1. pPst 9.5 kb- smallest plasmid Found only in Y. Pestis
Encodes for plasminogen activator (pla)
Cell surface protease
Has coagulase activity evident only at temperatures below 30°C, which initiates blockage in flea and forms fibrin matrix that anchors bacteria to proventiculus of flea
At 37°C exhibit fibronolytic activities, lyses fibrin at bite site and allows dissemination of bacterium
Definitely plays a role in flea transmission, not sure how important the gene product is in mammalian cells.

2. pPst Other possible functions of Plasminogen activator
Produces excess plasmin that causes ineffective structures between inflammatory cells and fibrin
Reduces chemoattractants at the infection site possibly via inhibition of IL-8.
Encodes bacteriosin pesticin (pst) and its immunity protein (pim)

3. pCD1 68-75 kb (depending on the strain)
Responsible for encoding anti-host genome
Y.Pestis, Y.pseudotuberculosis, and Y. Enterocolitis both possess this plasmid
Encodes the Low Calcium Response System (LCRS)- controlled by Temperature and Calcium concentration
Low Calcium response V antigen-LcrV
Yersinia Outer membrane proteins- Yops
Specific Yop chaperones- Syc
Yop secretion proteins/Type III secretion system- Ysc
This plasmid is largely responsible for encoding an anti-host genome. It is necessary for virulence in Y.Pestis and Y. Psuedotuberculosis. LcrV, Yops…all are factors encoded on this plasmid. Im going to talk about each of these factors and each of their function in detail in the next couple of slides.

4. pCD1 LcrV/V antigen LcrV protein plays a role in regulating the LCRS
It is a protective antigen that is associated with resistance to phagocytosis
May play an immunosuppressive role, by inhibiting cytokine production
Induces IL-10 production by macrophages this downregulates host’s immune response
Mutes inflammatory responses by inhibiting cytokine production

5. pCD1 Yop proteins Altogether there are 29 Yop proteins
Yop effector proteins protect Y.pestis from macrophages by destroying phagocytic signaling capabilities
There are 6 Yops which directly or indirectly cause disease:
Yop E, Yop H, Yop J, Yop O, Yop M, Yop B and D
These yops listed here are termed effectors.
Yop E through depolymerizing actin microfilaments causes the collapse of the cytoskeleton.

6. How Yops Thwart Host Immune System
■ Yop E
■ Indirectly depolymerizes actin microfilaments
■ Yop H
■ a protein tyrosine phosphatase capable of dephosphorylating host proteins such as p125FAK and p130Ca at focal adhesions, inhibiting signal transduction necessary for phagocytosis
■ Yop J
■ Binds to MAP kinase family-downregulating TNF-

7. More Yops Yop O/YpkA Yop M Yop B and D
Ser/Thr kinase that probably interferes with signaling pathways
Yop M
Prevents thrombin platelet aggregation
Mutes inflammatory response by sequestering thrombin
Yop B and D
Translocate effectors across cell membrane

8. Yops Regulation Temperature and Ca2+
When placed in 37°C and in a medium deprived of Ca2+, Y. Pestis ceases growth and expression of Yops is induced.
Yops expression and secretion is induced in Ca2+-containing media by a local signal that occurs at the site of contact between the pathogen and the eukaryotic cell. This leads to the polarized transfer of at least five Yops into the target cell.

9. pCD1 Syc proteins Small Yop binding proteins
Not all Yops are syc associated
Act as chaperones by preventing Yop degradation
Act as “secretion pilots” leading the Yops to their location of secretion

10. pCD1 Ysc/ Type III secretion system
Yops is mediated by Type III secretion system called Ysc
Ysc Type III secretion system consists of:
the core Ysc mechanism for secretion through the two bacterial membranes
a delivery apparatus (YopB, YopD, YopK, LcrV)
control elements (LcrE also called YopN and TyeA at the surface and LcrG in the cytosol)
anti-host effector proteins (YopE, YopH, YopM, YpkA and YopJ).

11. Ysc/Type III secretion system
Model for Ysc-dependent and independent secretion of LcrV. LcrV, YopB, YopD, and possibly other proteins (X) are secreted by contact-activated Ysc secretion channels and assemble into an apparatus that mediates targeting of secreted Yops across the plasma membrane and into the cytoplasm of eukaryotic cells. LcrV shed from these structures diffuses into the surrounding medium. The LcrV-transporting contact-activated translocator (VCAT) can also secrete LcrV from Yersinia. Upon contact with the plasma membrane, LcrV not involved in secretion or targeting of Yops is secreted across the inner membrane (IM), periplasm (PP), and outer membrane (OM) of Y. pestis through the VCAT mechanism composed of non-pCD1-encoded proteins. LcrV secreted in this manner is delivered directly to associated eukaryotic cells. [Return to Article]
J. Bacteriol.J. Virol.Eukaryot. CellMicrobiol. Mol. Bio. ReviewAll ASM Journals

12. Yersinia’s Deadly Kiss

13. All Together Now! Yersiniabaktin=Yersiniabactin (Ybt)
Koagulase and Plasminaktivierung= Plasminogen activator
Pestizin=Pesticin
pTox=pFra
pCPC=pPst
pYV=pCD1
Fra1=F1
Mureintoxin=Murine Toxin
Phagozyte=phagocyte

14. Clinical Aspects Focus on Bubonic Plague
Signs and symptoms
Complications
Differential Diagnosis
Diagnosis
Treatment
Prognosis
Prevention
Vaccine Development

15. Presenting Plague Patients%

16. Signs & Symptoms Within 2-7 days of Infection:
■ Severe malaise or prostration ---75%
■ High fever, headache ---20%-85%
■ Chills %
■ Vomiting ---25%-49%
■ Altered mentation ---26%-38%
■ Abdominal pain %
■ Other: bladder distention, apathy, confusion, fright, anxiety, oliguria,
anuria, tachycardia, hypotension, leukocytosis
Abrupt onset
Presenting symptoms include prostration or severe malaise (75%), headache (20%–85%), vomiting (25%–49%), chills (40%), altered mentation (26%–38%), cough (25%), abdominal pain (18%), and chest pain (13%).
Other manifestations of bubonic plague include bladder distention, apathy, confusion, fright, anxiety, oliguria, and anuria. Tachycardia, hypotension, leukocytosis, and fever are frequently encountered.
Prostration = extreme physical exhaustion/weakness
Septicemia (intermittent at first, rapidly becomes constant)
A femoral bubo (a), the most common site of an erythematous, tender, swollen, lymph node
bubo, the inflammatory swelling of one or more lymph nodes, usually in the groin; the confluent mass of nodes, if untreated, may suppurate and drain pus

17. Signs & Symptoms 6-8 hours following onset of symptoms:
■ Pain/tenderness at regional lymph nodes
enlarge to be called buboes
- extremely painful
- occur in groin*, axilla or cervical areas
- usually occur in 1 region, but multiple can
be seen
- may drain pus if left untreated
■ ulcer or skin lesions at site of flea bite
<10% of cases

18. Complications ■ 2 Septicemic Plague due to hematogenous spread
(23% of patients)
- Purpuric Lesions
- Acral Necrosis
- DIC
disseminated
intravascular
coagulation
- Convulsions
- Shock
Bubonic plague invades bloodstream… patient develops secondary septicemic plague
Sepsis Syndrome with Multi Organ Involvement
DIC: tissue necrosis and bleeding following uncontrolled activation of clotting factors & fibrinolytic enzymes
Death due to endotoxic shock
Purpuric lesions
diffuse, hemorrhagic
changes in skin
(darkened skin changes
="black death”)
Acral necrosis

19. Complications ■ 2 Pneumonic Plague (5-15% of patients)
- begins as interstitial pneumonitis;  bacteria in interstitium
- show signs and symptoms prior to developing advanced
pneumonitis:
High fever
Severe bronchopneumonia
Chest pain
Coughing or spitting up blood
Difficulty breathing
■ Infection of Buboes
■ Meningitis (< 5%)
■ Death

20. Distinguishing 1 from 2
Pneumonic Plague
■ no buboes
■ 1-3 day incubation period
■ infectious pneumonitis from onset of
symptoms
■ sputum production common
■ less severe evidence of disease
in organs other than lungs
■  bacteria in alveoli
■ tracheal & bronchial mucosal hemorrhages
■ can lead to 2 Septicemic Plague
symptoms
Sudden onset of fever, chills, headache, body pains, weakness, chest discomfort & tightness; Coughing with sputum production; Difficulty breathing; Coughing or spitting up blood; GI symptoms; Pharyngitis
- direct inhalation of Y. Pestis

21. Distinguishing 1 from 2
Septicemic Plague
■ no buboes
■ 1-4 day incubation period
■ more GI symptoms
■ can also lead to 2 Pneumonic
Plague (25%)
■ meningitis is 4x more common
symptoms
Fever and chills
Extreme exhaustion
Bleeding disorder
Necrosis of small vessels
Hemorrhagic Skin lesions
(purpuric lesions, acral necrosis)
Gangrene of extremities (nose or digits)
DIC Convulsions
Shock
- direct bloodstream inoculation with Y. Pestis

22. Differential Diagnosis
Bubonic Plague
■ Chancroid
■ Primary genital herpes
■ 1 or 2 syphilis
■ Strangulated inguinal
hernias
■ Streptococcal or
staphylococcal adenitis
■ Tularemia
■ Cat scratch disease
■ Mycobacterial infection
■ Lymphogranuloma
venereum