Human-pathogenic Yersinia species Prof. James B. Bliska

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1 Human-Pathogenic Yersinia Species, Ph.D. Department of Molecular Genetics and Microbiology Stony Brook University 1 Scientific classification 2 Common features Gram-negative rod-shaped bacteria Facultative intracellular pathogens Adept at circumventing innate immunity (especially Y. pestis) Single circular chromosome of approximately 4,600,000 nucleotides 12 genomes presently sequenced: Y. enterocolitica (1) Y. pestis (7) Y. pseudotuberculosis (4) Circular plasmid of approximately 70,000 nucleotides pyv or pcd1 Encodes a type III secretion system 3 1

2 Classification of strains Y. enterocolitica 6 biogroups (1A, 1B, 2, 3, 4 and 5) Y. pseudotuberculosis 21 serogroups based upon variations in O antigen of LPS Y. pestis 5 groups or biovars (Orientalis, Mediaevalis, Antiqua, Microtus, Pestoides) 4 Carniel, E. Microbes and Infection, 3, 2001, Overview of Yersinia infections Ingested or M cell in small intestine e Macrophage Rare event Brendan Wren,

3 Intestinal Yersinia infections Septicemia Enterocolitis Mesenteric lymphadenitis Adapted from Salyers and Whitt, Bacterial Pathogenesis, ASM Press 7 Intestinal Yersinia infections: transmission Linked to ingestion of contaminated foods, water, and milk; Contaminated foods include pork, tofu, and poultry May occur directly from dogs, cats, and swine No role for airborne route of infection or for insect vectors in the transmission of disease Reports of person-to-person spread are conflicting and generally are not observed in large outbreaks 8 Intestinal infections: outcomes Diarrhea Most common clinical manifestation The usual presentation is characterized by diarrhea, low-grade fever, and abdominal pain lasting 1-3 weeks; Diarrhea may be bloody in severe cases Vomiting Present in approximately 40% of cases Mesenteric adenitis, mesenteric ileitis, or acute pseudoappendicitis These manifestations are characterized by fever, abdominal pain, tenderness of the right lower quadrant, and leukocytosis 9 3

4 Intestinal infections: outcomes cont. Reactive arthritis Most commonly reported in Scandinavia, polyarticular arthritis can occur after infection with Y. enterocolitica The onset of joint symptoms typically occurs 1-2 weeks after gastrointestinal illness and occurs in approximately 2% of patients Erythemanodosum manifests as painful raised red or purple lesions, primarily on the lower extremities Septicemia Most commonly in patients who have predisposing conditions; Iron overload 10 Plague: overview About total cases/year in U.S Mainly southwestern Unites States Bubonic most common form Only 1-2 cases per year of pneumonic form 11 Transmission modes of Y. pestis Pneumonic plague Bubonic plague Adapted from Salyers and Whitt, Bacterial Pathogenesis, ASM Press 12 4

5 Unique features of Y. pestis Lacks O antigen Contains 2 additional plasmids required for unique aspects of disease pmt1 Phospholipase: required for survival in flea mid-gut F1 protein: promotes transmission following flea bite ppcp1 Pla: plasminogen activator and adhesin/invasin Required for dissemination to lymph nodes after flea bite Promotes bacterial growth in lungs during pneumonic plague 13 Unique features of Y. pestis cont. Pigmentation (pgm) segment Genes within (hms) code for extracellular matrix important for biofilm formation in flea Pgm segment encoded in Y. pseudotuberculosis but hms genes not highly expressed Two genes inactivated in Y. pestis RcsA, a negative regulator of biofilms The gene for a glycosylhydrolase (NghA) that cleaves beta-linked N-acetylglucosamine residues and reduces biofilm formation 14 Plague: bubonic Inguinal, axillary, or cervical lymph nodes most common 80% can become bacteremic 60% mortality if untreated 15 5

6 Plague: pneumonic From aerosol or septicemic spread to lungs Person-to-person transmission by respiratory droplet 100% mortality if untreated 16 Plague: pneumonic cont. Incubation: 1-3 days Sudden onset headache, malaise, fever, myalgia, cough Pneumonia progresses rapidly to dyspnea, cyanosis, hemoptysis Death from respiratory collapse/sepsis Pneumonic infiltrate of pneumonic plague 17 Attributes of a successful bacterial pathogen 1. Adhere to/invade into host cells 2. Regulate gene expression in response to environmental cues 3. Avoid, subvert or co-opt host immune responses 4. Acquire nutrients to replicate 18 6

7 Adhesion and invasion determinants Pili Psa/Myf pili Afimbrial adhesins Ail Major role in serum resistance Lui et al., Infect. Immun. 74, YadA (2006) Encoded on plasmid; Gene inactivated in Y. pestis Binds ECM proteins Invasin Gene (inva) inactivated in Y. pestis Important role in allowing enteropathogenicyersinia to invade M cells overlying Peyer s patches 19 Mechanism of Yersinia invasion Role in the disease process is to allow bacterial penetration of the gut-associated lymphoid tissue Small intestinal mucosa (ileum) Adapted from Salyers and Whitt, Bacterial Pathogenesis, ASM Press 20 Invasin 986 amino acid outer-membrane protein D4-D5domains sbind with high affinity to several β 1 integrins (α 3, α 4, α 5, α 6 and α V ) Hamburger Z. A. et al., Science, 286, 291-5, (1999) 21 7

8 Mechanism of invasin-promoted uptake Integrin clustering activates signaling pathway Actin rearrangements lead to zippering of membrane around bacterium 22 Systemic infection in mice may bypass lymph node colonization Septicemia Enterocolitis Adapted from Salyers and Whitt, Bacterial Pathogenesis, ASM Press Mesenteric lymphadenitis 23 Regulate gene expression Transcription factors RovA, member of the MarR/SlyA family of winged-helix transcription factors Regulates expression of invasin and Psa Important for virulence in intestinal and bubonic Yersinia infections PhoP, two-component regulator Required for survival in macrophages VirF, AraC-like activator of the ysc-yop virulon encoded on pyv/pcd1 24 8

9 Avoid, subvert or co-opt host immune responses After translocation across M cells, the bacteria encounter macrophages or dendritic cells 25 Localization of Y. pseudotuberculosis in rabbit M cell and macrophage 4 hours post infection Fujimura et al. J. Clin. Electron Microscopy Extracellular location of Y. enterocolitica in rabbit Peyer s patches 24 hr post infection Lian et al., J. Med. Microbiol

10 Simplified Yersinia pathogenesis model Resist phagocytosis PMN Extracellular replication Naive Mφ Release? Escape Avoid intracellular killing Apoptosis 28 Summary of vacuole trafficking studies Y. pestis-containing vacuoles acquire markers of late endosomes/lysosomes (between 1.5 to 8 hr post infection) PhoP-regulated genes required for survival in phagosomes Spacious phagosome formation associated with replication at 8 hr post infection Does not require other known virulence factors (pcd1, Pla) 20h post infection in a mouse bone-marrowderived macrophage Grabenstein et al., Infect. Immun Model of YCV trafficking Pinocytosis Phagocytosis Early.. endosome Late endosome ? Lysosomes Spacious phagosome..... ph 7.. Nucleus Golgi Grabenstein et al., Infect. Immun Pujol et al., Infect. Immun

11 Simplified Yersinia pathogenesis model PMN Extracellular replication Naive Mf Release? Escape Avoid intracellular killing Apoptosis 31 pcd1 encodes type III protein secretion system Yops, LcrV 37 C Adapted from: Cornelis, Nature Reviews, 2002 Thanassi and Hultgren, Curr. Opin. Cell. Biol., Yersinia type III secretion system Yersinia 37 C Yersinia YopB/YopD YopH PTPase Effectors Host cell YopM LRR YopE GAP YopJ Acetyl transferase YopO GDI YopT Cys protease 33 11

12 Overview of Yop effectors: antagonize phagocytosis, inhibit cytokine production, induce apoptosis Navarro et al., Actin cytoskeleton rearrangement target Rho GTPases PTPase counteracts phagocytosis Acetyl Transferase inhibits MAPK and NF-κB pathways Viboud and Bliska, 2005 Function unknown 35 The type III injectisome From Cornelis, G.R., Nature Reviews,

13 LcrV Mueller et al., Science Acquire nutrients to replicate Carniel, E. Microbes and Infection, 3, 2001, The Yersinia HPI comprises genes involved in uptake of iron HPI encodes genes involved in the synthesis and transport of the siderophore yersiniabactin The siderophore-fe 3+ complex recognizes a specific bacterial outer membrane receptor; Proteins located in the periplasm and the inner membrane of the cell wall transport the complex into cytosol HPI is found in other enterobacteria such as Escherichia coli, Klebsiella, Citrobacter and Salmonella Miller, M.C. et al., Journal of Inorganic Biochemistry, 100, 2006,

14 Some open questions in Yersinia pathogenesis Other than pmt1 and ppcp1, what are the critical genetic differences between Y. pestis and Y. pseudotuberculosis that account for differences in virulence? How does Yersinia block phagosome acidification How does translocation of type III effector proteins occur? 40 Thank you Members of the Yersinia research community who provided data discussed in this presentation Listeners 41 References pmt1 Phospholipase: Hinnebusch B.J., et al., Science, 269, (2002) F1 protein: Sebbane F., et al., Infect Immun. 77, (2009) Hms genes and biofilm production PMID: PMID: Invasin PMID: Dissemination of Y. pseudotuberculosis from intestine to blood stream in mice PMID: RovA PMID:

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