Dr. Manuel Varela: Yersinia pestis

May 15, 2017 by

An Interview with Dr. Manuel Varela: Yersinia pestis


Michael F. Shaughnessy –


1) Dr. Varela, today, I would like to ask you about Alexandre Yersin, who apparently was well known for his studies of the plague bacterium that was interestingly enough, named after him.  What do we know about his birth and early life?


In 1863, Alexandre Emile John Yersin, known as the discoverer of the plague-causing bacterium Yersinia pestis, was born on September 22 in the municipality of Aubonne, located in eastern Switzerland. Just a few weeks prior to his birth, Yersin’s father, a natural science educator, died suddenly. Thus, Yersin’s mother and his two siblings (Franck and Emilie) moved to Morges, Switzerland, where as a single parent she founded a school for girls. As a child it is reported that Yersin had an interest in the natural sciences, which was the study of the biological, physical, geological, and chemical world. Yersin went to high school in Lausanne, Switzerland. He then began his studies in medicine while in Lausanne but then moved to Marburg, Germany, and later Paris, France, where he took his medical degree in 1888.


Dr. Yersin then took a post in the laboratory of the famous Dr. Prof. Louis Pasteur at the Ecole Normale, having been invited by Dr. Emile Roux to be his (Roux’s) assistant in Pasteur’s laboratory. While there, Dr. Yersin took part in the discovery of Pasteur’s famous rabies vaccine. In 1889, Yersin became a citizen of France, apparently in order to be allowed to treat patients there. Yersin’s parents were French, and this may have had an influence in his decision, as well. During that same year, Dr. Yersin spent a few months working in the laboratory of the famous Dr. Robert Koch, in Berlin, Germany, acquiring training as a bacteriologist. Next, Yersin moved back to Paris to work and collaborate with Roux at the newly established Institute of Pasteur, where they studied diphtheria caused by the bacterium called Corynebacterium diphtheriae and discovered the diphtheria toxin.


2) I suspect everyone, at some time or another has heard about the “Black Death” and various forms of the plague.  In the old days and Middle Ages, obviously sanitation was not the same as today. But what were some of the contributing factors to the “plague bacterium”?


The plague disease in humans is caused by the Yersinia pestis bacterium, a bacillus-shaped Gram-negative microbe sharing the same bacterial family as the Escherichia coli, Salmonella enterica, and the Shigella dysenteriae bacteria. The plague is considered to be a so-called zoonotic disease, because it involves transmission to humans from a variety of animals.


Fleas, for example, then carry the bacteria from the animals to people.


There are three kinds of the plague. The first is the most common and is called the bubonic plague, characterized by the so-called bubo lesion (the medical term for bubo is adenitis) on, for example, the arms and neck of its patients or in places where lymph nodes are found. The bacterium is transmitted to humans by fleas and other insects having emerged from rodents and other small animals.


The pneumonic plague is a severe lung infection also transmitted by fleas and rodents but also by person-to-person contact and via the airborne mode of transmission.


The aggressively lethal septicemic plague, where the bacteria actively multiply in the patient’s blood, occasionally arises from first acquiring the bubonic or pneumonic plague forms.


In our history, there have been three so-called great plague pandemics, i.e., world-wide presence of the disease, all pandemics of which are well-known and documented. The first great pandemic is referred to as the urban or the Justinian plague, having started in Egypt in 541-545 AD, during the reign of the Byzantine emperor, Justinian I, and lasting well into the 6th and 8th centuries, killing, by some accounts, an estimated 100 million victims in its wake.


In fact, Justinian I himself is thought to have suffered from the plague, but he survived the disease. It is further believed, however, that the Justinian plague pandemic contributed to the eventual fall of the Roman Empire.


The second plague pandemic, called the medieval plague or “Black Death” by later historians and the “Great Mortality” by its victims who perished from it, started approximately in 1320-1353 and lasting well into the 18th century with multiple epidemics throughout Europe and Asia, killing an estimated 20 to 200 million people, decimating, in particular, the European population and profoundly affecting its societal stability.


The term “Black Death” has an uncertain origin. Some say the term arose because of the black color of the dead tissue apparent in the septicemic patients, but this explanation is suspect because the septicemic plague was relatively rare. Another explanation holds that the Black Death term arose because of an error in translation.


The story starts with the Swedes in 1555 and the Danes in 1605, and later by historian Johannes Isaacus Pontanus in 1631, all of whom were apparently under the misapprehension that the Latin terms used to describe early plague epidemics by Seneca and de Corbeil (atra mors), or by Simon de Covino (mors nigra), meant “Black Death” or “Big Death” when these early writers probably more likely meant to say “Great Mortality” after the shockingly substantial numbers of deaths associated with the plague. In any case, the origin of the term “Black Death” remains a mystery.


The third, or modern plague pandemic, is said to have started in 1894 in Hong Kong, lasting well into the early 20th century, and killing an estimated 200 million people. Some believe this pandemic is present today. It is this pandemic, however, from which Yersin is said to have isolated the bacterium known as the causative agent of the plague disease.


Altogether, it is widely believed that the plague has wiped out approximately one-fourth to one-third of the world’s human population, thus permanently altering the course of history. The significance of this Y. pestis microbe, therefore, cannot be understated.


3) How did he come to discover this bacterium that was ultimately named after him?


The story of how Dr. Yersin discovered the plague-causing bacterium begins when he travelled to Hong Kong in June of 1894, shortly after the start of the modern or third great plague pandemic. Yersin’s purpose in making the trip was to investigate the emerging outbreak there of the bubonic plague, which by the time Yersin arrived had already killed an estimated 60,000 people there.


Yersin was not immediately welcomed in Hong Kong, however, as local officials had already appointed the well-respected and very famous Japanese physician and bacteriologist Dr. Shibasaburo Kitasato, who had arrived in Hong Kong three days earlier, to lead the investigation. Dr. Kitasato had worked in the eminent Robert Koch’s laboratory and had discovered the tetanus-causing bacterium, and he had worked in Emil von Behring’s laboratory, developing potential antisera therapies for the tetanus, anthrax and diphtheria—all serious diseases. Kitasato had even been nominated for the very first Nobel in the category of physiology or medicine.


Consequently, Yersin, prevented from conducting autopsies, had trouble even obtaining plague tissue samples, reportedly going so far as to bribe handlers of dead bodies of the plague victims to “appropriate” specimens. Eventually, perhaps reluctantly, Yersin was allowed to proceed with his investigation, but only after repeated appeals to the local Hong Kong officials.


Once fully accepted, Yersin went to work with his investigation of the outbreak.


First, he acquired samples, properly this time, of the buboes tissues from the autopsied corpses of the Hong Kong plague victims. Next, he found bacteria within the buboes tissues while studying them under the microscope. They were bacillus-shaped (i.e., rod or stick-like) in his microscope. Yersin then isolated the bacilli bacteria by culturing them on a special peptone agar culture medium in an incubator. Next, using his purified bacterial cultures, he inoculated test animals, guinea pigs, mice and rats, with his purified bacteria. The inoculated laboratory animals died. Then Yersin examined the tissues and organs of the dead animals and found the same bacillus-shaped bacteria in the dead animals, using his microscope.


He quickly returned to Paris and shared his findings, presenting them first at a scientific conference (a congress) and then publishing them in the journal Annals of the Institute Pasteur, in 1894. It is reported that Yersin even personally informed both Drs. Roux and Pasteur of his discovery.


4) What were the initial reactions?


In short, the reactions were mixed. In certain circles, Yersin’s plague bacillus was immediately accepted as the causative agent of the second great pandemic. On the other hand, Dr. Kitasato had also essentially performed the same type of investigation as Yersin, identifying the plague bacterium from the blood of plague victims and animals. In fact, Kitasato had even published his work in the prestigious journal Lancet, prior to Yersin’s publication. All told, it is becoming increasingly clear that both Yersin and Kitasato deserve credit as co-discoverers of the plague bacterium, each investigator having worked independently of the other.


It is not entirely clear why Yersin, and not Kitasato, received the bulk of the credit for the historical discovery. The microbe was, after all, named to honor only Yersin. One explanation that has been put forth is perhaps that Kitasato’s preparation was not completely pure and contaminated by other bacteria, in particular, with the so-called Streptococcus. Another idea is that, in his paper, Kitasato had stated in Japanese that his bacilli, while appearing like Streptococcus, were not to be mistaken as such, as the bacteria were bacillus in shape, but perhaps that that proper phrasing was missed in the subsequent translation. The literature today frequently lists both Yersin and Kitasato as co-discoverers of the plague microbe.


At the time of his publication, the plague bacteria were named by Yersin as Pasteurella pestis, but other names had been given for these same bacteria, such as Bacterium pestis, Bacillus pestis, and Pestisella pestis. The term “pestis” is Latin for plague. The scientific name we have come to know today, Yersinia pestis, was established in the mid-20th century, again mainly as a tribute to Yersin.


After his historical discovery, Yersin focused his collaborative research efforts with Drs. Albert Calmette and Emile Roux on developing a plague antiserum, which unfortunately had poor efficacy. Yersin moved to Vietnam where he established a school of medicine. Yersin died in 1943 on the first of March, at 79 years of age.


Kitasato, on the other hand, while losing out on the Nobel in 1901 to his collaborator, von Behring, for their work on diphtheria, nonetheless made other significant discoveries. For example, with his student, Shiga Kiyoshi, they isolated the causative agent of dysentery, the bacterium known as Shigella dysenteriae. Seventy eight years old, Kitasato died in Tokyo, Japan, on the 13th of June in 1931.


Neither Yersin nor Kitasato received the Nobel for their discoveries.


For many years, there was a controversial issue about whether the Y. pestis was the causative agent of the first two plague pandemics, the Justinian and the medieval plagues, as well as the third pandemic, from which Yersin and Kitasato had demonstrated. In the first two instances, corpses from both of these pandemics had been exhumed from their graves, and their teeth pulp examined for the presence of the Y. pestis contagion.


After all of these years, only the teeth pulp is thought to harbor any remaining plague bacteria. The Y. pestis bacteria were indeed found in the samples tested. Thus, because Y. pestis has been definitively shown to be responsible for the emergence of all three pandemics, it demonstrates that this one pathogenic bacterial species alone has killed more human beings in history than any other contagion known to humankind.


5) What impact does this discovery have on us today? Are there any variants of these bacteria still in existence, and are they resistant to antibiotics?


Oddly enough, the plague remains only a relatively minor problem today, with approximately 10 cases per year in the US. Beginning in 2011, the DNA genomes of the Y. pestis bacteria from each of the pandemics were pieced together and later completely sequenced and compared to the genomes of modern Y. pestis. Surprisingly, the ancient genomes did not differ widely with its modern genomic counterparts. In fact, only about 100 nucleotide differences were observed between the ancient (deadly) and modern (not so deadly) Y. pestis genomes.


The question, thus, arose as to why the ancient form of the bacterium was so deadly in ancient times while its modern and mostly genetically identical descendant is, relatively speaking, virtually harmless today. The immediate obvious answers are that the fleas, rodents and other so-called vectors of disease transmission are now sufficiently controlled to block the contagion today; another idea was that perhaps all of the people who succumbed to the plague and died could no longer pass along their susceptible genes into the next generations while those who survived were able to propagate their immune genes into the succeeding generations, and now modern human beings are consequently and simply immune to the plague. A third notion is that improvements in medicine, such as antibiotics and vaccines, plus sanitation, and personal hygiene practices, were all effective enough to countermand the modern plague.


Evidence for a fourth explanation, however, was published in 2015 to account for the vast differences in virulence between the ancient and modern plague bacteria. In the study, investigators closely examined the DNA sequences of the ancient versus the modern genomes of the two Y. pestis bacterial forms and found two sequence variations between the ancient deadly and the modern harmless bacteria. In the first genetic variation, the ancient plague bacterium acquired a new plasmid containing a gene, called pla, which encodes a so-called plasminogen activator protein, Pla, a protease enzyme that breaks-up another inactive protein, called plasminogen, thus converting it into an active form of the protein, called serine protease plasmin.


This active plasmin then allows the ancient bacteria to escape from clots that are originally meant to contain the microbes. Now the escaped bacteria move on to infect the lymph nodes and grow within them, swelling up and forming the characteristic buboes.


The second variation of these ancient pla gene-containing microbes had a new mutation at a single-point within the pla gene that made the resulting protease work even more efficiently than before in order to greatly enhance the virulence of the ancient plague strain.  The new mutation caused the bacteria to convert from a pneumonic form to a more deadly form that’s readily transmitted by coughing or flea bites.


Thus, even though Yersin’s plague bacterium has a variety of virulence factors to cause disease, subtle sequence variations in the ancient plague bacterium genome were all that were needed to have a profound enhancement in starting pandemics, in order to further its deadly effects. Somehow, the dangerous ancient bacteria lost or neutralized their deadly genomic variations to pacify their modern counterparts, making the plague a rare infectious disease nowadays.


One potentially alarming impact has to do with the notions of bioterrorism and of the “weaponizing” of the plague bacterium by certain countries. Thus, care, prevention, and bio-defensive measures are often called for in circumstances where individuals working in this area may be at risk.


Regarding your inquiry about antibiotics and resistance to these agents, modern plague is often effectively treated with the antibiotic streptomycin. If streptomycin resistant variants of Y. pestis emerge, however, then alternative therapy is invoked with the chloramphenicols, sulfamethoxazole-trimethoprim, and the tetracyclines. In general, many bacteria have developed resistance mechanisms to all of these antimicrobials, but do not appear to be a significant problem for confounding treatment efforts against the modern plague, yet. In some geographical pockets, drug- and multi-drug resistant strains have emerged, but spread of infection through these populations will be prevented with vaccines.


6) What have I neglected to ask?


I think one interesting incident to point out is that, of the many hundreds of millions of deaths which were, in fact, due directly to the Y. pestis plague bacterium, it had been mistakenly blamed for a serious outbreak of cholera in London during the 1850s. The rumors began when the excavations had been started for London’s new sewer system. Apparently, during the dig, a large number of human skeletons had been unearthed. Some of the corpses were still in the process of decaying.


One London area in particular was suspect, called the Golden Square district. Prior to its development the same area had been called Craven’s Pest Field, and 200 years earlier it had been the site of a mass grave and burial grounds from the multitude of plague deaths that had occurred. The field contained the human remains of about 4000 bodies, buried unceremoniously in mass pits.


Craven’s field had been left untouched for many years because of the fear of releasing the contagion. Until, of course, urban development encroached, making the area the new Golden Square district, with townhouses. The area was now suffering a severe cholera epidemic in the late 1850s.


Some had speculated that the ghosts of the Great Mortality had had their resting places disturbed by the sewer excavations, or that because of the humiliating way in which they had been buried (aligned improperly and in mass graves) the ghosts had wreaked their vengeance by bestowing the cholera upon the Londoners of the Golden Square district.


Another explanation (which was more reasonable but nonetheless incorrect) was that the sewer construction had inadvertently contaminated the city with plague contagion from the physical disruption of the Black Death burial site. To provide evidence that there was no plague contamination from the sewer project, a map of London had been drawn showing the plague pit, the sewer lines, and the houses of cholera victims.


There was no connection between the plague burial pit and the cholera or the sewer lines. There WAS, however, a connection between the water supply and sewage contamination of the cholera contagion, but that’s another story!

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