BALL STATE UNIVERSITY

Yersinia pestis
The Bubonic Plague
Jessica A. Eddy
April 18th, 2012

BIO454: Genomes
Instructor: Dr. Blakey

Yersinia pestis, the Bubonic Plague, was vicious to Europe in the 1300s and
although it still exists today, it does not seem to be such a horror. There
are 4 main strains that have been sequenced, but they are categorized
into 3 main subtypes. Scientists currently do not agree with category
placements, and researchers are looking into the differences and
similarities between these strains to better categorize them into subtypes.
Scientists are attempting to understand what has changed in the strains of
Y. pestis. Different approaches are being used to understand this
bacterium because the evolution of pathogens can give insight to future
problems. Yersinia pestis has had to evolve to get where it is today, and
scientists need to alter the approaches used to understand this bacterium,
but this is not an easy task since new technologies are still necessary to
produce more accurate results. Strains of Yersinia pestis have been
sequenced, which is providing more understanding, but there is always
more to be learned and discovered. Understanding the past is a valuable
way to control the future.

Every day, there is a risk that a disease will infiltrate the human body
and in Europe, this became a living nightmare. In the 1300s, Europe was
attacked by a pandemic that is infamous still today: the Black Death.
Yersinia pestis is the culprit responsible for the death of 200 million people
since the Dark Ages (Dawes 2001). According to Ewen Callaway (2011),
geneticists have reproduced the genome of Y. pestis from East Smithfield, a
burial ground created for the plagues victims. Today, this bacterium is found
primarily in fleas and the rats that carry them and help facilitate the spread
of the plague, but not everyone agrees the bacterium was transmitted only
in this manner when the Black Plague reigned. Although Y. pestis is widely
believed to be involved in all plague epidemics since the Black Death,
scientists are struggling to understand how the strain is related to other
plague epidemics (Holmes 2011). This struggle is due to many obstacles that
must be circumnavigated or resolved before proceeding.
The Yersinia genus includes Y. pseudotuberculosis and Y. enterocolitica.
Y. pestis is derived from Y. pseudotuberculosis. Y. pestis has three main
subtypes: antiqua, medievalis, and orientalis. These subtypes are also called
biovars. Subtypes are determined by the capability using glycerol and
reducing nitrate (Chain 2006). There are four classic sequenced strains of Y.
pestis: CO92, KIM, Antiqua, and Nepal. Figure 1 demonstrates clearly the
believed relationship between Y. pseudotuberculosis, 91001, a humanavirulent strain, and the 4 classic sequence strains of Y. pestis. sSNPs
locations between the different strains imply a single biovar is inaccurate
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phylogenetic representation (Chain 2006). The study from Eppinger et al.

(2010) agrees with the aforementioned
statement saying the method does not reflect
the true evolutionary relationship the best.
Three virulence plasmids, pMT, pCD,
and pPCP and a single circular chromosome
are generally affiliated with classical Y. pestis
strains (Chain 2006). Differences in the size
of plasmids were usually accredited to
intrachromosomal deletions and the
addition of genomic fragments. In
strains from the Western United
States, the pMT-PCP plasmid is made
up of the murine toxin plasmid
interrupted by two tandemly repeated
copies of the pPCP plasmid which explains the bigger size of the

Chain 2006

plasmid and the different G-C content in a head-to-tail arrangement

(Eppinger 2010). Table 1 lists strain-specific gene inactivations and or
deletions that were discovered; some of these are specific to only two of the
five genomes (Chain 2006).
Challenges assault scientists and just as Y. pestis has had to evolve to
survive, scientists need to evolve the approaches used to understand this
bacterium. To obtain 700-year-old bacterial DNA is arduous in its own right,
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but the possibility of contamination elevates the level of difficulty (Holmes

2011). An important improvement in gene isolation is the use of a molecular
capture assay that helps identify Y. pestis compared to the other DNA filling
the background. Although the molecular capture assay is helpful, there are
other technological barriers that still need to be resolved so that scientists
can perform in vivo experiments. The plague bacillus does not have a strong
cell wall and cannot endure rigorous purifications that are necessary for
isolation (Dawes 2001). The polymerase chain reaction (PCR) technique is a
sensitive method and was used by a team to obtain the Y. pestis bacterium
from the teeth of a child and two adults. Some are worried about
contamination because, according to Callaway (2011), 108 teeth from 61
individuals have not produced a trace of the bacterium. There are arguments
that Thomas Gilbert, the one who attempted to replicate the experiment, did
not follow the methods properly, but many skeptics agreed with Gilberts
study (Callaway 2011). Sharon DeWitte and Hendrik Poinar were unsatisfied
with PCR and decided to wait until better techniques were available. The
technique they decided to use to sequence DNA damaged by being in the
earth for too long was next-generation DNA sequencers, machines that read
short parts of DNA (Callaway 2011). These sequencers gave Svante Pbo
the ability to draft the sequence of the Neanderthal but did not prove perfect
for Y. pestis; therefore, they turned to another technique to aid them:
targeted capture (Callaway 2011). Targeted capture utilizes lab-synthesized
bait DNA to omit other DNA sequences while gathering the desired old DNA
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(Callaway 2011). This method allowed Johannes Krause and Poinar to extract
and sequence Y. pestis (Callaway 2011). Rino Rappuoli, the vice president of
vaccine research at Chiron, says that genomic approaches can be very
important alternatives to common, primarily empirical approaches and
particularly when strain differences in surface protein sequences and crossreactivity with human proteins have caused problems (Dawes).
An issue other than technology that confronts scientists is the inability
to know which alterations are important in respect to evolution and function
(Dawes 2001). Little appears to be altered in the different strains of Y. pestis
from the Black Plague compared to contemporary versions, so teams are
now looking for other alterations that might explain the fierceness of the
Black Death (Callaway 2011). Genomes from the medieval era appear to lack
singular alterations when compared to contemporary genomes (Bos 2011).
Researchers are attempting to replicate the Black Death pathogen (Calloway
2011).
Although the restoration of the Black Plague may sound dangerous,
there are precautions and reasons that society should be safe. Calloway
states that the plague can be treated with antibiotics and precautions are in
place (Calloway 2011). Also, the conditions of Europe in the 1300s are
drastically different than today. Environmental and epidemiological
determinants assisted in the downfall of Europe (Callaway 2011). Sick
soldiers returning from war used ports that opened up new frontiers to the
plague (Callaway 2011). In addition, the soldiers were not the only ones ill;
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the communities soldiers returned to were sapped due to years of cold, wet
weather and poor nutrition (Callaway 2011). Genetic alterations possibly are
the cause for dissimilarities in manifestation and asperity (Bos 2011). Mark
Achtman, a plague-evolution expert at University College Cork in Ireland,
says that it is possible the plague was not spread by fleas on rats as it is
today but by other animals capable of amplifying the transmission, or
another pathogen provided help (Calloway 2011). We have antibiotics and
take precautions that reduce fear, but it is good researchers are focusing on
ancient pathogens because there are many benefits that can come from the
studies.
Scientists perform experiments, not just to attain money, but to gain
knowledge and benefit for the world. Ancient pathogens have the potential
to provide insight about current or future outbreaks (Calloway 2011). Another
advantage of trying to learn about old pathogens is the possibility for
researchers to recognize pathogens that attacked ancient crops and map the
movement and evolutionary connections with modern strains (Calloway
2011). Sequencing also holds the promise to help establish vaccines and
therapeutics, and as more bacterial pathogens are sequenced, the chance to
use these sequences for vaccines is augmented (Dawes 2001). Genomic
data from old microbes can clarify mechanisms of the evolution of pathogens
and adjustment of emerging or re-emerging contaminations (Bos 2011). The
evolution of Y. pestis has taken more than a few years to develop, and

scientists will need more than a few years to understand exactly what took
place.
The evolution of pathogens can be very helpful indicators of what
might happen in the future, and this is one reason why Y. pestis is studied. Y.
pestis is a blood-borne pathogen that can be lethal if not treated and is
closely related to Y. pseudotuberculosis which tends to cause non-fatal gut
disease (Dawes 2001). It is believed the two have gone off in separate
directions only 1,500 to 20,000 years ago (Dawes 2001). A somewhat far
away predecessor of both Y. pestis and Y. pseudotuberculosis, Y.
enterocolitica causes non-fatal gut disease too (Dawes). Although Y. pestis
and Y. pseudotuberculosis both derive from the same ancestor Y.
enterocolitica, Y. pseudotuberculosis causes non-fatal gut disease like Y.
enterocolitica which is more distantly related than Y. pestis that is dissimilar
in that it can be lethal; researchers want to know why (Dawes).
Diseases have fascinated researchers and scientists for a long time
because they are something that more than likely are not going away and
more likely will resurface. Y. pestis is not gone from the world, but if it
attempts to strike with the same ferocity as in Europe, solutions should be
ready and waiting. There have been and will be problems along the way, but
technologies are created in response so that science may get around said
problems and garner better results. Y. pestis has been sequenced and is
providing insight, but there is always more to be learned and discovered.

Hopefully, our understanding will evolve and grow to fit the ever-changing
pathogens.

References
Bos, K. I., et al, 2011 A draft genome of Yersinia pestis from victims of the
Black Death. Nature 478: 506-510.
Callaway, E., 2011 Plague genome: The Black Death decoded. Nature 478:
444-446.
Chain, P., et al, 2006 Complete Genome Sequence of Yersinia pestis Strains
of Antiqua and Nepal516: Evidence of Gene Reduction in an Emerging
Pathogen. Journal of Bacteriology 188: 4453-4463.
Dawes, H., 2001 Yersinia pestis: Sequence sheds light on the plagues past.
Current Biology 11: R949-R951.
Eppinger, M., et al, 2010 Genome Sequence of the Deep-Rooted Yersinia
pestis Strain Angola Reveals New Insights into the Evolution and Pangenome
of the Plague Bacterium. Journal of Bacteriology 19: 1685-1699.
Holmes, E. C., 2011 Genomics: Plagues progress. Nature 478: 465-466.

PowerPoint also used:

Achtman M., et al., 1999 Yersinia pestis, the cause of plague, is a recently
emerged clone of Yersinia pseudotuberculosis. PNAS 96: 14043-14048.
Castle, K., et al, 2011 A National Park Service Managers Reference Notebook
on Plague (Yersinia pestis). National Park Service. ii-97.
World Map image:
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