The Good Housekeeper: GBV-C Co-infection with HIV

I know that the study of infectious diseases seems grim. Bacteria are increasingly becoming antibiotic resistant, baby parasites nest in your brain and tuberculosis spreads to your toes. I get it – all this devotion to the study of what gruesome-thing-du-jour is in your blood and saliva isn’t sunshine, puppies and rainbows. So in the interests of maintaining public optimism, I offer you GBV-C, a virus that has been found to offer a protective, antiviral effect against HIV infection. Yes, that HIV.

GBV-C is a member of the Flaviviridae family of viruses; a truly distinguished and deadly lot that includes hepatitis C, yellow fever, West Nile virus, dengue and a slew of nasty encephalitis-causing viruses that roost in mosquitoes and ticks. GBV-C, however, appears to exist within the body as a benign, harmless infection. It provokes no identifiable clinical symptoms of disease and seems to be the black sheep of the noxious Flaviviridae family (2).

The one thing that makes this virus notable, even outstanding!, is that for those who are co-infected with HIV and GBV-C have been shown to live significantly longer than those without GBV-C (3). If HIV is an unwelcome houseguest in the human body, then infection with GBV-C is like waking up one day and unexpectedly finding a housekeeper doing the dirty dishes and containing the mess created by this destructive guest.

A model showing the morphology of the GBV-C virus. The human body’s antibody to the E2 envelope glycoprotein may assist in stymieing HIV infection. Source: the Physician Research Network. Click for source.

GBV-C is spread in ways similar to the transmission of HIV: through sex, birth and blood. Human infection with this virus is common, with 1–8% of healthy blood donors showing evidence of infection worldwide (1). So common in fact that isolated indigenous populations in Papua New Guinea have been found to harbor the virus (3). The global distribution and evolution of the major genotypes of this virus echoes human migration patterns, indicating that this is one old viral dude that’s been hitchhiking along with humans for millions of years (3).

GBV-C provides a beneficial, antiviral effect by stymieing HIV’s advances into immune cells and spurning its replication efforts in the body. It does so in a dizzying number of ways (reader beware: it’s gonna get molecular up in here). The infection stimulates messenger signals known as cytokines to activate a cellular response and consume HIV infected cells (otherwise known as the T helper 1 (Th1) cellular response system). The virus also encourages the innate immune response by boosting pathogen-fighting interferons; this innate response can be considered one of the “first responders” of the immune system as it responds to nearly every type of infection while also activating the more advanced components of the immune system to respond to any bacterial/viral/protozoal invaders.

GBV-C stops the human cells that HIV infects, CD4+ T cells, from killing themselves in the programmed-cell-death process known as apoptosis. Apoptosis is induced by HIV-infected T cells and can affect uninfected and infected cells alike; GBV-C protects these cells maintaining their population levels. This discovery is particularly noteworthy as a crash in the population of CD4+ T cells (“CD4+ counts”) is a hallmark of immune deficiency and disease progression from HIV to AIDS.

On the left, a T cell is covered in budding HIV virus particles and will soon undergo programmed-cell-death or apoptosis. On the right, a close-up of the budding virions. Infection with GBV-C may halt this process and prevent a population crash in the numbers of CD4+ cells. Image source: Roingeard P & Brand D (1998) Budding of Human Immunodeficiency Virus. N Engl J Med. 339(32). Click for source.

The GBV-C virus causes a decrease in the cellular presence of the receptors that HIV uses to gain entry into the cell (in science speak: there’s a down-regulation of the CCR5 and CXCR4 chemokine receptors); this is akin to temporarily removing the doors of your house to predators and thieves instead of just locking them shut. In addition, GBV-C infection seems to decrease the presentation of special “activation markers” on the surface of  T cells, interfering with signaling pathways that activate the production of T cells (2). Unfortunately, it’s not entirely clear what effect this “reduced immune activation” has on the immune system and whether this assists or hinders the immune system’s response to HIV infection. Certain proteins produced by GBV-C also inhibit HIV replication (1)(3).

Overall, co-infection with the two viruses leads to increased survival of HIV-infected patients and decreased mortality (1). Not too shabby! How it does this is largely uncertain – is it due to the presence of GBV-C virus particles (known as viremia) or the body’s response to GBV-C infection and its concomitant production of antibodies against the GBV-C virus (namely the anti–GBV-C envelope glycoprotein (E2) antibody)? The fact that 15-43% of HIV-positive people show active GBV-C infection and that another 31-55% show evidence of previous infection makes it difficult to disentangle these two questions (1). However, by all accounts, what we have here looks like a symbiotic relationship between GBV-C and humans.

Two researchers behind most of the work into GBV-C have suggested that this delightful house keeper may more accurately be called the “good boy virus”, a phrase that heartily captures its place in a HIV-positive body (2). The Flaviviridae family needs all the good PR it can get and this GBV-C do-gooder might be the trick.

Though it’s unlikely HIV physicians are going to start recommending GBV-C infection to HIV-infected patients anytime soon, the work of this benign virus has intriguing implications for future virology research. Study of the virus’s housekeeping efforts may yield new targets for anti-HIV medications and alternative avenues for future research. Perhaps this virus could even be genetically-modified to magnify its HIV-fighting ways and be employed as a form of HIV treatment? We already do the opposite of this with many virulent pathogens, using weak or killed viruses as vaccines, and there are physicians out there that advocate for the use of bacteria-killing viruses; using bacteriophages to fight stubborn infections was an especially popular treatment in Eastern Europe and the former Soviet Union in the 1950s (4). In the face of mounting antibiotic resistance across the globe, using do-gooder viruses like GBV-C may soon be the only option we have left.

Resources

The United State’s FDA does not screen donated blood for GBV-C. Dozens of studies have failed to find any link whatsoever between GBV-C infection and disease and, as such, an estimated 1000 Americans a day receive blood with GBV-C virions or antibodies (2). I covered blood screening for infectious diseases in this article here.

The Phage Therapy Center in the Republic of Georgia is the pioneer in providing bacteriophage treatment for a smorgasbord of bacterial infections.

A 2006 review looks at natural resistance to HIV in certain individuals, whether that be from GBV-C, the host immune response or host genetics. It’s free!

References
1. Xiang J et al. (2004) Inhibition of HIV-1 replication by GB virus C infection through increases in RANTES, MIP-1, MIP-1, and SDF-1. Lancet. 363(9426): 2040-6
2. Bhattarai N & Stapleton JT. (2012) GB virus C: the good boy virus? Trends Microbiol. 20(3):124-30
3. Polgreen PM et al. (2003) GB virus type C/hepatitis G virus: a non-pathogenic flavivirus associated with prolonged survival in HIV-infected individuals. Microbes Infect. 5(13): 1255–1261
4. Sulakvelidze A et al (2001) Bacteriophage Therapy. Antimicrob. Agents Chemother. 45(3): 649-659. Accessible here.

Polgreen, P., Xiang, J., Chang, Q., & Stapleton, J. (2003). GB virus type C/hepatitis G virus: a non-pathogenic flavivirus associated with prolonged survival in HIV-infected individuals Microbes and Infection, 5 (13), 1255-1261 DOI: 10.1016/j.micinf.2003.08.006

Chronicle of a Death Foretold: Human Sentinels for Disease Outbreaks

The bodies of dead crows were found littered through back yards, overgrown meadows, and public parks. We like to attach meanings to events, to craft symbols out of the mysterious and unknown. Birds, in particular, are favorite auguries of ours. These mass crow die-offs were an ugly, sinister sign – what could this mean, mass deaths of creatures customarily seen as living omens of death and the plague?

Unusual as it is, dead animals dropping from the sky isn’t uncommon. But the crows were different. In the summer of 1999, the massive die-off of Corvus brachyrhynchos heralded the entry of West Nile Virus (WNV) into the northeastern United States and its ensuing epidemic. Shortly after reports of clusters of dead crows crept into local health departments, news of a curious encephalitis sending many people to the emergency rooms in New York, New Jersey and Connecticut started popping up.

A flock of dead crows found in the state of Jharkhand, India in December of 2011. Crows can be susceptible to many avian viruses that threaten humans; in this case, these crows tested positive for H5N1 infection. Click for image source.

It is presumed that WNV had arrived from Israel, incubating in mosquito eggs hidden in rubber tires destined for transport to the United States. Local birds were soon infected by these foreign mosquitoes, then local mosquito populations acquired the virus by feeding on infected birds and soon the infection was introduced to Americans enjoying their summer outdoors. Now Americans had a novel type of sentinel, joining the sun, the sizzling barbecue and humid nights: when WNV sends people to emergency rooms around the country, we have unequivocal proof that summer has arrived.

Certain species of animals are sentinels for particular viral diseases; clusters of their deaths are red flags for emerging outbreaks and bioterrorism events that may soon spread to humans. The use of animals for this purpose can go beyond the canary in the coal mine, serving as bright flashes of information in uncertain times. Strangely acting horses can alert veterinarians to eastern equine encephalitis and Hendra virus, two very nasty viruses that are fatal to both humans and horses. Gorilla families hemorrhaging to death in Uganda can portend the emergence of Ebola in a nearby village. When pigs in Malaysia and Bangladesh are found coughing, slumped in mud, Nipah virus might soon make its leap into humans.

Depending on their profession, humans can be sentinels for outbreaks too. Many people ought to consider themselves lucky to work at a desk, rather than working face-to-face everyday with dozens of strangers or animals. When clusters of emergency room physicians and nurses, EMTs, or veterinarians multiply fall ill from odd illnesses, the CDC sits up and pays attention.

A local Red Cross group in Kikwit, Zaire in 1995 responding to the Ebola epidemic following the deaths of many hospital personnel. Click for image source.

In Zaire in 1995, entire teams of medical professionals, from hospital technicians to nurses to physicians, succumbed to the newly emerging Ebola virus; over 70% of the initial cases were nosocomial, hospital-borne infections among medical personnel (1). Since 1994, a series of sporadic outbreaks of the respiratory Hendra virus among horses have infected horse trainers and veterinarians in Australia, killing at least three people (2). The H1N1 pandemic in the spring of 2009 infected over 25% of medical professionals working in both the adult and pediatric emergency departments, significantly eclipsing the infection rates of other hospital departments (3). Most recently, the measles epidemic in New Zealand last fall spread to ambulance workers; a quarantine was implemented in one company to prevent spread to future patients and medical staff (4). An amplification of an infectious disease among medical professionals working on the front-lines of medical care is, unluckily, just another occupational hazard.

What other occupations might find themselves unintentionally waving this epidemiological flag? Flight attendants busily crossing the globe on international flights, handing out meager rations of peanuts to travelers. Bartenders and waitresses hurrying away in busy restaurants in major urban cities. Bushmeat hunters searching for wild game and loggers working in old-growth jungle and rain-forests, exposing themselves to fleeing animals and mosquitoes harboring unknown viruses. Hustling prostitutes. Teachers in elementary schools fending off the sticky hands of their students.

Hundreds of pigs culled during the Nipah epidemic in Malaysia in 1999. Image: BBC News Online. Click for image source.

This concept of human sentinels is crucial in light of the changing landscape of of diseases today – SARS, Nipah virus and Hendra virus are all diseases that have emerged as significant public health threats only in the past few decades. Zoonotic of origin, these guys love to flip back and forth between humans and wild and domestic animals in their natural transmission cycle. In a sense, outbreaks in certain occupations that traverse the line between the domesticated world and the wild can serve as a “yardstick” for measuring the health and stability of their surrounding ecosystem, an indicator of environmental disruption that encourages transmission of zoonotic diseases (5).

That wild and domestic animals may announce the onset of a zoonotic outbreak in humans is the very reason that public health agencies long for close working relationships between veterinarians and local health agencies. Shaping an effective disease surveillance program requires looking two-ways, at diseases and deaths in both humans and animals. Thankfully, we have a few existing groups that can pinpoint the origins of emerging diseases – the ProMed emails alerts of unusual disease incidences and outbreaks, the One Health initiative looks at human, animal and environmental health, along with the emerging viral research by the Global Viral Forecasting Initiative. Instead of looking to the skies for signs, we can take an interdisciplinary approach to disease surveillance by looking around us. Check out the resources below for some of the organizations doing just that.

RESOURCES

Automated Epidemiologic Geotemporal Integrated Surveillance System or AEGIS is a syndromic surveillance system for the Massachusetts Department of Public Health. It performs automated, real-time surveillance for bioterrorism and naturally occurring outbreaks.

The Global Viral Forecasting Initiative, brainchild of virologist extraordinaire Nathan Wolfe, is developing a global early warning system to prevent novel pandemics. By monitoring emerging viruses in humans and animals, the Initiative can molecularly track the development of novel zoonotic viruses. Super neat stuff.

Promed Mail sends email alerts on the latest infectious diseases and epidemics around the world.

The Global Avian Network for Surveillance or GAINS surveillance system has an electronic database of over 100,000 birds that monitors avian health worldwide, specifically the transmission of flu viruses among wild birds.

The Canary Database is an online database for biomedical research examining animals as sentinels of zoonotic, environmental, and toxic effects that are applicable to human health. It is pretty thorough and a great reference site. I’ve certainly had my fun tooling around on it.

Additionally, read this rather upsetting article from the Journal of Infectious Diseases describing the Ebola outbreak in Zaire and the difficulties in diagnosing the disease as, one by one, medical personnel died from the hemorrhagic fever.

REFERENCES

(1) Waterman T (1999) Ebola Zaire Outbreaks. Tara’s Ebola Site  [Accessed March 23, 2012]
(2) WHO (July 2009)  Hendra Virus Fact sheet N°329 WHO Media Centre [online] Available here. [Accessed Mar 20, 2012.]
(3) One News/NZN. (28/9/2011) Measles spreads to ambulance staff. ONE News/ NZN  [online] Available here. [Accessed Mar 20, 2012.]
(4) Santos CD, Bristow RB, Vorenkamp JV. (2010) Which health care workers were most affected during the spring 2009 H1N1 pandemic? Disaster Med Public Health Prep. 4(1):47-54
(5) Cook A, Jardine A and Weinstein P. (2004) Using Human Disease Outbreaks as a Guide to Multilevel Ecosystem Interventions. Environ Health Perspect. 112(11):  1143-46

Cook, A., Jardine, A., & Weinstein, P. (2004). Using Human Disease Outbreaks as a Guide to Multilevel Ecosystem Interventions Environmental Health Perspectives, 112 (11), 1143-1146 DOI: 10.1289/ehp.7122

Nobel Prizes, Tropical Medicine & One Nazi Sympathizer

Nobel Prizes! We all want one, don’t we? While fantasizing about heavy gold medallions and the Swedish Nobel Assembly, I wondered how many of the Nobel Laureate prizes in Physiology and/or Medicine have gone towards scientists studying infectious diseases, immunology and the tropical medicine field. Snooze button alert, am I right? This is the product of a one-track mind so you have my apologies. But! If it’s any consolation, there’s a story hidden in this article of a Nobel Laureate Nazi sympathizer that infected mental patients with malaria to cure them of their psychoses. Science!

Let’s get down to numbers. As of 2011, this prize has been awarded 101 times to 199 scientists for their research in this field.  In recent years, the Nobel Prize has often been awarded jointly to two or three scientists for either collaborative research or for research that is similar in scope and specialty. Of those 101 occasions, 22 of these awards have been given to 39 researchers for their work in infectious diseases, virology, parasitology and/or immunology. Only one woman is on this list, Françoise Barré-Sinoussi, for her incredible work with Luc Montagnier in identifying HIV at the height of the HIV/AIDS epidemic in the 1980s.

The Nobel Medal for Physiology or Medicine is embossed with the Genius of Medicine "holding an open book in her lap, collecting the water pouring out from a rock in order to quench a sick girl's thirst". Description from NobelPrize. Click for image source.

(FYI, there have been nine years in which the Nobel Prize ceremony and awards did not occur due to the disruption of the two World Wars – in 1915 to 1918, 1921, 1925 and 1940 to 1942. Those Nazi’s again!)

So roughly one-fifth of the Nobel Prizes in medicine have gone to scientists making breakthroughs in the study of infectious diseases or in the immunological response to those diseases, indicating how crucial this particular field of medicine is to the scientific community. We’re not talking dermatology or podiatry here, folks.

In the early years, the Assembly awarded prizes for game-changing advances in the research of big killer diseases of the day – diptheria, tuberculosis and malaria, to name a few. The development of a yellow fever vaccine (1951), discoveries of vital antibiotics that are still used today globally (1939, 1945 and 1952), and the finding that HPV causes cervical cancer (2008) are just a few of the discoveries that the Nobel Assembly considered so revelatory to the field of medicine. Until the past century, how infectious diseases entered the body, their mechanisms of pathogenesis and even their treatment was largely unknown. Malaria is transmitted by mosquitoes? Not known till 1899 and Sir Ronald Ross’s discovery of the parasite hiding inside the stomach of an Anopheles mosquito was justly recognized by the Nobel Assembly in 1902. Lice transmit typhus? Charles Nicolle’s finding won him a good amount of money and worldwide recognition/obscurity in 1928. In 1954, a trio of American researchers were awarded the Nobel for identifying a method to culture the polio virus, launching the beginning of the end of polio’s affliction on humanity.

Many of the awarded discoveries laid the foundation for productive research that has led to profound advancements in disease and pest control, the emergence of the field of public health, development of the pharmaceutical industry and the industrial production of vaccines, antibiotics, and antivirals. Life-changing stuff, ya’ll. Flipping through this list is a reminder of how far we’ve come in the past century in establishing our domain over bacteria and viruses that previously wiped us out with shocking regularity. In fact, some of their findings and methods are so commonplace, hell! even banal, in today’s laboratories that it’s like stepping into a science time-warp reading about their discoveries. Thanks to these scientists, we possess the complex knowledge of genetics, immunology, antibiotics, vaccines and their respective methods to identify, culture and combat new diseases.

The list of winners is below and includes the year they were awarded, their country of origin and the award’s justification from the Nobel Assembly in quotes; those quotes come from the official website of the Nobel Prize. I’ve included in the list a few winners awarded for their work in immunology and antibiotics; while these are not strictly related to infectious diseases, their research laid a strong foundation for future infectious disease work and, well, they’re related and it’s important so get over it.

Scroll past this engrossing list that I’ve compiled for your disease-seeking pleasure.

1901
Emil Adolf von Behring (Germany) “for his work on serum therapy, especially its application against diphtheria, by which he has opened a new road in the domain of medical science and thereby placed in the hands of the physician a victorious weapon against illness and deaths”. Victorious weapon! I’d think the flowery language alone should do the trick. In any case, von Behring developed a diphtheria antitoxin, known as a “serum”, by repeatedly injecting a horse with diphtheria toxin.

"A Future Pharmacy". An editorial cartoon mocking Emil Adolf Von Behring and his discovery of serum therapy, in which a diphtheria antitoxin was developed by repeatedly exposing a horse to the diphtheria toxin. Click for source.

1902
Sir Ronald Ross (United Kingdom) for his discovery that malaria is transmitted by mosquitoes.

1905
Robert Koch (Germany) “for his investigations and discoveries in relation to tuberculosis”.

1907
Charles Louis Alphonse Laveran (France) “in recognition of his work on the role played by protozoa in causing diseases”.

1927
Julius Wagner-Jauregg (Austria) “for his discovery of the therapeutic value of malaria inoculation in the treatment of dementia paralytica”. In 1927, Wagner-Jauregg received the  Prize for his discovery of pyrotherapy, the treatment of mental disease by febrile diseases. Besides being a Nobel Laureate, Wagner-Jauregg was also a Nazi sympathizer, anti-semite and racial hygienist. A charmer of a man who forced sterilization upon the mentally ill and the criminal and the so-called “father of fever therapy”(1).

Mustachioed Wagner-Jauregg stands behind a mental patient receiving malarial pyrotherapy. Click for image source.

Since 1887, he had been investigating the phenomenon of how fever, as induced by tuberculosis or St. Anthony’s Fire, may reduce the effects of psychosis caused by tertiary syphilis. Bacterial infections didn’t seem to do the trick in terms of bringing about recurrent high fevers and so in 1917 Wagner-Jauregg turned to the malaria parasite. Turns out, malaria is an excellent specimen for this sort of thing and led to its usage as an established treatment for paralytic dementia and neurosyphilis. An application of quinine would end the fever therapy, eliminating the infection and leaving the patient free of his psychoses. For this hot discovery, Wagner-Jauregg is the only psychiatrist to have ever won the Nobel for investigative fieldwork on mental illness.

1928
Charles Jules Henri Nicolle (France) for his discovery that lice transmitted typhus. Nicholle was a master researcher of infectious diseases and made many discoveries regarding the pathology and epidemiology of brucellosis, leishmaniasis, measles, rinderpest, scarlet fever, Mediterranean spotted fever, toxoplasmosis, trachoma and tuberculosis. Nicholle’s typhus work was performed in Tunisia using chimpanzees and toque macaques (2).

Vials and tablets of prontosil, an industrial dye discovered to have antibacterial properties by Gerhard Domagk. Click for source.

1939
Gerhard Domagk (Germany) “for the discovery of the antibacterial effects of prontosil”. Prontosil was the first drug to be found effective against bacterial infections and Domagk made quick use of this fact – as a streptococcal infection threatened his daughter, his treatment with prontosil prevented the therapeutic amputation of her arm.

1945
Sir Alexander Fleming (United Kingdom), Sir Ernst Boris Chain (United Kingdom), and Howard Walter Florey (Australia) “for the discovery of penicillin and its curative effect in various infectious diseases”.

1951
Max Theiler Union (South Africa) “for his discoveries concerning yellow fever and how to combat it”. He had an enormous forehead. His award is the only one that has gone towards a virus vaccine.

1952
Selman Abraham Waksman (United States) for his discovery of streptomycin, the first antibiotic effective against tuberculosis”. Waksman coined the term “antibiotic”.

There's really no reason why I'm including this photo of Selman Waksman here aside from the fact that it's so gloriously Science!-y. Lab coat, check. Flask of something liquid, check. Mood lighting, check. Click for image source.

1954
John Franklin Enders, Thomas Huckle Weller and Frederick Chapman Robbins (all from the United States) “for their discovery of the ability of poliomyelitis viruses to grow in cultures of various types of tissue”. Their work laid the groundwork for Jonas Salk’s development of the polio vaccine.

1958
Joshua Lederberg (United States) “for his discoveries concerning genetic recombination and the organization of the genetic material of bacteria”.

1966
Peyton Rous (United States) “for his discovery of tumour-inducing viruses”. In 1911, Rous found that some cancers can be transmitted by viruses, known as retroviruses. Forty years later, he was awarded for his work.

1969
Max Delbrück, Alfred D. Hershey and Salvador E. Luria (all from the United States), “for their discoveries concerning the replication mechanism and the genetic structure of viruses”. The trio discovered that bacterial resistance to bacteriophages (viruses that infect bacteria) was a result of random mutations.

1972
Gerald M. Edelman (United States) and Rodney R. Porter (United Kingdom) used immunoglobins sourced from human blood to identify the chemical structure of antibodies.

1976
Baruch S. Blumberg and D. Carleton Gajdusek (United States) “for their discoveries concerning new mechanisms for the origin and dissemination of infectious diseases”. These guys were awarded for their research on kuru, a disease caused by human prions. Fun fact: Blumberg also discovered the hepatitis B virus and developed the diagnostic test and vaccine for it.

1984
Niels K. Jerne (Denmark), Georges J.F. Köhler (Federal Republic of Germany) and César Milstein (Argentina and the United Kingdom) “for theories concerning the specificity in development and control of the immune system and the discovery of the principle for production of monoclonal antibodies”.

1989
J. Michael Bishop and Harold E. Varmus (United States) “for their discovery of the cellular origin of retroviral oncogenes”.

1997
Stanley B. Prusiner (United States) “for his discovery of Prions – a new biological principle of infection”.

2005
Barry J. Marshall and J. Robin Warren (Australia) “for their discovery of the bacterium Helicobacter pylori and its role in gastritis and peptic ulcer disease”. Marshall is famously known for downing culture medium loaded with H. pylori to directly prove that the bacterium is responsible for causing ulcers.

2006
Andrew Z. Fire and Craig C. Mello (United States) “for their discovery of RNA interference, [otherwise known as] gene silencing by double-stranded RNA”.

2008
Harald zur Hausen (Germany) “for his discovery of human papilloma viruses causing cervical cancer”, along with Françoise Barré-Sinoussi and Luc Montagnier (France) “for their discovery of the human immunodeficiency virus”.

Resources

The Nobel Prize website has a fleshed-out timeline for every Laureate and includes their biography, a short article about their research, the lecture they delivered to the Nobel Assembly and much much more.

References
(1) Howes OD et al (2009) Julius Wagner-Jauregg, 1857–1940. Am J of Psy. 166(4): 409.
(2) Schultz MG and Morens DM. (2009) Charles-Jules-Henri Nicolle. Emerg Infect Dis. 15(9): 1520-22. Accessed online.

Schultz, M. (2009). Charles-Jules-Henri Nicolle Emerging Infectious Diseases, 1519-1522 DOI: 10.3201/eid1509.090891