Working group Prof. Dr. med. Barbara C. KahlCystic fibrosis is one of the most prevalent hereditary diseases in the Caucasian population. Cystic fibrosis (CF) patients suffer from recurrent and chronic bacterial airway infections, which eventually lead to lung insufficiency and preterm death. Staphylococcus aureus is one of the first pathogens, which can be isolated from the airways of CF patients and in most patients, the same clone persists for many years. The main focus of our research group is the persistent S. aureus colonization/infection in the airways of CF patients. We are studying the mechanisms of adaptation and microevolution during long-term persistence of S. aureus in the airways of CF patients. Therefore, we characterize virulence regulators and genes in clinical S. aureus isolates including small colony variants (SCVs). Furthermore, we investigate the epidemiology and colonization dynamics of S. aureus in CF patients, the adaptive processes of S. aureus during persistence, which include characterization of virulence patterns in in vitro cell culture models.
Group leader Prof. Dr. med. Barbara C. Kahl
Postdocs Dr. rer. nat. Cathrin Baum
Dr. rer. nat. Nina Hirschhausen
Dr. rer. nat. Claudia Neumann
PhD student Desirée Block (B.Sc., M.Sc.)
Cand. B.Sc. Barbara Müther
Cand. med. Johannes Birtel
Technicians Susanne Deiwick
Students Lena Raidt (SHK)
Felix Stader (SHK)
Projects1. Infection genomics: Host-pathogen interactions: Effects of secreted proteins of Staphylococcus aureus on cells and components of the immune system.
(B. Kahl, M. Hussain, G. Peters, funded by BMBF)Duration: 0 1.09.2010 - 30.08.2013Staphylococcus aureus pathogenicity is determined by the expression of a large set of virulence factors. Extracellular proteins constitute a reservoir of virulence factors. The function of at least 70% of the extracellular proteins encoded by S. aureus is not yet clear and a possible role in virulence has to be elucidated. The present project focuses on the characterization of these particular proteins. Mutants of the corresponding genes, recombinant proteins and corresponding polyclonal antibodies will be generated and used for functional characterization. The functional analysis includes binding to cells of the immune system and non-professional phagocytes (epithelial and endothelial cells), the response of these cells to the proteins, the humoral and cellular immune response, interaction with components of the innate immune system and virulence in animal models. With this, we want to gain new insights into immune evasion mechanisms of S. aureus. Moreover, we will define potential candidates for the development of new vaccines aiming at mitigating or preventing S. aureus infections. Finally, a protein array will be developed which can be used for detection of early S. aureus infections. 2. Pathophysiology of staphylococci in the post-genomic era: Staphylococcus aureus in the airways of cystic fibrosis patients, a human model for long-term adaptive interaction
(A. Mellmann, B. Kahl, funded by Transregional Collaborative Research Centre 34) Duration: 01.07.2010 – 30.06.2014 The lungs of cystic fibrosis (CF) patients are often infected by a predominant Staphylocccus aureus clone for many years. During persistent infection, S. aureus needs to adapt to the hostile environment of the lungs. In this project, we study adaptive mechanisms of S. aureus isolates recovered from the airways of individual CF patients several years apart using different approaches: comparison of proteome and transcriptome, assessment of the role of small non-coding RNAs, and investigation of the virulence potential of isolates. On the host site, the immune proteome of CF patients in response to chronic S. aureus infection will be analyzed. 3. In vivo evolution of Staphylococcus aureus during chronic airway infection of cystic fibrosis (CF) patients.
(B. Kahl, funded by the Interdisciplinary Clinical Research Center (IZKF), Münster) Duration 01.01.2009 – 31.12.2011 One of the first pathogens that colonizes and infects the respiratory tract of CF patients is Staphylococcus aureus. The certified CF center in the Pediatric Department of the Universtity Clinics in Münster cares for more than 80 CF patients. Within the last years we were able to show that S. aureus colonization and infection in CF patients is chronic, persists for many years and in most patients is caused by a predominant S. aureus clone. We are interested whether long-time persistence of S. aureus in the CF lung accompanies genomic adaptations of strains to this environment. For that, we access a unique collection of S. aureus airway isolates from CF patients who have been persistently infected/colonized. Early and late S. aureus isolates of 3 patients recovered 12 years apart, were chosen for whole genome sequencing. Clonal identity of the respective strain pairs was ensured by MLST, PFGE, agr-, and spa-typing prior to WGS. The genome sequences of early and late isolates were compared to detect SNPs (short nucleotide polymorphism), insertions, and deletions. Sanger resequencing will be performed to confirm genomic alterations and a potential impact of detected mutations on protein composition and functionality. In parallel, virulence of the strain pairs will be compared focusing on biofilm formation, adhesive and invasive properties, the capacity to induce apoptosis and host inflammatory response, as well as the virulence potential in an in vivo murine chronic pneumonia model. 4. Dissection of Staphylococcus aureus infection from colonization in cystic fibrosis (CF) patients, a non-interventional, prospective, longitudinal multicenter study
(B. Kahl, H. Wittkowski, funded by Mukoviszidose e.V.; ClinicalTrials.gov Identifier: NCT00669760) Duration: 01.01.2008 – 31.12.2011S. aureus is not only one of the first pathogens infecting the airways of CF patients, but also a highly prevalent microorganism (>60% of all CF patients; European and American CF registries), which often persists for several years in the respiratory tract of CF patients. The purpose of this study is to dissect infection by S. aureus from colonization. Therefore, the following non-interventional, prospective, longitudinal multicenter study will be conducted to verify the following hypothesis: CF patients with high bacterial loads are more likely to be infected by S. aureus than patients with low bacterial loads. The primary endpoint is the bacterial load of sputum and throat cultures. Secondary endpoints are: Nasal carriage, molecular analysis of S. aureus (monoclonal/polyclonal); Serum: S. aureus-specific antibodies, S100A12, IL-8, TNF-a; Sputum: S100A12, IL-8, myeloperoxidase; S. aureus therapy regimens; Lung function analysis: FEV1, DFVC, DMEF25; BMI development Inclusion criteria are S. aureus cultures for more than 6 months within the last year, children (>6 years) and patients, who are able to perform lung function tests. Exclusion criteria are P. aeruginosa and/or B. cepacia cultures for more than 6 months within the last year before recruitment or during the study period. In addition to microbiological investigations and clinical laboratory tests, the actual clinical situation will be evaluated and reported during the study period. The results of this observational study will be used to carefully plan a clinical interventional study. Furthermore, it might be possible to characterize a subpopulation of patients, which is at particular risk for S. aureus infections.
LiteraturAcapsular clinical Staphylococcus aureus isolates lack agr function
Fischer J, Lee JC., Peters G, Kahl BC.
Clin Microbiol Infect. 2013 Oct 17. doi: 10.1111/1469-0691.12429. [Epub ahead of print] Extended Staphylococcus aureus persistence in cystic fibrosis is associated with bacterial adaptation.
Hirschhausen N, Block D, Bianconi I, Bragonzi A, Birtel J, Lee JC, Dübbers A, Küster P, Kahl J, Peters G, Kahl BC.
Int J Med Microbiol. 2013 Dec;303(8):685-92 Intravenous antibiotics given for 2 weeks do not eradicate persistent Staphylococcus aureus clones in cystic fibrosis patients.
C. Andersen, B.C. Kahl, H.V. Olesen, S. Jensen-Fangel, N. Nerskov-Lauritsen
Clin Microbiol Infect. 2013 Sep 17. DOI: 10.1111/1469-0691.12406 Predictive diagnostics for Escherichia coli infections based on the clonal association of antimicrobial resistance and clinical outcome.
Tchesnokova V, Billig M, Chattopadhyay S, Linardopoulou E, Aprikian P, Roberts PL, Skrivankova V, Johnston B, Gileva A, Igusheva I, Toland A, Riddell K, Rogers P, Qin X, Butler-Wu S, Cookson BT, Fang FC, Kahl B, Price LB, Weissman SJ, Limaye A, Scholes D, Johnson JR, Sokurenko EV.
J Clin Microbiol. 2013 Sep;51(9):2991-9. doi: 10.1128/JCM.00984-13. Epub 2013 Jul 10 Small colony variants (SCVs) of Staphylococcus aureus - A bacterial survival strategy.
Infect Genet Evol. 2013 May 27. pii: S1567-1348(13)00205-0. doi: 10.1016/j.meegid.2013.05.016. [Epub ahead of print]1. Kahl BC Impact of Staphylococcus aureus on the pathogenesis of chronic cystic fibrosis lung disease. (2010) Int J Med Microbiol 300:514-519. 2. Loffler B, HussainM, Grundmeier M, Bruck M, Holzinger D, Varga G, Roth J, Kahl BC, Proctor RA, and Peters G (2010) Staphylococcus aureus Panton-Valentine leukocidin is a very potent cytotoxic factor for human neutrophils. PLoS Pathog 6:e1000715.3. Baum C, Haslinger-Löffler B, Westh H, Boye K, Peters G, Neumann C, Kahl BC (2009) Non-spa typable clinical Staphylococcus aureus strains are natural occuring protein A mutants. J Clin Microbiol 47:3624-3629.4. Wellinghausen N, Chatterjee I, Berger A, Niederfuehr A, Proctor RA, Kahl BC (2009) Characterization of clinical Enterococcus faecalis Small Colony Variants. J Clin Microbiol 2009, 47:2802-2811. 5. Mainz JG, Naehrlich L, Schien M, Käding M, Schiller I, Mayr S, Schneider G, Wiehlmann L, Cramer N,Pfister W, Kahl BC, Beck JF, Tümmler B (2009) Concordant genotype of upper and lower airways P.aeruginosa and S.aureus isolates in cystic fibrosis. Thorax 64:535-540.
6. Eickel V, Kahl B, Reinisch B, Dübbers A, Küster P, Brandt C, Spellerberg B (2009) Emergence of respiratory Streptococcus agalactiae isolates in cystic fibrosis patients. PLoS ONE 4:e4650. doi:101371/journal.pone.0004650.7. Besier S, Zander J, Kahl BC, Kraizy P, Brade V, Wichelhaus TA. The thymidine-dependent small-colony variant phenotype is associated with hypermutability and antibiotic resistance in clinical Staphylococcus aureus isolates. Antimicrob Agents Chemother 2008, 52:2183-9.8. Chatterjee I, Kriegeskorte A,Fischer A, Deiwick S, Theimann N, Proctor RA, Peters G, Herrmann M, Kahl BC (2008). In vivo mutations of thymidylate synthase (thyA) are responsible for thymidine-dependency in clinical small colony variants (TD- SCVs) of Staphylococcus aureus. J Bacteriol, 190:834-42. 9. Kahl BC, Peters G (2007) Microbiology. Mayhem in the lung. Science 315: 1082-1083. 10. Hörnig-Franz I, Kahl BC, Tebbe W, Kersting C, Bürger H, Nolte K, Becker K, Bulla N, Debus O, Rabe H, Harms E (2007) Pneumonie mit Staphylococcus aureus (PVL-Gen positiv). Letal verlaufende nekrotisierende Pneumonie bei einem 12-jährigen immunkompetenten Mädchen. Monatsschr Kinderheilkd 155: S10-5. 11. Chatterjee I, Herrmann M, Proctor RA, Peters G, Kahl BC (2007) Enhanced post-stationary phase survival of a clinical thymidine-dependent small colony variant of Staphylococcus aureus results from lack of functional TCA cycle. J Bacteriol 189: 2936-2940. 12. Ridder-Schaphorn S, Ratjen F,Dübbers A, Häberle J, Falk S, Küster P, Schuster A, Mellies U, Löwe B, Reintjes R, Peters G, Kahl BC (2007) Nasal Staphylococcus aureus carriage is nota risk factor for lower airway infection in young cystic fibrosis patients. J Clin Microbiol 45: 2979-84. 13. Goerke C, Gressinger M, Endler K, Breitkopf C, Wardecki K, Stern M, Wolz C,Kahl BC (2007) High phenotypic diversity in infecting but not in colonizing Staphylococcus aureus populations. Environ Microbiol 9:3134-3142.14. Proctor RA, von Eiff C, Kahl BC, Becker K, McNamara P, Herrmann M, Peters G (2006) Small colony variants: a pathogenic form of bacteria that facilitates persistent and recurrent infections. Nat Rev Microbiol 4: 295-305.15. Kahl BC, Mellmann A, Deiwick S, Peters G, Harmsen D (2005) Variation of the polymorphic region X of the protein A gene during persistent airway infection of cystic fibrosis patients reflects two independent mechanisms of genetic change in Staphylococcus aureus. J Clin Microbiol 43: 502-505. 16. Kipp F, Kahl BC, Becker K, Baron EJ, Proctor RA, Peters G, von Eiff C (2005) Evaluation of two chromogenic agar media for recovery and identification of Staphylococcus aureus small colony variants. J Clin Microbiol 43: 1956-1959.17. Haslinger-Löffler B, Kahl BC, Strangfeld K, Grundmeier M, Wagner B, Fischer U, Cheung AL, Peters G, Schulze-Osthoff K, Sinha B (2005) Multiple virulence factors are required for Staphylococcus aureus-induced apoptosis in endothelial cells. 7: 1087-1097.
18. Kahl BC, Belling G, Becker P, Chatterjee I, Wardecki K, Hilgert K, Cheung AL, Peters G, Herrmann M (2005) Thymidine-dependent Staphylococcus aureus small colony variants (SCV) are associated with extensive alterations in regulator and virulence gene expression profiles. Infect Immun 73: 4119-4126.19. von Eiff C, Kahl BC, Kipp F, Herrmann M, Peters G, Becker K (2004) Staphylococcus aureus „Small Colony Variants“- das „zweite“ Gesicht eines klinisch wichtigen Erregers. Hyg Mikrobiol 2: 25-29. 20. Goerke C, Matias y Papenberg S, Dasbach S, Dietz K, Ziebach R, Kahl BC, Wolz C (2004) Increased frequency of genomic alterations in Staphylococcus aureus during chronic infection is in part due to phage mobilization. J Infect Dis 189: 724-734.21. Kahl BC, Belling G, Reichelt R, Herrmann M, Proctor RA, Peters G (2003) Thymidine-dependent small colony variants of Staphylococcus aureus exhibit gross morphological and ultrastructural changes consistent with impaired cell separation. J Clin Microbiol 41: 410-413.22. Kahl BC, Duebbers A, Lubritz G, Haeberle J,Koch HG, Ritzerfeld B, Reilly M, Harms E, Proctor RA, HerrmannM, Peters M (2003) Population dynamics and persistence of Staphylococcus aureus in cystic fibrosis patients. J Clin Microbiol 41: 4424- 4427.23. Kahl BC, Becker K, Friedrich AW, Clasen J, Sinha B, von Eiff C. Peters G (2003) Agr- dependent bacterial interference has no impact on long-term colonization of Staphylococcus aureus during persistent airway infection of cystic fibrosis patients. J Clin Microbiol 41: 5199-5201. 24. Sinha B, Hussain M, Kahl BC, Heilmann C, Peters G, Herrmann M (2003) Adhärenz und zelluläre Invasion von Staphylococcus aureus. Hyg Mikrobiol 2: 80-84. 25. Proctor RA, Dalal SC, Kahl B, Brar D, Peters G, Nichols WW. Two diarylurea electron transport inhibitors reduce Staphylococcus aureus hemolytic activity and protect cultured endothelial cells from lysis. Antimicrob Agents Chemotherap, 2002, 46:2333-2336. 26. Kahl BC, Goulian M, van Wamel W, Herrmann M, Simon SM, Kaplan G, Peters G, Cheung AL. Staphylococcus aureus RN6390 replicates and induces apoptosis in a pulmonary epithelial cell line. Infect Immun 2000, 68:5385-5392.27. von Eiff C, Vaudaux P, Kahl BC, Lew D, Schmidt A, Peters G, Proctor RA. Blood stream infections caused by small colony variants of coagulase-negative staphylococci following pacemaker implantation. Clin Infect Dis 1999, 29:932-934. 28. Kahl B, Herrmann M, Schulze Everding A, Koch HG, Becker K, Harms E, Proctor RA, Peters G (1998). Persistent Infection with small colony variant strains of Staphylococcus aureus in patients with cystic fibrosis.J Infect Dis; 177:1023-9.