufrecht erhalten. Das angewendete Testverfahren konnte im Vergleich zwischen Gesunden und KHK-Patienten eine
prolongierte InsulinÀmie zeigen. Diese könnte an der Atherogenese beteiligt sein. Trotz VerlÀngerung der InsulinÀmie wurde die
FettsÀuresuppression nicht, wie bei den Gesunden Probanden verstÀrkt, was eine reduzierte Insulinwirksamkeit darstellen könnte.
Die Ergebnisse rechtfertigen eine mehr fettstoffwechselbezogene Sichtweise der Pathogenese des Metabolischen Syndroms. Die
Messung der Körperkonstitution könnte bei der RisikoabschÀtzung von Stoffwechselstörungen hilfreich sein. Insbesondere die Messung
des intraabdominalen Fettgewebes ist aufgrund der erhöhten StoffwechselaktivitÀt dabei interessant. Die prospektive Bedeutung solcher
Belastungstests mĂŒĂte anhand gröĂer angelegter Studien mit der Möglichkeit der Bestimmung von Normalbereichen ĂŒberprĂŒft werden
Exposure Risk for Infection and Lack of Human-to-Human Transmission of Mycobacterium ulcerans
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Deciphering the genetic basis for polyketide variation among mycobacteria producing mycolactones.
BACKGROUND: Mycolactones are immunosuppressive and cytotoxic polyketides, comprising five naturally occurring structural variants (named A/B, C, D, E and F), produced by different species of very closely related mycobacteria including the human pathogen, Mycobacterium ulcerans. In M. ulcerans strain Agy99, mycolactone A/B is produced by three highly homologous type I polyketide megasynthases (PKS), whose genes (mlsA1: 51 kb, mlsA2: 7.2 kb and mlsB: 42 kb) are found on a 174 kb plasmid, known as pMUM001. RESULTS: We report here comparative genomic analysis of pMUM001, the complete DNA sequence of a 190 kb megaplasmid (pMUM002) from Mycobacterium liflandii 128FXT and partial sequence of two additional pMUM replicons, combined with liquid chromatography-tandem mass spectrometric (LC-MS/MS) analysis. These data reveal how PKS module and domain differences affecting MlsB correlate with the production of mycolactones E and F. For mycolactone E these differences from MlsB in M. ulcerans Agy99 include replacement of the AT domain of the loading module (acetate to propionate) and the absence of an entire extension module. For mycolactone F there is also a reduction of one extension module but also a swap of ketoreductase domains that explains the characteristic stereochemistry of the two terminal side-chain hydroxyls, an arrangement unique to mycolactone F CONCLUSION: The mycolactone PKS locus on pMUM002 revealed the same large, three-gene structure and extraordinary pattern of near-identical PKS domain sequence repetition as observed in pMUM001 with greater than 98.5% nucleotide identity among domains of the same function. Intra- and inter-strain comparisons suggest that the extreme sequence homogeneity seen among the mls PKS genes is caused by frequent recombination-mediated domain replacement. This work has shed light on the evolution of mycolactone biosynthesis among an unusual group of mycobacteria and highlights the potential of the mls locus to become a toolbox for combinatorial PKS biochemistry.RIGHTS : This article is licensed under the BioMed Central licence at http://www.biomedcentral.com/about/license which is similar to the 'Creative Commons Attribution Licence'. In brief you may : copy, distribute, and display the work; make derivative works; or make commercial use of the work - under the following conditions: the original author must be given credit; for any reuse or distribution, it must be made clear to others what the license terms of this work are
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