Category Archives: Mediterranean

Reservoirs of Salt Adapted Yersinia pestis

The Arab Maghreb is one of the most arid environments to host plague reservoirs. The most recent study on the area highlights the proximity of plague foci to salt water, either the Mediterranean Sea, Atlantic Ocean or importantly inland salt lakes (Malek et al, 2016). These inland salt springs, called chotts, are saltier than the ocean. They were specifically able to cultivate Y. pestis from high salt soil and isolate a high salt tolerant strain of Yersinia pestis from Algeria. Plague foci across North Africa were found at an average of 0.89 km from salt water, while the average distance from fresh water is 4.6 km.

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Plague sites 1940-2015. Red = plague sites, Blue = salt water lakes, Yellow = fresh water. Cropped from Malek et al, 2016.

They also note the importance of L-form Yersinia pestis in their environmental samples. L-form bacteria are an understudied cell wall deficient state that quite a few bacteria, including Yersinia pestis, use for long term survival. The L-form of Y. pestis may be important in environmental persistence. Because they are believed to have a slower reproduction rate,  the L-form may also play a role in altering the molecular clock of some strains. To date,  publications that focus on L-form Y. pestis have been in either Russian or Chinese. It seems clear that the L-form is found in some instances in Asia as well. Importantly, some L-form bacteria can regain their cell wall and return to active ‘normal’ growth.

Soil osmolarity is the key feature that allows (or requires) the L-form to persist. Withstanding osmotic tensions is the primary role of the cell wall. Without the cell wall, the cell loses its ‘normal’ shape, taking on a spherical shape determined by hydrophobic-hydrophilic interactions (like oil and water). As the cell membrane is primarily made of phospholipid, its the L-form shape resembles a sturdy oil globule or a liposome (B below). This was apparent by gram stain when the normal individual short rod-shaped (coccobacilli) cells transformed into clusters of completely round (cocci) cells. This was confirmed under the electron microscope where the change is very apparent.

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They also isolated a strain, Algeria3, a descendant of the third pandemic, from soil containing 4% salt, that can grow in a 15% salt broth. Other Algerian isolates that were not found in high salt soils experimentally survived as well in high salt media if the salt content was ramped up in a step-wise fashion. Growth in high salt conditions altered their protein production to increase those related to osmoregulation, metabolism, outer membrane proteins and others of unknown function.  Osmoregulation genes changes are a direct response to the higher salt concentration. The L-form cells are clearly still metabolically active.

Taken together these protein profiles suggest that it has adapted to survive in the salty soil with the ability to adjust its structure and function as necessary to persist.  They note that other plague reservoirs are in regions of the world with salt lakes or other salty sources, but more environmental sampling will be necessary to determine if this is a universal Y. pestis capability. This all has obviously important implications for plague ecology.


Reference

Malek, M. A., Bitam, I., Levasseur, A., Terras, J., Gaudart, J., Azza, S., et al. (2016). Yersinia pestis halotolerance illuminates plague reservoirs. Scientific Reports, 7, 1–10.

A winter’s worth of work

Its well into spring now and my blogging has perhaps hit an all time low. I have been working on a project that I will write about more later this year. I’ve been reading a lot about environmental history, not the usual material for this blog. Some of it is listed below. It’s a sample of the kind of thing that I need to be read to understand disease in the past. I think it will be worth it eventually even if pollen diagrams and geology diagrams are not very exciting. 

I do have quite a few ideas for new posts, so I will be back…soon. 

A sampling of some of my recent reading:

Büntgen, U., Myglan, V. S., Ljungqvist, F. C., McCormick, M., Di Cosmo, N., Sigl, M., et al. (2016). Cooling and societal change during the Late Antique Little Ice Age from 536 to around 660 AD. Nature Geoscience, 1–7. http://doi.org/10.1038/ngeo2652

Mitchell, P. D. (2015). Human Parasites in Medieval Europe: Lifestyle, Sanitation and Medical Treatment. Advances in Parasitology (Vol. 90, pp. 389–420). Elsevier Ltd. http://doi.org/10.1016/bs.apar.2015.05.001

Mitchell, P. D. (2016). Human parasites in the Roman World: health consequences of conquering an empire. Parasitology, 1–11. http://doi.org/10.1017/S0031182015001651

Brogolio, G.P. 2015. Flooding in Northern Italy during the Early Middle Ages: resilience and adaption, in Post-Classical Archaeologies. 5: 47-68.

Galassi FM, Bianucci, R., Gorini, G., Giacomo M. Paganotti. G.M., Habicht, M.E., and Rühli, F.J. 2016. The sudden death of Alaric I (c. 370–410AD), the vanquisher of Rome: A tale of malaria and lacking immunity, European Journal of Internal Medicine. http://dx.doi.org/10.1016/j.ejim.2016.02.020 [Ahead of Print]

Mensing, S. A., Tunno, I., Sagnotti, L., Florindo, F., Noble, P., Archer, C., et al. 2015. 2700 years of Mediterranean environmental change in central Italy: synthesis of sedimentary and cultural records to interpret past impacts of climate on society, in Quaternary Science Reviews, 116(C), 72–94.

Sadori, L., Giraudi, C., Masi, A., Magny, M., Ortu, E., Zanchetta, G., & Izdebski, A. 2015. Climate,  environment and society in southern Italy during the last 2000 years. A review of the environmental, historical and archaeological evidence, in Quaternary Science Reviews, 1–16.

Li, Y.-F., Li, D.-B., Shao, H.-S., Li, H.-J., & Han, Y.-D. (2016). Plague in China 2014—All sporadic case report of pneumonic plague. BMC Infectious Diseases, 1–8. http://doi.org/10.1186/s12879-016-1403-8

Statskiewicz, A. (2007). The early medieval cemetery at Aschheim-Bajuwarenring: A Merovinigan population under the influence of pestilence? In Skeletal series and their socio-economic context (pp. 35–56).

The Schistosoma in the Reliquary

The 800th anniversary of the birth of Saint Louis, King of France, in 2014 provided an opportunity to obtain a sample of his relics for “scientific identification”. With all relics the chain of custody and its backstory are critical for evaluation. Most of Louis’ relics held in the Basilica of Saint-Denis  were destroyed during the religious violence of  sixteenth century Paris. Fortunately the process of preserving and transferring Louis home to France from the site of death on crusade in Tunis, North Africa left bits of him in several locations.  Part of the preservation process used at the time removed his intestines and other internal organs to be embalmed separately while the remainder of the body was boiled to clean the bones to return to Paris. The bones were enshrined in Paris, while the heart and some viscera were enshrined at the abbey of Monreale near Palermo by his brother Charles, King of Sicily, who oversaw the preparation of the body and its transport back to France.

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13th century Reliquary of Saint Louis, Basicilla of St. Dominic, Bologna Italy. (Source: Photo of Georges Jansoone, public domain on Wikipedia)

During a stop over at Bologna en route to Lyon and then Paris, some of his viscera were removed and interned in the Basilica of Saint Dominic. In 1297 Louis, who had died on 25 August 1270, was officially canonized as Saint Louis of France.  A portion of these visceral relics were given for the consecration of the cathedral of Turin in 1895, and these were transferred to the cathedral of Versailles in 1985. It is from this visceral relic that the 2 g specimen was obtained for scientific evaluation.

Microscopic Examination

The plan is to do a full “medical and forensic anthropological analysis” of the remains. The first result released by Phillipe Charlier’s team is the discovery of a semicircular parasite viewed by Scanning Electron Microscope (SEM) analysis, identified as a male Schistosoma based on its size and morphology.  Schistomsoma are a sexually dimorphic flat worm, also known as a blood fluke, that inhabit the capillaries of the abdomen (mesentery or bladder plexus depending on species) and release their eggs into either feces or urine. The eggs hatch in fresh water and must pass through a fresh water snail before emerging as larvae that can inhabit a mammalian host. Only about 50% of the eggs produced actually exit the body.  The adult worms and eggs that do not reach the feces or urine can cause extensive inflammation resulting in granulomas and fibrosis (scar tissue) to the abdominal organs (liver, spleen, intestines, bladder) and the blood vessels of the abdomen causing an accumulation of fluid in the peritoneal cavity.  Eggs that do breach into the lumen of the intestine cause chronic blood loss into the lumen producing chronic bloody stools. In the worst cases the blood loss can cause anemia.

Saint Louis
Source: Charlier, Bouchet, Weil & Bonnet, 2015.

Compare to a SEM preparation of a modern (non-mummified) male Schistosoma:

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A C-shaped male schistosoma; the smaller female resides in the canal. (Source: David Williams, Illinois State University made public domain, Wikipedia)

King Louis had not been in Tunis long enough for him to contracted schistosomiasis upon his arrival only a month before his death. When and where he contracted the flat worm infestation is open to more speculation.  Schistosoma have been observed in archaeological remains of one individual from 9th century France, but have not yet been commonly observed. Charlier et al. (2015) suggest that Louis’ previous crusade in North Africa between 1250 and 1254, spending some time imprisoned in Damietta, Egypt, is the most likely period for start of the infection. If this is true, then Louis would have had a chronic infection for about 20 years at the time of his death. Charlier’s team do not believe that Schistosomiasis contributed to his death.

So far they have not observed any other parasites in the sample. This is not necessarily surprising considering that they have not yet identified the anatomic source of the specimen. It is not possible to even guess at the anatomic source from the crumbling, blackened specimen pictured in their study (fig. 1). Their analysis is continuing.

Debating Saint Louis’ Cause of Death

As soon as the schistosoma report was published, the debate on the cause of Louis’ death began in the letters of Forensic Science, Medicine and Pathology (where the report was published). So lets begin with the best account of Louis’ death, and go from there.

Beyond skirmishing and entrenching the camp nothing was done, as King Louis was awaiting the arrival of his brother Charles of Anjou (now King of Sicily). Whilst they were waiting encamped, John Tristan fell sick, and died on board one of the ships on August 3rd. A few days later the Legate also died and many other persons, some of fever, some of dysentery. Philip, the King’s eldest son, fell sick with fever; and the King was taken with dysentery (the complaint to which he nearly succumbed in his first Crusade) and died on August 25th. (Guillaume de Nangis account in the Memoirs of Lord Joinville, Book 4, Ch. 4)

Strangely, the plague has traditionally been claimed as Louis’ cause of death. This is completely unfounded since the Black Death will not bring epidemic Yersinia pestis back to the Mediterranean for another 77 years! There is nothing in the account above to suggest plague. This has rightly been dismissed as Louis’ cause of death.

Eric Faure wrote a letter arguing that malaria was a possible cause based on reports of Louis’ history of tertian fevers dating back to the 1242. Faure suggests that Louis went on his first crusade in thanksgiving for surviving “cerebral malaria with a coma” after a relapse in 1244. Cerebral malaria is usually caused by Plasmodium falciparum, which is not a chronic (relapse causing) infection. If Louis suffered relapses of malaria contracted in France then it was most likely Plasmodium vivax, which rarely causes cerebral malaria. Whether or not Louis had cerebral malaria in the 1240s, this doesn’t really inform of his his health in 1270.  Faure also notes that some of the men on Louis’ last crusade had intermittent fevers suggestive of malaria. Faure reaches too far suggesting that the dysentery was a symptom of malaria. Gastrointestinal symptoms are possible but rare in malaria and usually then in children. Philippe Charlier responded with a letter dismissing Faure’s suggestion to look for Plasmodium in the remains, because they would not have caused Louis’ death. Following the report in Lord Joinville’s memoir (quoted above), Charlier reports in his letter that his team is now looking for evidence of bacteria, viruses or amoeba in the embalmed “intestines” that are more likely to be the cause of the “dysenteric syndrome” reported in “Louis and his court”.

I will be watching for the final report, but the idea of a single enteric pathogen being the cause of death may not really represent reality. Based on Joinville’s memoir is appears that the “court” was suffering from a variety of camp diseases found in most medieval armies on prolonged campaigns. In such a situation, co-infection is highly likely particularly with chronic parasites. Indeed, Louis was probably not the only one in camp with schistosomiasis lingering from previous travels.  Although I doubt malaria caused Louis’ dysentery, it is quite possible he was suffering from chronic malaria and that it contributed to weakening his health, making him more susceptible to other pathogens. Indeed co-infection with Schistosoma and Plasmosdium could have made him quite anemic.  It would still be worth knowing if Louis had an active malaria infection, even if Shigella or another enteric pathogen was the primary cause of death.

References:

Charlier, P., Bouchet, F., Weil, R., & Bonnet, B. (Oct. 2015). Schistosomiasis in the mummified viscera of Saint-Louis (1270 AD). Forensic Science, Medicine, and Pathology, 1–2. http://doi.org/10.1007/s12024-015-9722-4

Faure, E. (Dec. 2015). The infections of Saint-Louis: possible involvement of malaria.[Letter]  Forensic Science, Medicine, and Pathology, 1–1. http://doi.org/10.1007/s12024-015-9732-2

Charlier, P. (2016). Neither plague nor malaria, but dysentery as a cause of death for St. Louis. [Letter]  Forensic Science, Medicine, and Pathology, 1–1. http://doi.org/10.1007/s12024-015-9738-9

The Memoirs of Lord Joinvilletranslated by Ethel Wedgwood, E-text. University of Virginia library

Louis IV of France, Wikipedia.

WHO fact sheet: Schistosomiasis

Schistosoma, Wikipedia

Michael Walsh, Schistosomiasis on the Infection Landscapes blog. See this page for the best description of the medical effects of schistosomiasis.