Using genomics to track the dissemination of Yersinia pestis strains

1. Using genomics to track the dissemination of Yersinia pestis strains

2. Transmission cycle of plague A useful diagram illustrating the transmission cycle of the plague by Neil Chamberlain “Transmission Cycles of Plague” is available for educational use from microbelibrary.org

3. Y. pestis and plague • Nearly all plague infections are zoonotic with humans being accidental hosts • Two different transmission cycles of Y. pestis infection: • zoonotic • human to human • Zoonotic (pertaining to animals) plague: transmission due to the bite of infected fleas or contact with infected animals • Zoonotic plague is further divided into: • urban • sylvatic (rural) plague • The second type involves the pneumonic transmission of infection between humans. Information from Neil Chamberlain “Transmission Cycles of Plague” www.microbelibrary.org

4. Sylvatic plague cycle • Infection of wild mammals, (usually rodents), involves the passing of plague from mammal to mammal by fleas (Xenopsylla cheopis) infected with Y. pestis. • Mammal to mammal passage of Y. pestis can occur by direct contact but is less common. • Many of these mammals are relatively resistant to the lethal effects of Y. pestis infection. • Squirrels, rabbits, and field rats can all serve as long-term reservoirs for the plague bacterium (enzoonotic plague). • Other mammals are highly susceptible to the lethal effects of Y. pestis, resulting in rapid spread of plague among the animals with large numbers dying in the community (epizoonotic plague; e.g., prairie dogs). • If humans enter these areas they acquire the infection via flea bites or direct contact with infected animals.

5. Urban plague cycle • Involves the passing of plague from mammal to flea to mammal. • Mammal to mammal passage of Y. pestis can occur by direct contact but is less common. • These mammals, frequently rats, are usually highly susceptible to the lethal effects of Y. pestis. • As a result, large numbers of these animals die (epizoonotic plague). • The infected fleas, lacking their more common rodent hosts, then bite and infect humans. • Direct contact with infected animals can also result in human infection.

6. Vehicle of transmission: Fleas • The flea draws viable Y. pestis organisms into its intestinal tract. These organisms multiply in the flea and block the flea's proventriculus. • Some Y. pestis in the flea are then regurgitated when the flea gets its next blood meal, thus transferring the infection to a new host. • While growing in the flea, Y. pestis loses its antiphagocytic capsular layer. • The male oriental rat flea, Xenopsylla cheopis, is the primary vector of plague in most large plague epidemics in Asia, Africa, and South America. Both male and female fleas can transmit the infection.

7. Human infections • Following flea bites or by direct contact with infected animals; usually result in bubonic plague. • Most of the unencapsulated organisms are phagocytosed (ingested) and killed by polymorphonuclear leukocytes (PMNs or neutrophils) in the human host. • A few bacilli are taken up by tissue macrophages. • The macrophages are unable to kill Y. pestis and provide a protected environment for the organisms to synthesize their capsular and other virulence factors. • The Y. pestis are taken to the draining lymph nodes by the macrophages.

8. Human infection cont. • The encapsulated intracellular organisms then kill the macrophage and are released into the extracellular environment, where they resist phagocytosis by the PMNs. • The Y. pestis multiply rapidly and cause a hemorrhagic inflammatory response making the lymph nodes hot, swollen, tender, and hemorrhagic. • This gives rise to the characteristic black buboes responsible for the name of this disease: bubonic plague.

9. Human infection cont. • In a relatively short period of time the bacteria get out of the swollen lymph nodes and into the bloodstream. • Then they infect the liver, spleen, and lungs. Bacteremia (bloodstream infection; septicemic plague) occurs rapidly if not treated and mortality rates can be as high as 75%. • A severe bacterial pneumonia can develop: pneumonic plague. Patients expel large numbers of viable organisms during coughing fits. • Other humans inhale these viable organisms and develop pneumonia. • The spread of Y. pestis via droplets from human to human is also known as the demic plague cycle. • Spread from person to person can be very rapid and has in the past caused epidemics with mortality rates over 90%, if untreated.

10. Historic 3 pandemics of plague -Pandemic is defined as an epidemic that spreads throughout the human population across a large region such as a continent or worldwide -1st pandemic ~550 A.D. Confined mainly to Africa and some parts of the Middle east -2nd pandemic originated in Central Asia and spread via trading routes into Europe (Killed ~30-60% of European population) -3rd pandemic started in 1850’s in China’s Yunnan province and was confined mainly to Asia

11. Identifying the causative agent of the plague -French researcher Paul-Louis Simond postulated a connection between human and rodent plague and identified the flea as a possible vector -In 1894, in Hong Kong, bacteriologist Alexandre Yersin isolated the responsible bacterium (Yersinia pestis) and determined the common mode of transmission -A short time later, Japanese physician and researcher Shibasaburo Kitasato independently identified the plague bacillus (after mis-identifying the bacterium at an earlier point).

12. As of 2010 there are 7 complete and 14 draft Y. pestis genomes Traditionally the strains are classified as biovars (Antiqua, Mediaevalis, Orientalis, and other) based on the following phenotypic characteristics: -Antiqua = East Africa: (glycerol positive, arabinose positive, and nitrate positive) -Mediaevalis = Central Asia: (glycerol positive, arabinose positive, and nitrate negative) -Orientalis = Central Asia (glycerol negative, arabinose positive, and nitrate positive) -other (ie Microtus, Pestoides) = not consistent for above phenotypes

13. Paleomicrobiology -The prefix paleo comes from the Greek work palaios, meaning “ancient” -Bacterial colonization of dental pulp can occur during bacteremia -Bacteremia (also known as plague septicaemia with Y. pestis) is the presence of bacteria in the blood

14. Extraction of bacterial DNA from dental pulp -Some historians believed that a flu-like virus and not Y. pestis was responsible for the 1st and 2nd pandemics -DNA detected in dental pulp confirm that Y. pestis was the cause -Which biovar(s) are most similar to the Y. pestis strain(s) from the dental pulp from the corpses?

15. 11 categories of virulence factors for Y. pestis -Adhesion -Antigen -Effectors -Insect effectors -Host barriers -Up regulated in human plasma -Iron acquisition -Phage component -Regulator -Secretion -Invasion

16. Known or putative virulence factors for Y. pestis separated into categories (n=148 genes)

17. Use of genomic tools to study Y. pestis Concepts in this module that you will address: #1) Mutations that affect the production of a fully functional gene product that has phenotypic consequences (insertions, deletions, single nucleotide polymorphisms [SNP’s]) to study the genes glpD, napA, and araC #2) Paleomicrobiology investigation: Determine which biovar(s) have the most similar genes to the amplified sequences from the dental pulp of 3 corpses. #3) Use of genome alignments: Determine an island that is unique to the four genomes for strains that infect humans and is absent in Y. pestis strain 91001. #4) Determine the conservation of a virulence factor in the five strains in the genome alignment. Determine if it is a fully functional product in strain 91001.