Monday, January 14, 2013

LigB of Leptospira interrogans: Avoiding or exploiting complement?

LigB has turned out to be a versatile surface protein for Leptospira interrogans.  The protein is one of several that the spirochete uses to stick to the extracellular matrix, a critical step in colonizing host tissues.  In addition, LigB's ability to bind fibrinogen may help L. interrogans spread within the host by slowing clot formation.  According to separate studies from the U.S. and Brazil published last year, LigB also helps L. interrogans fend off attack by the host complement system.

What's the evidence that LigB protects Leptospira from complement?  L. interrogans, like many pathogens, survives even when complement is present.  Its resistance to complement is reflected by its ability to survive in human serum, which contains all the complement components necessary to assemble the deadly membrane attack complex within exposed microbial membranes.  On the other hand, the nonpathogen Leptospira biflexa, which lacks the ligB gene, is rapidly killed by human serum.  The U.S. study demonstrated that transformation of L. biflexa with a plasmid expressing LigB allowed the nonpathogen to survive in human serum diluted to 5%, a concentration that easily killed L. biflexa harboring a similar plasmid lacking ligB.

How exactly does LigB protect Leptospira from the onslaught of complement?  A defensive strategy deployed by many pathogens is to seize host complement regulators, which are present to prevent complement activation on host cells.  The Brazilian study showed that LigB grabs several complement regulators that diminish the levels of the key complement proteins C3b and C4b on the bacterial surface.  Two of these regulators are the factor H protein and the C4-binding protein (C4BP).  Both complement regulators break apart the C3 convertases, the enzyme complexes that generate C3b from C3.  Factor H and C4BP also serve as cofactors for the protease factor I, which cleaves C3b and C4b into smaller pieces to prevent their assembly into the C5 convertase.  The C5 convertase is what triggers assembly of the membrane attack complex.  As expected, the investigators found that the breakdown of C3b and C4b by factor I in the presence of its complement regulators was accelerated when LigB was present.

LigB is not the only Leptospira protein that captures complement regulators.  The proteins LenA (originally called LfhA) and LcpA bind factor H and C4BP, respectively.  The importance of factor H in protecting L. interrogans can be seen in the bar graph below.  Survival of L. interrogans was poor in serum lacking factor H.  Addition of factor H to the serum up to the concentration found in blood (500 μg/ml) enhanced survival of the spirochete.

Figure 1 from Castiblanco-Valencia et al. (2012).  Survival of L. interrogans in factor H-depleted serum with 500 μg/ml factor H (FH) is set to 100%.

Surprisingly, the U.S. study revealed that a small segment of LigB grabbed C3b and C4b.  Why would L. interrogans risk capturing complement proteins while simultaneously collecting complement regulators that inactivate those same proteins?  Other pathogens do just fine capturing complement regulators without actively grabbing complement components.  It's possible that C3b and C4b are inactivated more effectively by the complement regulators when all components are bound to LigB.

There's another possibility that should be considered.  What went unmentioned in both papers is that C3b and its cleavage products, which may remain attached to the bacterial surface, are opsonins recognized by phagocytes aiming to grab and engulf microbial intruders.  However, several intracellular pathogens use complement receptors as an entry point to invade macrophages.  L. interrogans has been shown to survive within cultured macrophages and even remained intact within macrophages in a zebrafish model, as I've explained in another post.  Is it possible that L. interrogans uses C3b to grab and invade macrophages?

Featured papers

Castiblanco-Valencia MM, Fraga TR, da Silva LB, Monaris D, Abreu PAE, Strobel S, Jozsi M, Isaac L, & Barbosa AS (2012). Leptospiral immunoglobulin-like proteins interact with human complement regulators factor H, FHL-1, FHR-1, and C4BP. Journal of Infectious Diseases, 205 (6), 995-1004 DOI: 10.1093/infdis/jir875

Choy H (2012). Multiple activities of LigB potentiate virulence of Leptospira interrogans: Inhibition of alternative and classical pathways of complement. PLoS ONE, 7 (7) DOI: 10.1371/journal.pone.0041566

Other helpful papers

Verma A, Hellwage J, Artiushin S, Zipfel PF, Kraiczy P, Timoney JF, and Stevenson B (March 2006).  LfhA, a novel factor H-binding protein of Leptospira interrogansInfection and Immunity 74(5):2659-2666.  Link

Barbosa AS, Monaris D, Silva LB, Morais ZM,  Vasconcellos SA,  Cianciarullo AM, Isaac L, and Abreu PAE (July 2010).  Functional characterization of LcpA, a surface-exposed protein of Leptospira spp. that binds the human complement regulator C4BP.  Infection and Immunity 78(7):3207-3216.  DOI: 10.1128/IAI.00279-1010.1128/IAI.00279-10

Related posts

Thursday, January 3, 2013

Spirochete research: 2012 in review

Here are some of my favorite spirochete papers from 2012.  Direct access to all research articles (some behind a paywall) is provided via the DOI links.  Where present, the links above the citations lead to my blog posts about the studies.


Two distinct regions of the Borrelia burgdorferi BBK32 lipoprotein sequentially mediate binding to the vessel wall in vivo during escape of the spirochete from the bloodstream.
  • Moriarty TJ, Shi , Lin Y-P, Ebady R, Zhou H, Odisho T, Hardy P-O, Salman-Dilgimen A, Wu J, Weening EH, Skare JT, Kubes P, Leong J, and Chaconas G (December 2012).  Vascular binding of a pathogen under shear force through mechanistically distinct sequential interactions with host macromolecules.  Molecular Microbiology 86(5):1116-1131.  DOI: 10.1111/mmi.12045

The Leptospira interrogans LigB protein protects the spirochete from complement by capturing complement regulatory proteins.
  • Castiblanco-Valencia MM, Fraga TR, da Silva LB, Monaris D, Abreu PAE, Strobel S, Jozsi M, Isaac L, and Barbosa AS (March 15, 2012).  Leptospiral immunoglobulin-like proteins interact with human complement regulators factor H, FHL-1, FHR-1, and C4BP.  The Journal of Infectious Diseases 205(6):995-1004.  DOI: 10.1093/infdis/jir875
  • Choy HA (July 2012).  Multiple activities of LigB potentiate virulence of Leptospira interrogans: inhibition of alternative and classical pathways of complement.  PLoS One 7(7):e41566. DOI: 10.1371/journal.pone.0041566

A B. burgdorferi lipase with hemolytic activity in vitro:
  • Shaw DK, Hyde JA, and Skare JT (January 2012).  The BB0646 protein demonstrates lipase and haemolytic activity associated with Borrelia burgdorferi, the aetiological agent of Lyme disease.  Molecular Microbiology 83(2):319-334.  DOI: 10.1111/j.1365-2958.2011.07932.x


Borrelia burgdorferi needs the alternative sigma factor RpoS to flee from the tick's midgut
  • Dunham-Ems SM, Caimano MJ, Eggers CH, and Radolf JD (February 2012).  Borrelia burgdorferi requires the alternative sigma factor RpoS for Dissemination within the vector during tick-to-mammal transmission.  PLoS Pathogens 8(2):e1002532.  DOI: 10.1371/journal.ppat.1002532


Video microscopy of B. burgdorferi swimming around in gelatin and mouse tissue:
  • Harman MW, Dunham-Ems SM, Caimano MJ, Belperron AA, Bockenstedt LK, Fu HC, Radolf JD, and Wolgemuth CW (February 21, 2012).  The heterogeneous motility of the Lyme disease spirochete in gelatin mimics dissemination through tissue.  Proceedings of the National Academy of Sciences USA 109(8):3059-3064.  DOI: 10.1073/pnas.1114362109

L. interrogans sheath protein homologs that are not needed for flagellar sheath formation:
  • Lambert A, Picardeau M, Haake DA, Sermswan RW, Srikram A, Adler B, and Murray GA (June 2012).  FlaA proteins in Leptospira interrogans are essential for motility and virulence but are not required for formation of the flagellum sheath.  Infection and Immunity 80(6):2019-2025.  DOI: 10.1128/IAI.00131-12


A close look at the ultrastructure of Leptospira without the artifacts generated by conventional electron microscopy:
  • Raddi G, Morado DR, Yan J, Haake DA, Yang XF, and Liu J (March 2012).  Three-dimensional structures of pathogenic and saprophytic Leptospira species revealed by cryo-electron tomography.  Journal of Bacteriology 194(6):1299-1306.  DOI: 10.1128/JB.06474-11


B. burgdorferi BicA, a protein that protects the spirochete from the toxic effects of copper and iron:
  • Wang P, Lutton A, Olesik J, Vali H, and Li X (December 2012).  A novel iron- and copper-binding protein in the Lyme disease spirochaete.  Molecular Microbiology 86(6):1441-1451.  DOI: 10.1111/mmi.12068


Inflammatory spirochete debris left behind following antibiotic treatment for Lyme disease
  • Bockenstedt LK, Gonzalez DG, Haberman AM, and Belperron AA (July 2, 2012).  Spirochete antigens persist near cartilage after murine Lyme borreliosis therapy.  The Journal of Clinical Investigation 122(7):2652-2660.  DOI: 10.1172/JCI58813

A critical analysis of a study that demonstrated persistence of B. burgdorferi in infected rhesus monkeys that were treated with antibiotics:
  • Wormser GP, Baker PJ, O'Connell S, Pachner AR, Schwartz I, and Shapiro ED (July 2012).  Critical analysis of treatment trials of rhesus macaques infected with Borrelia burgdorferi reveals important flaws in experimental design.  Vector-borne and Zoonotic Diseases 12(7):535-538.  DOI: 10.1089/vbz.2012.1012
  • Embers ME, Barthold SW, Borda JT, Bowers L, Doyle L, Hodzic E, Jacobs MB, Hasenkampf NR, Martin DS, Narasimhan S, Phillippi-Falkenstein KM, Purcell JE, Ratterree MS, and Philipp MT (January 2012).  Persistence of Borrelia burgdorferi in rhesus macaques following antibiotic treatment of disseminated infection.  PLoS One 7(1):e29914.  DOI: 10.1371/journal.pone.0029914

A tale of two more studies: topical antibiotics applied to tick bites to prevent Lyme disease
  • Wormser GP, Daniels TJ, Bittker S, Cooper D, Wang G, and Pavia CS (March 15, 2012).  Failure of topical antibiotics to prevent disseminated Borrelia burgdorferi infection following a tick bite in C3H/HeJ mice.  The Journal of Infectious Diseases 205(6):991-994.  DOI: 10.1093/infdis/jir382


Not so golden?  Microscopic agglutination test for diagnosis of leptospirosis
  • Limmathurotsakul D, Turner EL, Wuthiekanun V, Thaipadungpanit J, Suputtamongkol Y, Chierakul W, Smythe LD, Day NPJ, Cooper B, and Peacock SJ (August 1, 2012).  Fool’s gold: Why imperfect reference tests are undermining the evaluation of novel diagnostics: A reevaluation of 5 diagnostic tests for leptospirosis.  Clinical Infectious Diseases 55(3):322-331.  DOI: 10.1093/cid/cis403


Do nonspiral spirochetes help clean our environment?
  • Caro-Quintero A, Ritalahti KM, Cusick KD, Loffler FE, and Konstandtinidis KT (May/June 2012).  The chimeric genome of Sphaerochaeta: Nonspiral spirochetes that break with the prevalent dogma in spirochete biology.  mBio 3(3):e00025-12.  DOI: 10.1128/mBio.00025-12
  • Ritalahti KM, Justicia-Leon SD, Cusick KD, Ramos-Hernandez N, Rubin M, Dornbush J, and Loffler FE (January 2012).  Sphaerochaeta globosa gen. nov., sp. nov. and Sphaerochaeta pleomorpha sp. nov., free-living, spherical spirochetes.  International Journal of Systematic and Evolutionary Microbiology 62(Pt 1):210-216.  DOI: 10.1099/ijs.0.023986-0



Biofilms of the Lyme disease spirochete
  • Sapi E, Bastian SL, Mpoy CM, Scott S, Rattelle A, Pabbati N, Poruri A, Burugu D, Theophilus PAS, Pham TV, Data A, Dhaliwal NK, MacDonald A, Rossi MJ, Sinha SK, and Luecke DF (October 2012).  Characterization of biofilm formation by Borrelia burgdorferi in vitroPLoS One 7(10):e48277.  DOI: 10.1371/journal.pone.0048277



Looking for the syphilis spirochete in ancient bones
  • Montiel R, Solorzano E, Diaz N,  Alvarez-Sandoval BA, Gonzalez-Ruiz M, Canadas MP, Simoes N, Isidro A, and Malgosa A (May 2012).  Neonate human remains: A window of opportunity to the molecular analysis of syphilis.  PLoS One 7(5):e36371.  DOI: 10.1371/journal.pone.0036371

Presenting flawed studies directly to the public to bypass the scientific peer-review process:
  • Armelagos GJ, Zuckerman MK, and Harper KN (March 2012).  The science behind pre-Columbian evidence of syphilis in Europe: research by documentary.  Evolutionary Anthropology 21(2):50-57.  DOI: 10.1002/evan.20340


Here are two excellent review articles that appeared during the past year:
  • Radolf JD, Caimano MJ, Stevenson B, and Hu LT (February 2012).  Of ticks, mice and men: understanding the dual-host lifestyle of Lyme disease spirochaetes.  Nature Reviews Microbiology 10(2):87-99.  DOI: 10.1038/nrmicro2714
  • Charon NW, Cockburn A, Li C, Liu J, Miller KA, Miller MR, Motaleb MA, and Wolgemuth CW (2012).  The unique paradigm of spirochete motility and chemotaxis.  Annual Reviews of Microbiology 66:349-370.  DOI: 10.1146/annurev-micro-092611-150145