Paleomicrobiology is a recent science. Paleomicrobiology describes the history of infectious diseases through research of ancient microbes. The dentistry with the tooth in particular has contributed to the progress of this science. The dental pulp is very well protected in the centre of the tooth. Molecular techniques such as “suicide PCR” and Multiple Spacer Typing have identified and characterized micro-organisms in the ancient samples. The detection of bacterial DNA from ancient dental pulp provided the scientific evidences for diagnosis of the infectious diseases of the past. The cooperation of paleomicrobiology and paleodontology also contributed the new knowledge to human pathology

Gérard Aboudharam

Michel Drancourt

Tung Nguyen-Hieu

Paleomicrobiology is a recent science. Paleomicrobiology describes the history of infectious diseases through research of ancient microbes. The dentistry with the tooth in particular has contributed to the progress of this science. The dental pulp is very well protected in the centre of the tooth. Molecular techniques such as “suicide PCR” and Multiple Spacer Typing have identified and characterized micro-organisms in the ancient samples. The detection of bacterial DNA from ancient dental pulp  provided the scientific evidences for diagnosis of the infectious diseases of the past. The cooperation of paleomicrobiology and paleodontology  also contributed the new knowledge to human pathology

Directory of Open Access Journals

     N g u y e n - H i e u   e t   a l .                                                                                                   R E V I E W      Bull Int Assoc Paleodont. Volume 5, Number 1, 2011     www.paleodontology.com     48 Mini Review:  Dental Pulp as a Source for Paleomicrobiology  • Tung Nguyen-Hieu, Gérard Aboudharam, Michel Drancourt •  Unité de Recherche sur les Maladies Infectieuses et Tropicales Emergentes, UMR CNRS 6236, IRD198, IFR48, Faculté de médecine, Université de la Méditerranée, Marseille, France  Address for correspondence: Nguyen Hieu Tung  Unité de Recherche sur les Maladies Infectieuses et Tropicales Emergentes Faculté de médecine, Université de la Méditerranée Marseille, France E-mail: nghtung2183@yahoo.fr    Bull Int Assoc Paleodont. 2011;5(1):48-54.  Abstract Paleomicrobiology is a recent science. Paleomicrobiology describes the history of infectious diseases through research of ancient microbes. The dentistry with the tooth in particular has contributed to the progress of this science. The dental pulp is very well protected in the centre of the tooth. Molecular techniques  such  as  “suicide  PCR”  and  Multiple  Spacer  Typing  have  identified  and  characterized micro-organisms in  the  ancient  samples.  The  detection of  bacterial DNA from  ancient dental pulp  provided the scientific evidences for diagnosis of the infectious diseases of the past. The cooperation of paleomicrobiology and paleodontology  also contributed the new knowledge to human pathology.   Keywords: Paleomicrobiology; Ancient Teeth; Dental Pulp; Microbial Detection  Introduction In 1993, after the first molecular detection of Mycobacterium tuberculosis in ancient skeletons (1), paleomicrobiology    appeared  as  a  new  discipline  for  the  identification  and  characterization  of microorganisms  in  the  ancient  samples.  Frozen  tissues,  mummies,  skeletons  or  bones  were  the ancient  remains  often  used  in  paleomicrobiological  studies.  However,  the  major  limitations  of  the ancient DNA (aDNA) study consist in the degradation and fragmentation of aDNA, contamination of modern DNA or external sources. Moreover, aDNA extraction from mummified tissue and bones is complicated because rehydration and decalcification are necessary steps. Therefore, another  source of ancient specimens that possibly limited the disadvantages above, were proposed: dental pulp (2), Figure 1. Vascularization of this organ is important, proportionally comparable with the human brain (3). Pathogenic bacteria circulating in blood may occur and colonize the dental pulp. A guinea-pig      N g u y e n - H i e u   e t   a l .                                                                                                   R E V I E W      Bull Int Assoc Paleodont. Volume 5, Number 1, 2011     www.paleodontology.com     49 model has confirmed the possibility to detect bacterial DNA in dental pulp after experimental infection by Coxiella burnetii. This experimental model has also confirmed that pathogenic bacteria in blood had penetrated and colonized the dental pulp and that could then be used for diagnosis of bacteremia (4,5). Moreover, Bartonella henselae DNA and Bartonella grahamii DNA had been isolated from the dental pulp of the cats that had been bacteremic for Bartonella sp. then euthenased and buried one year previously, according to French army veterinary records (6). These results indicate that dental pulp was an excellent specimen for diagnosis of ancient bacteremia.  Figure 1 Ancient DNA recovered from dental pulp.  Paleomicrobiology of the dental pulp: methods In 1998, a team of researchers in France had first introduced dental pulp collected from ancient teeth as a suitable tool for the diagnosis of ancient infectious diseases (7). The advantages of using dental pulp for paleomicrobiological study is its better preservation in a closed cavity.  In the closed cavity, the soft tissues are preserved from environmental contamination and attacks of degrading agents.  Moreover, the DNA extraction from soft tissues of the dental pulp is simple (2). An experimental animal model infected with Coxiella burnetii had also confirmed the possibility of finding bacterial DNA in dental pulp 20 days after bacteremia (4). A second guinea-pig model infected with Coxiella burnetii showed persistence of the bacteria in the dental pulp (5). The pulp can then be used as a tool for retrospective diagnosis when there was a bacteremia. Afterwards, this team  proposed the “suicide PCR” for molecular identification of bacterial DNA from ancient teeth with strict criteria to avoid any risk  of  contamination  and  Multiple  Spacer  Typing  for  the  genotyping  of  ancient  bacteria  (8).  The important criteria of “suicide PCR” included: 1- The primers are used only once, 2-There were no positive controls, 3-The negative controls should be added, 4-Negative results are tested by other primers, 5-Positive results are confirmed by sequencing (9). The suicide PCR principle was used by another Greek team in 2006. This team using dental pulp has shown that the plague of Athens was in      N g u y e n - H i e u   e t   a l .                                                                                                   R E V I E W      Bull Int Assoc Paleodont. Volume 5, Number 1, 2011     www.paleodontology.com     50 fact  typhoid fever (10). Rapid Diagnostic Test was an immunodetection method developed by another team for special diagnosis of plague from ancient remains  (11). From 1998 to the present day, several independent teams using these methods,  detected multiple pathogens from ancient dental pulp to confirm the epidemics of the past, Table 1.  Table 1 Microorganisms detected from ancient teeth Microorganisms  Infectious diseases  References Anelloviridae  Virus 25 Bartonella henselae  Cat-scratch disease 24 Bartonella quintana  Trench fever 17-19;21;22 Mycobacterium leprae  Leprosy 23 Mycobacterium tuberculosis  Tuberculosis 23 Rickettsia prowazekii  Typhus 21;22 Samonella enterica serovar Typhi  Typhoid fever 10 Yersinia pestis  Plague 7-9;11;13-18   Paleomicrobiology of the dental pulp: contributions In history, the three plague pandemics were cited as humanity’s disaster, in which an epidemic known as Black Death  occurred in 1347 and then spread from the Caspian Sea to almost all European countries. Over the next few years, Black Death had killed an estimated 17 – 28 million Europeans, representing approximately one third of the population at the time (12). In 1998, Yersinia pestis DNA had first been  identified in dental pulp of the ancient teeth collected from skeletons excavated from 16th and 18th century graves of persons suspected of having died of plague  (7). Genotyping had then yielded Y. pestis Orientalis strain  (8,13). These results were later reinforced by other independent teams to confirm the presence of plague in Medieval Europe  (11,14-16). Interestingly, two recent studies detected co-infection of the body louse,  Bartonella quintana, along with Yersinia pestis in ancient  dental  pulp,  suggesting  that  the  body  louse    may  have  been  a  vector  for  interhuman transmission of medieval plague pandemics  (17-18). Bartonella quintana is the causative agent of trench fever disease and transmitted by lice, fleas and ticks to humans. Molecular detection of B. quintana in a 4000-year-old human tooth (southeastern France)  indicated  that  B.  quintana  bacteremia  had  occurred  in  prehistoric  humans  19.  Typhus epidemic had  been  demonstrated  in  the  20th  century  as  “a  best friend”  of  wars.  In the  past,  this epidemic spread rapidly in Europe from Spain to France and Italy following the wars between Emperor Charles V and King François I (20). The city of Douai in northern France was besieged from 1710 to 1712 during the war of Spanish succession when France and Spain opposed the other nations. The causative agent of typhus, Rickettsia prowazekii strain Madrid E type B, and B. quintana DNA had been isolated from ancient dental pulp of the individuals buried in Douai. This molecular detection      N g u y e n - H i e u   e t   a l .                                                                                                   R E V I E W      Bull Int Assoc Paleodont. Volume 5, Number 1, 2011     www.paleodontology.com     51 allowed the researchers to confirm the presence of the typhus epidemic in Douai from 1710 – 1712 (21).  Typhus was  later  a  constant  companion of  Napoleon’s  army  in  all  European wars.  In 2006,  evidence of louse-transmitted diseases in soldiers of Napoleon’s Grand Army in Vilnius was reported when R.  prowazekii  and  B.  quintana DNA had  been amplified  by  “suicide PCR”  from dental  pulp collected from the remains of the soldiers. These results showed that louse-borne infectious diseases had affected nearly one-third of Napoleon’s soldiers buried in Vilnius and suggested a possible cause of the French retreat from Russia  (22), Figure 2 and 3. Mycobacterium  tuberculosis  and  Mycobacterium  leprae  had  also  been  detected  in  ancient  teeth recovered from a 1st century tomb in Jerusalem, Israel. The findings had suggested the co-infection of these  two  mycobacteria  in ancient  human populations (23).  Moreover DNA of  Samonella  enterica serovar  Typhi  had  been  successfully  identified  in  dental  pulp  of  ancient  teeth  collected  from Kerameikos cemetery. This discovery had demonstrated that typhoid fever could be the cause of the plague of Athens in 430 – 426 BC (10). Interestingly, molecular detection of Bartonella henselae in dental pulp of 800-year-old cats suggested the occurrence of Cat-scratch disease within a medieval population in France because archaeological and historical data indicated that these cats had lived in close contact with people (24). A recent study had surprisingly reported the detection of an ancient virus (Anelloviridae DNA) from 200-year-old dental pulp recovered from ancient teeth of Napoleon’s soldiers in Kaliningrad, Russia (25).    Figure 2 and 3 Vilnius, 1812, Pictures of Yann ARDAGNA (UMR 6578 CNRS Université de la Méditerranée-CNRS-EFS        N g u y e n - H i e u   e t   a l .                                                                                                   R E V I E W      Bull Int Assoc Paleodont. Volume 5, Number 1, 2011     www.paleodontology.com     52 Perspectives X-rays were used to detect and locate the target teeth in the jaws (Figures 4 and 5). The ancient tooth was usually bisected longitudinally for the recovery of dental pulp (Figures 6, 7 and 8). This method led to the destruction of its morphological structure and possible external contamination . “Orthograde entrance technique” is an “original method” to collect dental pulp with morphological conservation of ancient teeth that will be used afterwards by anthropologists  (26).  However, this technique has some disadvantages due to the relatively fragile, permeable and weak nature of the tooth root. The first method  is  probably  the  simplest  and  most  effective  despite  the  disadvantage  of  morphological changes of the tooth, it is necessary to restore the tooth by bonding techniques. The  ancient  proteins  seem  to  be  more  resistant  than  aDNA.  Lipids  such  as  mycolic  acids  of  M. tuberculosis also appear to be particularly robust  in ancient remains (27). Mass spectrometry had successfully identified specific bacterial proteins  of M. tuberculosis from archeological bone samples (28).  Therefore,  this  method,  detecting  microbial  non-nucleotidic  biomolecules  from  ancient  dental pulp,  may  hereafter  become  a  promising  approach  to  complement  aDNA  analysis  in paleomicrobiology. In conclusion, detection of pathogen traces from ancient teeth by using many paleomicrobiological methods has provided the scientific proofs to elucidate several infectious diseases in the past. This great cooperation of paleomicrobiology and paleodontology has also contributed  new knowledge of human pathology as well as  evidence of human history.    Figure 4 Mandible of a child about 9 years (Observance massgrave-Marseille 1724)  Figure 5 Radiography of the mandible and choice of the tooth (34 non-eruptive)       N g u y e n - H i e u   e t   a l .                                                                                                   R E V I E W      Bull Int Assoc Paleodont. Volume 5, Number 1, 2011     www.paleodontology.com     53    Figure 6 Realization of the fracture using a carborundum disc on handpiece  Figure 7 Recovery of the dental pulp   Figure 8 After the fracture and recovery of the pulp, tooth is ready to be glued  References 1.  Spigelman  M,  Lemma  E.  The  use  of  the  polymerase  chain  reaction  to  detect  Mycobacterium  tuberculosis  in  ancient skeletons. Int J Osteoarchaeol 1993;3:143. 2.  Drancourt M, Raoult D. Palaeomicrobiology: current issues and perspectives. Nat Rev Microbiol 2005;3:23-35. 3.  Kim S. Regulation of pulpe blood flow. J Dent Res 1985;64:590-596. 4.  Aboudharam G, Lascola B, Raoult D, Drancourt M. Detection of Coxiella burnetii DNA in dental pulp during experimental bacteremia. Microb Pathog 2000;28:249-254. 5.  Aboudharam G, Drancourt M, Raoult D. Culture of C. burnetii from the dental pulp of experimentally infected guinea pigs. Microb Pathog 2004;36:349-350. 6.  Aboudharam G, Vu DL, Davoust B, Drancourt M, Raoult D. Molecular detection of Bartonella spp. in the dental pulp of stray cats buried for a year. Microb Pathog 2005;38:47-51. 7.  Drancourt M, Aboudharam G, Signoli M, Dutour O, Raoult D. Detection of 400-year-old Yersinia pestis DNA in human dental pulp: an approach to the diagnosis of ancient septicemia. Proc Natl Acad Sci U S A 1998;95:12637-12640. 8.  Drancourt M, Roux V, Dang LV et al. Genotyping, Orientalis-like Yersinia pestis, and plague pandemics. Emerg Infect Dis 2004;10:1585-1592.      N g u y e n - H i e u   e t   a l .                                                                                                   R E V I E W      Bull Int Assoc Paleodont. Volume 5, Number 1, 2011     www.paleodontology.com     54 9.  Raoult D, Aboudharam G, Crubezy E,  Larrouy G, Ludes B, Drancourt M. Molecular identification by "suicide PCR" of Yersinia pestis as the agent of medieval black death. Proc Natl Acad Sci U S A 2000;97:12800-12803. 10. Papagrigorakis  MJ,  Yapijakis  C,  Synodinos  PN,  Baziotopoulou-Valavani  E.  DNA  examination  of  ancient  dental  pulp incriminates typhoid fever as a probable cause of the Plague of Athens. Int J Infect Dis 2006;10:206-214. 11. Bianucci R, Rahalison L, Massa ER, Peluso A, Ferroglio E, Signoli M. Technical note: a rapid diagnostic test detects plague in  ancient  human  remains:  an  example  of  the  interaction  between  archeological  and  biological  approaches  (southeastern France, 16th-18th centuries). Am J Phys Anthropol 2008;136:361-367. 12. Drancourt M, Raoult D. Molecular insights into the history of plague. Microbes Infect 2002;4:105-109. 13. Drancourt M, Signoli M, Dang LV et al. Yersinia pestis Orientalis in remains of ancient plague patients. Emerg Infect Dis 2007;13:332-333. 14. Bianucci R, Rahalison L, Peluso A et al. Plague immunodetection in remains of religious exhumed from burial sites in central France. Journal of Archaeological Science 2009;36:616-621. 15. Haensch S, Bianucci R, Signoli M et al. Distinct clones of Yersinia pestis caused the black death. PLoS Pathog 2010;6. 16. Wiechmann  I,  Grupe  G.  Detection  of  Yersinia  pestis  DNA  in  two  early  medieval  skeletal  finds  from  Aschheim  (Upper Bavaria, 6th century A.D.). Am J Phys Anthropol 2005;126:48-55. 17. Tran TN, Le Forestier C, Drancourt M, Raoult D, Aboudharam G. Co-detection of Bartonella quintana and Yersinia pestis in an 11th - 15th burial site, Bondy, France. Am J Phys Anthropol 2011. 18. Tran  TN, Signoli M, Fozzati L, Aboudharam G, Raoult D, Drancourt M. High throughput, multiplex pathogen detection authenticates plague waves in medieval Venice, Italy. PLoS One 2011;6:e16735. 19. Drancourt M, Tran-Hung L, Courtin J, Lumley H, Raoult D. Bartonella quintana in a 4000-year-old human tooth. J Infect Dis 2005;191:607-611. 20. Raoult D, Woodward T, Dumler JS. The history of epidemic typhus. Infect Dis Clin North Am 2004;18:127-140. 21. Nguyen-Hieu  T,  Aboudharam  G,  Signoli  M,  Rigeade  C,  Drancourt  M,  Raoult  D.  Evidence  of  a  louse-borne  outbreak involving typhus in Douai, 1710-1712 during the war of Spanish succession. PLoS One 2010;5:e15405. 22. Raoult D, Dutour O, Houhamdi L et al. Evidence for louse-transmitted diseases in soldiers of Napoleon's Grand Army in Vilnius. J Infect Dis 2006;193:112-120. 23. Matheson  CD,  Vernon  KK,  Lahti  A  et  al.  Molecular  exploration  of  the  first-century  Tomb  of  the  Shroud  in  Akeldama, Jerusalem. PLoS One 2009;4:e8319. 24. La VD, Clavel B, Lepetz S, Aboudharam G, Raoult D, Drancourt M. Molecular detection of Bartonella henselae DNA in the dental pulp of 800-year-old French cats. Clin Infect Dis 2004;39:1391-1394. 25. Bedarida S, Dutour O, Buzhilova AP, de MP, Biagini P. Identification of viral DNA (Anelloviridae) in a 200-year-old dental pulp sample (Napoleon's Great Army, Kaliningrad, 1812). Infect Genet Evol 2010. 26. Alakoc YD, Aka PS. "Orthograde entrance technique" to recover DNA from ancient teeth preserving the physical structure. Forensic Sci Int 2009;188:96-98. 27. Poinar  HN,  Stankiewicz  BA.  Protein  preservation  and  DNA  retrieval  from  ancient  tissues.  Proc  Natl  Acad  Sci  U  S  A 1999;96:8426-8431. 28. Boros-Major  A,  Bona  A,  Lovasz  G  et  al.  New  perspectives  in  biomolecular  paleopathology  of  ancient  tuberculosis:  a proteomic approach. Journal of Archaeological Science 2011;38:197-201.  

Mini Review:  Dental Pulp as a Source for Paleomicrobiology

Hrčak - Portal of scientific journals of Croatia

     N g u y e n - H i e u  e t  a l .                                                  R E V I E W      Bull Int Assoc Paleodont. Volume 5, Number 1, 2011     www.paleodontology.com   48 Mini Review:  Dental Pulp as a Source for Paleomicrobiology  • Tung Nguyen-Hieu, Gérard Aboudharam, Michel Drancourt •  Unité de Recherche sur les Maladies Infectieuses et Tropicales Emergentes, UMR CNRS 6236, IRD198, IFR48, Faculté de médecine, Université de la Méditerranée, Marseille, France  Address for correspondence: Nguyen Hieu Tung  Unité de Recherche sur les Maladies Infectieuses et Tropicales Emergentes Faculté de médecine, Université de la Méditerranée Marseille, France E-mail: nghtung2183@yahoo.fr    Bull Int Assoc Paleodont. 2011;5(1):48-54.  Abstract Paleomicrobiology is a recent science. Paleomicrobiology describes the history of infectious diseases through research of ancient microbes. The dentistry with the tooth in particular has contributed to the progress of this science. The dental pulp is very well protected in the centre of the tooth. Molecular techniques such as “suicide PCR” and Multiple Spacer Typing have identified and characterized micro-organisms in the ancient samples. The detection of bacterial DNA from ancient dental pulp  provided the scientific evidences for diagnosis of the infectious diseases of the past. The cooperation of paleomicrobiology and paleodontology  also contributed the new knowledge to human pathology.   Keywords: Paleomicrobiology; Ancient Teeth; Dental Pulp; Microbial Detection  Introduction In 1993, after the first molecular detection of Mycobacterium tuberculosis in ancient skeletons (1), paleomicrobiology  appeared as a new discipline for the identification and characterization of microorganisms in the ancient samples. Frozen tissues, mummies, skeletons or bones were the ancient remains often used in paleomicrobiological studies. However, the major limitations of the ancient DNA (aDNA) study consist in the degradation and fragmentation of aDNA, contamination of modern DNA or external sources. Moreover, aDNA extraction from mummified tissue and bones is complicated because rehydration and decalcification are necessary steps. Therefore, another  source of ancient specimens that possibly limited the disadvantages above, were proposed: dental pulp (2), Figure 1. Vascularization of this organ is important, proportionally comparable with the human brain (3). Pathogenic bacteria circulating in blood may occur and colonize the dental pulp. A guinea-pig      N g u y e n - H i e u  e t  a l .                                                  R E V I E W      Bull Int Assoc Paleodont. Volume 5, Number 1, 2011     www.paleodontology.com   49 model has confirmed the possibility to detect bacterial DNA in dental pulp after experimental infection by Coxiella burnetii. This experimental model has also confirmed that pathogenic bacteria in blood had penetrated and colonized the dental pulp and that could then be used for diagnosis of bacteremia (4,5). Moreover, Bartonella henselae DNA and Bartonella grahamii DNA had been isolated from the dental pulp of the cats that had been bacteremic for Bartonella sp. then euthenased and buried one year previously, according to French army veterinary records (6). These results indicate that dental pulp was an excellent specimen for diagnosis of ancient bacteremia.  Figure 1 Ancient DNA recovered from dental pulp.  Paleomicrobiology of the dental pulp: methods In 1998, a team of researchers in France had first introduced dental pulp collected from ancient teeth as a suitable tool for the diagnosis of ancient infectious diseases (7). The advantages of using dental pulp for paleomicrobiological study is its better preservation in a closed cavity.  In the closed cavity, the soft tissues are preserved from environmental contamination and attacks of degrading agents.  Moreover, the DNA extraction from soft tissues of the dental pulp is simple (2). An experimental animal model infected with Coxiella burnetii had also confirmed the possibility of finding bacterial DNA in dental pulp 20 days after bacteremia (4). A second guinea-pig model infected with Coxiella burnetii showed persistence of the bacteria in the dental pulp (5). The pulp can then be used as a tool for retrospective diagnosis when there was a bacteremia. Afterwards, this team  proposed the “suicide PCR” for molecular identification of bacterial DNA from ancient teeth with strict criteria to avoid any risk of contamination and Multiple Spacer Typing for the genotyping of ancient bacteria (8). The important criteria of “suicide PCR” included: 1- The primers are used only once, 2-There were no positive controls, 3-The negative controls should be added, 4-Negative results are tested by other primers, 5-Positive results are confirmed by sequencing (9). The suicide PCR principle was used by another Greek team in 2006. This team using dental pulp has shown that the plague of Athens was in      N g u y e n - H i e u  e t  a l .                                                  R E V I E W      Bull Int Assoc Paleodont. Volume 5, Number 1, 2011     www.paleodontology.com   50 fact  typhoid fever (10). Rapid Diagnostic Test was an immunodetection method developed by another team for special diagnosis of plague from ancient remains  (11). From 1998 to the present day, several independent teams using these methods,  detected multiple pathogens from ancient dental pulp to confirm the epidemics of the past, Table 1.  Table 1 Microorganisms detected from ancient teeth Microorganisms Infectious diseases References Anelloviridae Virus 25 Bartonella henselae Cat-scratch disease 24 Bartonella quintana Trench fever 17-19;21;22 Mycobacterium leprae Leprosy 23 Mycobacterium tuberculosis Tuberculosis 23 Rickettsia prowazekii Typhus 21;22 Samonella enterica serovar Typhi Typhoid fever 10 Yersinia pestis Plague 7-9;11;13-18   Paleomicrobiology of the dental pulp: contributions In history, the three plague pandemics were cited as humanity’s disaster, in which an epidemic known as Black Death  occurred in 1347 and then spread from the Caspian Sea to almost all European countries. Over the next few years, Black Death had killed an estimated 17 – 28 million Europeans, representing approximately one third of the population at the time (12). In 1998, Yersinia pestis DNA had first been  identified in dental pulp of the ancient teeth collected from skeletons excavated from 16th and 18th century graves of persons suspected of having died of plague  (7). Genotyping had then yielded Y. pestis Orientalis strain  (8,13). These results were later reinforced by other independent teams to confirm the presence of plague in Medieval Europe  (11,14-16). Interestingly, two recent studies detected co-infection of the body louse,  Bartonella quintana, along with Yersinia pestis in ancient dental pulp, suggesting that the body louse  may have been a vector for interhuman transmission of medieval plague pandemics  (17-18). Bartonella quintana is the causative agent of trench fever disease and transmitted by lice, fleas and ticks to humans. Molecular detection of B. quintana in a 4000-year-old human tooth (southeastern France) indicated that B. quintana bacteremia had occurred in prehistoric humans 19. Typhus epidemic had been demonstrated in the 20th century as “a best friend” of wars. In the past, this epidemic spread rapidly in Europe from Spain to France and Italy following the wars between Emperor Charles V and King François I (20). The city of Douai in northern France was besieged from 1710 to 1712 during the war of Spanish succession when France and Spain opposed the other nations. The causative agent of typhus, Rickettsia prowazekii strain Madrid E type B, and B. quintana DNA had been isolated from ancient dental pulp of the individuals buried in Douai. This molecular detection      N g u y e n - H i e u  e t  a l .                                                  R E V I E W      Bull Int Assoc Paleodont. Volume 5, Number 1, 2011     www.paleodontology.com   51 allowed the researchers to confirm the presence of the typhus epidemic in Douai from 1710 – 1712 (21). Typhus was later a constant companion of Napoleon’s army in all European wars. In 2006,  evidence of louse-transmitted diseases in soldiers of Napoleon’s Grand Army in Vilnius was reported when R. prowazekii and B. quintana DNA had been amplified by “suicide PCR” from dental pulp collected from the remains of the soldiers. These results showed that louse-borne infectious diseases had affected nearly one-third of Napoleon’s soldiers buried in Vilnius and suggested a possible cause of the French retreat from Russia  (22), Figure 2 and 3. Mycobacterium tuberculosis and Mycobacterium leprae had also been detected in ancient teeth recovered from a 1st century tomb in Jerusalem, Israel. The findings had suggested the co-infection of these two mycobacteria in ancient human populations (23). Moreover DNA of Samonella enterica serovar Typhi had been successfully identified in dental pulp of ancient teeth collected from Kerameikos cemetery. This discovery had demonstrated that typhoid fever could be the cause of the plague of Athens in 430 – 426 BC (10). Interestingly, molecular detection of Bartonella henselae in dental pulp of 800-year-old cats suggested the occurrence of Cat-scratch disease within a medieval population in France because archaeological and historical data indicated that these cats had lived in close contact with people (24). A recent study had surprisingly reported the detection of an ancient virus (Anelloviridae DNA) from 200-year-old dental pulp recovered from ancient teeth of Napoleon’s soldiers in Kaliningrad, Russia (25).    Figure 2 and 3 Vilnius, 1812, Pictures of Yann ARDAGNA (UMR 6578 CNRS Université de la Méditerranée-CNRS-EFS        N g u y e n - H i e u  e t  a l .                                                  R E V I E W      Bull Int Assoc Paleodont. Volume 5, Number 1, 2011     www.paleodontology.com   52 Perspectives X-rays were used to detect and locate the target teeth in the jaws (Figures 4 and 5). The ancient tooth was usually bisected longitudinally for the recovery of dental pulp (Figures 6, 7 and 8). This method led to the destruction of its morphological structure and possible external contamination . “Orthograde entrance technique” is an “original method” to collect dental pulp with morphological conservation of ancient teeth that will be used afterwards by anthropologists  (26).  However, this technique has some disadvantages due to the relatively fragile, permeable and weak nature of the tooth root. The first method is probably the simplest and most effective despite the disadvantage of morphological changes of the tooth, it is necessary to restore the tooth by bonding techniques. The ancient proteins seem to be more resistant than aDNA. Lipids such as mycolic acids of M. tuberculosis also appear to be particularly robust in ancient remains (27). Mass spectrometry had successfully identified specific bacterial proteins  of M. tuberculosis from archeological bone samples (28). Therefore, this method, detecting microbial non-nucleotidic biomolecules from ancient dental pulp, may hereafter become a promising approach to complement aDNA analysis in paleomicrobiology. In conclusion, detection of pathogen traces from ancient teeth by using many paleomicrobiological methods has provided the scientific proofs to elucidate several infectious diseases in the past. This great cooperation of paleomicrobiology and paleodontology has also contributed  new knowledge of human pathology as well as  evidence of human history.    Figure 4 Mandible of a child about 9 years (Observance massgrave-Marseille 1724)  Figure 5 Radiography of the mandible and choice of the tooth (34 non-eruptive)       N g u y e n - H i e u  e t  a l .                                                  R E V I E W      Bull Int Assoc Paleodont. Volume 5, Number 1, 2011     www.paleodontology.com   53   Figure 6 Realization of the fracture using a carborundum disc on handpiece  Figure 7 Recovery of the dental pulp   Figure 8 After the fracture and recovery of the pulp, tooth is ready to be glued  References 1. Spigelman M, Lemma E. The use of the polymerase chain reaction to detect Mycobacterium tuberculosis in ancient skeletons. Int J Osteoarchaeol 1993;3:143. 2. Drancourt M, Raoult D. Palaeomicrobiology: current issues and perspectives. Nat Rev Microbiol 2005;3:23-35. 3. Kim S. Regulation of pulpe blood flow. J Dent Res 1985;64:590-596. 4. Aboudharam G, Lascola B, Raoult D, Drancourt M. Detection of Coxiella burnetii DNA in dental pulp during experimental bacteremia. Microb Pathog 2000;28:249-254. 5. Aboudharam G, Drancourt M, Raoult D. Culture of C. burnetii from the dental pulp of experimentally infected guinea pigs. Microb Pathog 2004;36:349-350. 6. Aboudharam G, Vu DL, Davoust B, Drancourt M, Raoult D. Molecular detection of Bartonella spp. in the dental pulp of stray cats buried for a year. Microb Pathog 2005;38:47-51. 7. Drancourt M, Aboudharam G, Signoli M, Dutour O, Raoult D. Detection of 400-year-old Yersinia pestis DNA in human dental pulp: an approach to the diagnosis of ancient septicemia. Proc Natl Acad Sci U S A 1998;95:12637-12640. 8. Drancourt M, Roux V, Dang LV et al. Genotyping, Orientalis-like Yersinia pestis, and plague pandemics. Emerg Infect Dis 2004;10:1585-1592.      N g u y e n - H i e u  e t  a l .                                                  R E V I E W      Bull Int Assoc Paleodont. Volume 5, Number 1, 2011     www.paleodontology.com   54 9. Raoult D, Aboudharam G, Crubezy E, Larrouy G, Ludes B, Drancourt M. Molecular identification by "suicide PCR" of Yersinia pestis as the agent of medieval black death. Proc Natl Acad Sci U S A 2000;97:12800-12803. 10. Papagrigorakis MJ, Yapijakis C, Synodinos PN, Baziotopoulou-Valavani E. DNA examination of ancient dental pulp incriminates typhoid fever as a probable cause of the Plague of Athens. Int J Infect Dis 2006;10:206-214. 11. Bianucci R, Rahalison L, Massa ER, Peluso A, Ferroglio E, Signoli M. Technical note: a rapid diagnostic test detects plague in ancient human remains: an example of the interaction between archeological and biological approaches (southeastern France, 16th-18th centuries). Am J Phys Anthropol 2008;136:361-367. 12. Drancourt M, Raoult D. Molecular insights into the history of plague. Microbes Infect 2002;4:105-109. 13. Drancourt M, Signoli M, Dang LV et al. Yersinia pestis Orientalis in remains of ancient plague patients. Emerg Infect Dis 2007;13:332-333. 14. Bianucci R, Rahalison L, Peluso A et al. Plague immunodetection in remains of religious exhumed from burial sites in central France. Journal of Archaeological Science 2009;36:616-621. 15. Haensch S, Bianucci R, Signoli M et al. Distinct clones of Yersinia pestis caused the black death. PLoS Pathog 2010;6. 16. Wiechmann I, Grupe G. Detection of Yersinia pestis DNA in two early medieval skeletal finds from Aschheim (Upper Bavaria, 6th century A.D.). Am J Phys Anthropol 2005;126:48-55. 17. Tran TN, Le Forestier C, Drancourt M, Raoult D, Aboudharam G. Co-detection of Bartonella quintana and Yersinia pestis in an 11th - 15th burial site, Bondy, France. Am J Phys Anthropol 2011. 18. Tran TN, Signoli M, Fozzati L, Aboudharam G, Raoult D, Drancourt M. High throughput, multiplex pathogen detection authenticates plague waves in medieval Venice, Italy. PLoS One 2011;6:e16735. 19. Drancourt M, Tran-Hung L, Courtin J, Lumley H, Raoult D. Bartonella quintana in a 4000-year-old human tooth. J Infect Dis 2005;191:607-611. 20. Raoult D, Woodward T, Dumler JS. The history of epidemic typhus. Infect Dis Clin North Am 2004;18:127-140. 21. Nguyen-Hieu T, Aboudharam G, Signoli M, Rigeade C, Drancourt M, Raoult D. Evidence of a louse-borne outbreak involving typhus in Douai, 1710-1712 during the war of Spanish succession. PLoS One 2010;5:e15405. 22. Raoult D, Dutour O, Houhamdi L et al. Evidence for louse-transmitted diseases in soldiers of Napoleon's Grand Army in Vilnius. J Infect Dis 2006;193:112-120. 23. Matheson CD, Vernon KK, Lahti A et al. Molecular exploration of the first-century Tomb of the Shroud in Akeldama, Jerusalem. PLoS One 2009;4:e8319. 24. La VD, Clavel B, Lepetz S, Aboudharam G, Raoult D, Drancourt M. Molecular detection of Bartonella henselae DNA in the dental pulp of 800-year-old French cats. Clin Infect Dis 2004;39:1391-1394. 25. Bedarida S, Dutour O, Buzhilova AP, de MP, Biagini P. Identification of viral DNA (Anelloviridae) in a 200-year-old dental pulp sample (Napoleon's Great Army, Kaliningrad, 1812). Infect Genet Evol 2010. 26. Alakoc YD, Aka PS. "Orthograde entrance technique" to recover DNA from ancient teeth preserving the physical structure. Forensic Sci Int 2009;188:96-98. 27. Poinar HN, Stankiewicz BA. Protein preservation and DNA retrieval from ancient tissues. Proc Natl Acad Sci U S A 1999;96:8426-8431. 28. Boros-Major A, Bona A, Lovasz G et al. New perspectives in biomolecular paleopathology of ancient tuberculosis: a proteomic approach. Journal of Archaeological Science 2011;38:197-201.  

Mini Review: Dental Pulp as a Source for Paleomicrobiology

Abstract

Similar works

Full text

Available Versions

Directory of Open Access Journals

Hrčak - Portal of scientific journals of Croatia