1. Two Med Students in Search of Answers Tam-Linh Nguyen Elizabeth Leddy
The Plague
Two Med Students in Search of Answers
Tam-Linh Nguyen
Elizabeth Leddy

2. Intro
Patient Info
large swelling of lymph nodes at arm pit, neck and groin (buboes)
high fever
delerium
black splotches on skin
as the disease progressed, the buboes burst

3. Culture/Isolation of Organism
Organism taken from blood or buboes
Media used: blood agar or other enteric media
After 24 hours the colonies were pinpoint
After 48 hours the colonies were mm in diameter, and grey to greyish white, and slightly mucloid
In broth culture (bile or bile salts), clumps adhere to side of tube

4. Tests performed

5. Yersinia Species Three species of Yersinia cause disease in humans
Y. pseudotuberculosis and Y. enterocolitica are enteric food and water borne pathogens
acquired by ingestion of contaminated food
can cross the gastrointestinal mucosa
infection is localized

6. Y. pestis Discovered in 1894 by French bacteriologist Alexandre Yersin
Gram negative rod shaped bacilli
found in low frequency in wild rodent populations, such as Rattus rattus (small and black) and Rattus norreguis (large grey sewer rat)
Oriental rat flea Xenopsylla cheopis is prime transmitter of disease
members.aol.com/landmate/ biological.htm

7. Modes of Transmission Rattus rattus Rattus norvegicus
Xenopsylla cheopis

8. Y. pestis morphology Part of Enterobacteriaceae family
Optimal growth at 28 deg. C
Non-motile, slow in vitro growth
Closely related to E. coli by DNA-DNA hybridization
Unable to survive outside of a host organism

10. A little bit of history
6th Century (Justinian’s plague AD)

11. A little bit of history Black Death (1346 to nearly the 19th century)
this was also known as the “Great Dying” or the “Great Pestilence”
Responsible for the death of 30% of the European population
Third Pandemic (1894)

12. Transmission of Y. pestis
Disease endemic to rat species
Transmitted by fleas
The flea bites the rat, and ingests the organism
The Y. pestis replicates in the digestive tract of the flea
A solid mass forms which obstructs the fleas gut

13. Transmission of Y. pestis
plague/cheob6x4.htm

14. Transmission of Y. pestis
Flea can’t ingest blood
Gets increasingly hungry, therefore increasingly voracious
The flea attacks more rats, which are then infected with the microbe
When the rat population dwindles, the flea moves onto other available hosts, such as humans

15. Human to Human Transmission
Inhalation of infected respiratory droplets
When an infected human coughs, some microbe is released
Airborne pathogen can be inhaled by another human in close proximity
Accounts for ability of such a large outbreak in Europe

16. Stages of Disease
Bacteria travel through the blood to the nearest lymph nodes
In lymph nodes, Y. pestis is ingested by fixed macrophages
Y. pestis is able to grow in inactivated macrophages and replicate
Elicits an inflammatory response (the bubo)

17. Stages of Disease The inflammatory response - “bubo”
2000/09/12/52.asp

18. Stages of Disease
Bacteria from the bubo leak into the blood stream. (septicemic plague)
Lysis of the bacteria releases LPS, which causes septic shock
Eventually bacteria reach the lung, where they parasitize the lung macrophages (pneumonic plague)

19. Stages of Disease
At the pneumonic stage, the bacteria can be spread to others via aerosols. (respiratory droplets)
Direct inhalation at this point of the disease, induces more rapid development (than flea)
At this stage the bacteria has well developed virulence factors needed to colonize the human body

20. Stages of Disease

21. Virulence Mechanism of Y. pestis
Many genes necessary for virulence are found on plasmids
Due to similarity of Y. pestis and Y. pseudotuberculosis, it is possible to eliminate certain genes located on Y. enterocolitica as critical to causing plague

22. Virulence Mechanism of Y. pestis
The three similar Yersinia species have kbp plasmids
These plasmids carry a number of genes related to virulence

23. Virulence Mechanism of Yersinia
These virulence genes produce virulence factors that fall into four general categories:
Adhesion and invasion (YadA)
Execreted antiphagocytic proteins (Yops)
Proteins involved in processing and excreting Yops (Ysc)
Regulatory proteins (Lcr)

24. Virulence Mechanism of Y. pestis
Genes represented are known to be encoded on the 75-kbp plasmid of Yersinia species

25. Virulence Mechanism of Y. pestis
What do you think would happen if Y. pestis was cured of its virulence plasmids?

26. Virulence Mechanism of Y. pestis
What do you think would happen if Y. pestis was cured of its virulence plasmids?
A decrease in ability to colonize in the human body

27. Virulence Mechanism of Y. pestis
What do you think would happen if Y. pestis was cured of its virulence plasmids?
A decrease in ability to colonize in the human body
Disease not able to spread as easily

28. Virulence Mechanism of Y. pestis
Y. pestis has two more plasmids than Y. pseudotuberculosis and Y. enterocolitica
One 110 kbp plasmid
not much is known about the genes on this plasmid
one gene encodes a protein component of an antiphagocytic protein capsule (F1)
another encodes a toxin that kills mice, but has not been shown to be toxic in humans

29. Virulence Mechanism of Y. pestis
Fraction 1 (F1) is a capsule like antigen expressed at 37 deg. C
located on the large plasmid found only in Y. pestis
Involved in the ability of Y. pestis to prevent uptake by macrophages
Does not influence the general ability of phagocytic cells
Expression of F1 reduced the number of bacteria that interacted with macrophages

30. Virulence Mechanism of Y. pestis
The second plasmid (pPCP1) is a 9.5 kbp plasmid that encodes 3 proteins: pesticin, pesticin immunity protein, and plaminogen activator

31. Virulence Mechanism of Y. pestis
Plasminogen Activator (Pla)
multifunction surface protein
this aids in the spread of the disease throughout the human body
may also have a role in the insect mediated transmission of the bacteria

32. Chromosomally encoded virulence factors
Three chromosomally encoded virulence genes (yst, psa, inv)
the first encodes a heat stable enterotoxin
eventually leads to fluid build up in the intestine
the second is the main structural component of the Yersinia fimbriae
the third helps to colonize deep tissues (lymph nodes)

33. Yersinia Pestis Genome
magpie.genome.wisc.edu/browser/ Yersinia_pestis_circle.html

34. Special Traits of Y. pestis
Two extra plasmids account for higher virulence
Plasminogen
Hemin storage
iron reserve
also protect the bacteria from host’s defenses by making it seem more like a host component

35. Treatment and Prevention
Antibiotic Therapy
streptomycin, tetracycline, and chloramphenicol
all inhibit protein synthesis
essential to help the immune system clear the infection
the earlier the treatment is initiated, the better the outcome

36. Treatment and Prevention
Sanitation
keep rodent population down by proper disposal of garbage
eliminate crowded living conditions of substandard housing

37. Treatment and Prevention
Vaccines
effective but protects for less than a year
vaccine consists of whole killed Y. pestis cells
requires a series of injections over a 6 month period
live attenuated vaccines
injection of non-pathogenic mutant, derived from a fully virulent strain
safety questionable

39. Plague and relation to AIDS
CCR5 gene

40. “Ring Around The Rosy…”

41. References
Achtman, Mark, et al. “Yesinia pestis, the cause of plague…”. Proceedings of the National Academy of Sciences. Nov. 23, 1999
Balows, Albert. Ed. In chief Manual of Clinical Microbiology. 5th Edition. American Society for Microbiology. Washington D.C.
Du, Yidong, et al. “Role of fraction 1 antigen of Yersina pestis in inhibtion of phagocytosis.” Infection and Immunity. V70, no3. March 2002
Gerhardt, Philipp, ed in chief Methods for General and Molecular Bacteriology. American Society for Microbiology. Washington D.C.
Lederberg, Joshua, ed in chief. Encylcopedia of Microbiology. 2nd Edition, v3. Assoc. Press, New York
McEvedy, Colin. “The Bubonic Plague.” Scientific American. Feb v. 258 n2 p118

42. References
Revel, Paula A. “Yersinia virulence: more than a plasmid. FEMS Microbiology Letters. V206 no 2. Dec. 2001”
Salyers, Abigail A., and Dixie E. Whitt Bacterial Pathenogeneis, a Molecular Approach. American Society for Microbiology. Washington D.C.
Titball, Richard W, and E. Diane Williamson. “Vaccination against bubonic and pneumonic plague.” Elsevier. April 2001
Wieland, Felix. ed. “The Pla surface protease adhesin of Y. pestis mediates bacterial invasion into human endothelial cells”. FEBS Letters. V504, no.1-2. Aug. 2001