Tag Archives: Françoise Barré-Sinoussi

To Resign over an Editorial Decision You Disagree With

What would you do if you were serving on the editorial board of a scientific journal which had just published a manuscript that you knew was seriously flawed. Moreover, you knew that publication of the manuscript might seriously undermine global public health? That was the circumstance of cell biologist Klaudia Brix, Professor of Cell Biology, Jacobs University Bremen, Germany, when, in 2011, the Italian Journal of Anatomy and Embryology (IJAE)—the official publication of the Italian Society of Anatomy and Histology—published a paper by infamous AIDS denialist, Peter Duesberg, which reiterated his already discredited argument that HIV (the human immunodeficiency virus) does not cause AIDS (1). Brix resigned in protest from the IJAE editorial board. But why is that noteworthy? Remarkably, she was, for a time, the only member of the journal’s 13-person editorial board to do so, despite other members having similar misgivings over the decision to publish the paper. Afterwards, Heather Young, an anatomy and neuroscience researcher at the University of Melbourne, likewise resigned from the IJAE editorial board. Here is the background to this state of affairs.

Peter Duesberg is not the only AIDS denialist. However, he has been the most infamous of the AIDS denialists. HIV is a retrovirus, and Duesberg is the only AIDS denialist who also happens to be an expert retrovirologist. In fact, Duesburg was at one time a highly esteemed retrovirologist. In 1985 he was elected to the U.S. National Academy of Sciences; mainly for his 1970 discovery, with Peter Vogt, of the first known retroviral oncogene—the Rous sarcoma virus v-src.

AIDS denialist, Peter Duesberg
AIDS denialist, Peter Duesberg

Duesberg first put forward his denialist view in a 1987 paper in Cancer Research (2), which asserted that AIDS results from drug abuse, parasitic infections, malnutrition, and antiretroviral drugs. In Duesberg’s assessment, HIV is just another opportunistic infection. He has maintained that view since then, despite overwhelming evidence to the contrary. Consequently, he is looked upon as a pariah by the scientific community.

Even though Duesberg’s denialist views have been rejected by AIDS experts, Duesberg’s standing as a retrovirologist enabled him to yet influence some public health officials. In 2000, Duesberg was serving on a panel advising Thabo Mbeki (President of South Africa after Nelson Mandela) on how to manage the South African AIDS outbreak. Although Mbeki was an able and intelligent leader, he accepted Duesberg’s denialist view that HIV was not the cause of the South African AIDS epidemic. Thus, Mbeki allowed the South African outbreak to get completely out of control (3). Two independent studies later concluded that over 300,000 South African AIDS deaths would not have occurred if the Mbeki government’s public health policy had not followed the denialist view. Many thousands of South African AIDS victims, including infants, would have been spared infection if the government had publicized that AIDS is an infectious disease, and if it had made antiretroviral drugs available, particularly to pregnant women (1).  See Asides 1 and 2.

[Aside 1: The reasons why Mbeki assented to Duesberg’s denialist view are not clear. One possibility is that Mbeki held strong anti-colonialist and anti-West sentiments—born of having come of age during South Africa’s apartheid era—which led him to see his country’s AIDS crisis as a means by which the West sought to exploit his nation. To that point, he may have doubted the efficacy of expensive antiretroviral drugs, which were available only from large Western pharmaceutical companies. Moreover, the cost of treating the 5 million or more HIV-infected South Africans with those drugs would have exceeded the annual health department budget of his poverty-stricken nation by a factor of ten. Mbeki did accept that AIDS is the consequence of a breakdown of the immune system. But he was inclined to believe (or at least claimed) that poverty, bad nourishment, and ill health, rather than a virus, led that breakdown; a stance that enabled him to justify treating poverty in general, rather than AIDS in particular. Duesberg defended Mbeki in his publications, denying that hundreds of thousands of lives were lost in South Africa because of the unavailability of anti-retroviral drugs. But in 2002, after Mbeki suffered political fallout from the consequences of having acceded to Duesberg’s views, he tried to distance himself from the AIDS denialists, and asked that they stop associating his name with theirs.]

[Aside 2: The 2000 International AIDS Conference was taking place in Durban (a city in the South African province of KwaZulu-Natal) at the same time that Mbeki’s AIDS panel was convening in Johannesburg. Consequently, the denialist views expressed by Mbeki’s panel were also being heard in Durban. This prompted the so-called “Durban Declaration,” signed by over 5,000 scientists and physicians, and published in Nature, which proclaimed that the evidence that HIV causes AIDS is “clear-cut, exhaustive and unambiguous”.]

Well before Duesberg submitted his paper to IJAE, the arguments put forward in the paper had already been appraised and rebuffed by the scientific community. Indeed, the paper had previously been rejected by several other journals. The first submission was to the Journal of Acquired Immune Deficiency Syndromes (JAIDS), a peer-reviewed medical journal covering all aspects of HIV/AIDS. The JAIDS editors found that Duesberg’s contentions in the paper were based on a selective reading of the scientific literature, in which he dismissed all the vast evidence that HIV is the etiologic agent of AIDS. Not surprisingly, JAIDS rejected the paper, with one peer reviewer even warning that Duesberg and co-authors could face criminal charges if the paper were published.

After JAIDS rejected the paper, Duesberg  submitted a revised version to Medical Hypotheses (4). Like the original paper sent to JAIDS (as well as the version accepted by IJAE), the paper submitted to Medical Hypotheses contained data cherry-picked to cast doubt on HIV as the cause of AIDS. Nonetheless, Medical Hypotheses accepted the paper. However, the paper never went to press. But first, what was the explanation for the seemingly bizarre decision to accept the paper?

The answer laid in the fact that Medical Hypotheses was the only journal of its parent publisher, Elsevier, that did not use peer review; instead relying on its editorial board to select papers for publication. In any case, before the accepted paper went to press, prominent AIDS researchers, including Nobel laureate Francoise Barre-Sinoussi (co-discoverer that HIV is the cause of AIDS, 5), complained to Elsevier that the paper would have a negative impact on global healthcare, and requested that the paper be withdrawn.

Elsevier responded to these protests by asking the editors of another of its journals, The Lancet, to oversee a peer review of the paper. The Lancet editor sent the paper to five external reviewers, each of whom found that it contained numerous errors and misinterpretations, and that it could threaten global public health if it were published. Elsevier then permanently withdrew the paper.  Elsevier also instituted a peer-review policy at Medical Hypotheses (and fired the journal’s editor, who resisted the change).

The Medical Hypotheses incident resulted in more notoriety for Duesberg when the University of California, Berkley, where Duesberg is still a professor of molecular and cell biology, bought charges of misconduct against him for making false scientific claims in the paper, and for a conflict-of-interest issue. Apropos the latter, Duesberg did not reveal that co-author David Rasnick had earlier worked for Matthias Rath, a German doctor and vitamin entrepreneur, who sold vitamin pills as a therapy for AIDS. Duesberg was later cleared of both charges. But the next iteration of paper, to IJAE, did not respond to these allegations.

Duesberg regarded Elsevier’s actions as another example of “censorship” imposed by the “AIDS establishment.” Undeterred however, he submitted a revision of the paper to IJAE, which that journal then accepted, prompting Klaudia Brix and Heather Young to resign from that journal’s editorial board. The IJAE paper contained the same cherry-picked data and discredited assertions that were rejected earlier by JAIDS and Elsevier.  Moreover, publication of the paper still posed a threat to global public health. What then was behind the IJAE decision to publish?

Here is what happened. The paper was “peer-reviewed” by IJAE, but by only two reviewers; one of whom was Paolo Romagnoli, the IAJE editor-in-chief, who is neither a virologist or an epidemiologist but, instead, a cell anatomist. Consequently, the paper underwent only one external review, and there is no information regarding whether the lone external reviewer was an AIDS expert. One board member (who did not resign) later commented: “Only one [external] reviewer in my mind is not enough for manuscripts of a sensitive nature… (6)” [But this comment too is a bit troubling. Bearing in mind that the paper contained numerous errors and misinterpretations, would those have been okay if the paper were not of a “sensitive nature”?]

One also might ask why a journal that specialized in anatomy and embryology would consider a paper about the cause of AIDS. To that point; Klaudia Beix gave, as a reason for her resignation from the IJAE board, her belief that a journal should function within its scientific “scope” (6). So how did Romagnoli, the IJAE editor-in-chief, justify his decision to consider the paper?  He did so by asserting that it dealt with “issues related to the biology of pregnancy and prenatal development and with the tissues of the immune system (6).” But despite Romagnoli’s contention, the only mention of embryology in the paper was a short comment in the abstract: “We like to draw the attention of scientists, who work in basic and clinical medical fields, including embryologists, to the need of rethinking the risk-and-benefit balance of antiretroviral drugs for pregnant women, and newborn babies (1).”

As for Romagnoli’s reliance on only two reviewers, he justified that stance on the fact that the reviewers had concurring opinions. Moreover, he claimed that his criteria for selecting reviewers—apparently irrespective of their expertise—was to choose individuals (himself included) who he believed would not reject a paper merely because it challenged prevailing opinion.

But is there any possibility that Duesberg might be right? The answer is virtually none whatsoever. An earlier post noted: “…the evidence that HIV causes AIDS is, without exaggeration, overwhelming. Consider just the following. Data from matched groups of homosexual men and hemophiliacs show that only those who are infected with HIV ever develop AIDS. Moreover, in every known instance where an AIDS patient was examined for HIV infection, there was evidence for the presence of the virus. These data have been available for years, and Duesberg should have been aware of them. What is more, there has been the enormous success of antiretroviral therapy in changing AIDS from a nearly invariably fatal disease, into a manageable one, for many HIV-infected individuals (3).”

Even so, Duesberg is not regarded as a pariah by AIDS experts merely because his views concerning the connection between HIV and AIDS challenge accepted wisdom.  Instead, as asserted by Harvard University AIDS epidemiologist, Max Essex, Duesberg has sustained a “dangerous track of distraction that has persuaded some people to avoid treatment or prevention of HIV infection (6)”.

A scientist mounting a long-time challenge to the “establishment,” and being ridiculed for his views, before eventually being vindicated, makes for a very good story. However, such instances are very rare. Exceptions include Howard Temin (7) who hypothesized reverse transcription, and Stanley Pruisner (8) who hypothesized prions—infectious agents that contain no nucleic acid genome. Both researchers had to endure widespread ridicule for several years. But, and importantly, irrefutable evidence eventually accumulated to support their hypotheses. And, finally, both were awarded Nobel Prizes. But Duesberg has not been vindicated and, almost certainly, he  never will be.

References:

1. Duesberg PH, et al., 2011. AIDS since 1984: no evidence for a new, viral epidemic – not even in Africa. Italian Journal of Anatatomy and Embryololgy 116:73–92. http://fupress.net/index.php/ijae/article/view/10336/9525

2. Duesberg P, 1987. Retroviruses as carcinogens and pathogens: expectations and reality. Cancer Res. 47:1199–220. PMID3028606.

3. Thabo Mbeki and the South African AIDS Epidemic, Posted on the blog July 3, 2014.

4. Duesberg PH, et al., 2009. WITHDRAWN: HIV-AIDS hypothesis out of touch with South African AIDS – A new perspective. Medical Hypotheses. doi:10.1016/j.mehy.2009.06.024. PMID19619953.

5. Who Discovered HIV, Posted on the blog, January 24, 2014.

6.  Corbyn Z. 2012. Paper denying HIV–AIDS link sparks resignation: Member of editorial board quits as editor defends publication. Nature doi:10.1038/nature.2012.9926.

7. Howard Temin: “In from the Cold, Posted on the blog December 14, 2013.

8. Stanley Pruisner and the Discovery of Prions: Infectious Agents Comprised Entirely of Protein, Posted on the blog December 15, 2016.

 

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Harald zur Hausen, Papillomaviruses, and Cervical Cancer

Harald zur Hausen (1936- ) was awarded a share of the 2008 Nobel Prize in Physiology or Medicine for discovering that papillomaviruses cause cervical cancer. He received the award jointly with Luc Montagnier and Françoise Barré-Sinoussi, who were given their portion for discovering HIV (1). Before getting on with zur Hausen’s story per se, we begin with bit of earlier history.

zur hausen Harald zur Hausen in 2008

Genital warts are benign epithelial tumors that have been known and associated with sexual promiscuity since the time of the ancient Greeks. In 1907 these lesions were unequivocally proven to be an infectious disease by Italian researcher, G. Ciuffo, who showed that they can be transmitted by filtered extracts of wart tissue; a finding which also implied that the etiologic agent is a virus. Ciuffo inoculated himself to advance his case.

Ciuffo’s finding is relevant to our story since members of the papillomavirus family of DNA viruses are the cause of warts. What’s more, and importantly, some papillomaviruses  also cause malignant cervical carcinomas.

In 1933 Richard Shope, at the Rockefeller Institute, became the first researcher to isolate a papillomavirus, the cottontail rabbit papillomavirus. Shope went on to show that this virus is the cause of skin papillomas in its rabbit host. This finding by Shope was the first to demonstrate that a DNA virus can be tumorigenic.

Years earlier, in 1911, Peyton Rous discovered that an RNA virus—the Rous sarcoma virus (the prototype retrovirus)—causes solid tumors in chickens. Peyton Rous was Richard Shope’s friend and colleague at the Rockefeller Institute. In 1934 Shope asked Rous to characterize the warts that the rabbit papillomavirus induces in jackrabbits. Rous found those warts to be benign tumors that could progress to malignant carcinomas.

Despite the earlier findings of Ciuffo, Shope, and others, the notion that genital warts in humans is a sexually transmitted malady was slow to gain acceptance. Oddly, perhaps, recognition of that truth was prompted by a 1954 report that American servicemen, who had been serving in Korea, were transmitting genital warts to their wives upon returning to the U.S (T. J. Barrett, et al., J. Am. Med. Assoc. 154:333, 1954). [Sexually transmitted diseases were a long-standing problem in the military. Servicemen were most often infected by sex workers who frequented the vicinity of military quarters.]

The key discoveries of this tale are Harald zur Hausen’s 1983 and 1984 findings that two human papillomavirus subtypes, HPV-16 and HPV-18, together account for about 70% of all cervical cancers. Considering that more than 120 distinct HPV subtypes have been identified, the high degree of association of cervical carcinoma with only two of these subtypes provided compelling evidence for the viral etiology of this malignancy. Later studies showed that HPV-31, HPV-33, HPV-45, HPV-52, and HPV-58 are responsible for another 20% of cervical cancers. Indeed, an HPV infection is present in virtually all cervical carcinomas. See Aside 1.

[Aside 1: Cervical cancer was once the leading cause of cancer-related deaths in women in the United States. However, the number of cervical cancer deaths in the industrialized world decreased dramatically over the last 40 years, largely because of the Pap test, which can detect pre-cancer cervical lesions in their early stages. The CDC website reports 12,109 cervical cancer cases and 4,092 deaths from cervical cancer in the U.S. in 2011 (the most recent year for which data are available). Worldwide, cervical cancer was responsible for 275,000 deaths in 2008. About 88% of these deaths were in developing countries (J. Ferlay et al., Int. J. Cancer, 127:2893, 2010).]

Harald zur Hausen was a child in Germany during the Second World War, growing up in Gelsenkirchen-Buer, which was then a center for German coal production and oil refining and, consequently, a major target for allied bombing. [The city also contained a women’s sub-camp of the Buchenwald concentration camp. The Nazis used its prisoners for slave labor.] All members of zur Hausen’s family survived the war. However, zur Hausen’s primary education contained significant gaps because schools were closed during the allied bombing (2).

Despite the gaps in zur Hausen’s early education, he was keenly interested in biology and dreamed of becoming a scientist. Yet at the University of Bonn he opted to study medicine, rather than biology. After zur Hausen received his medical degree, he worked as a medical microbiologist at the University of Düsseldorf, where he enjoyed the opportunity that the University gave him to carry out research on virus-induced chromosomal aberrations.

Although zur Hausen was fascinated by his research, he was soon aware of the deficiencies in his scientific background. So, in 1966 he looked to enhance his proficiency as a scientist by securing a postdoctoral position in the laboratories of Gertrude and Werner Henle at the Children’s Hospital of Philadelphia.

The Henles were a German-born husband and wife research team, known for their work on flu vaccines. More apropos to our story, they are also known for demonstrating the link between the recently discovered Epstein-Barr virus (EBV; a herpesvirus) and infectious mononucleosis, as well as for showing that EBV is the etiologic agent of Burkitt’s lymphoma; a cancer found in parts of Africa. EBV was, in fact, the first virus associated with a cancer in humans. [Gertrude Henle’s mother was murdered by the Nazis in 1943.]

Although zur Hausen took part in the Henles’ experiments involving EBV, he did so grudgingly because he was intimidated by his inexperience in molecular biology. In his own words: “I urged Werner Henle to permit me to work with a different agent, namely adenovirus type 12, hoping that this relatively well established system would permit me to become acquainted with molecular methods. He reluctantly agreed. I started to work eagerly on the induction of specific chromosomal aberrations in adenovirus type 12-infected human cells…and, to please my mentor, I demonstrated electron microscopically the presence of EBV particles directly in… Burkitt’s lymphoma cells (2).”

In 1969 zur Hausen returned to Germany to take an appointment as an independent scientist at the University of Wurzburg. His research was now focused entirely on EBV. Specifically, he wanted to challenge the prevailing view that Burkitt’s lymphoma tumors are persistently infected with EBV (i.e., that the tumors continuously produce low levels of the virus).

I presume that zur Hausen was interested in this issue because it was reasonable to believe that EBV gene expression is necessary to maintain the neoplastic state of the Burkitt’s tumor cells. Persistent infection would be one means by which viral genes could be carried by the cells. But zur Hausen believed that EBV DNA might be maintained in Burkitt’s lymphoma cells, even if they did not produce EBV particles.

Werner Henle in Philadelphia (and also George Klein in Stockholm) sent zur Hausen a large number of Burkitt’s lymphoma cell lines and tumor biopsies to aid in his study. One of those cell lines, the Raji line of Burkitt’s lymphoma cells, did not produce EBV particles. Nevertheless, zur Hausen isolated sufficient EBV DNA from the Raji cells to prove that multiple copies of EBV DNA were maintained in them. This was the first time that tumor virus DNA was shown to be present in malignant human cells that were not producing virus. See Aside 2.

[Aside 2: In 1968 Renato Dulbecco and co-workers were the first to discover viral DNA integrated by covalent bonds into cellular DNA (J. Sambrook et al., Proc. Natl. Acad. Sci. U S A. 60:1288, 1968). They were studying cells transformed by the polyomavirus, SV40. Integration explained how SV40 genes could be stably maintained and expressed in transformed cells, in the absence of productive infection. Integration is now recognized as a key event in cell transformation by members of several virus families, including the polyomaviruses, papillomaviruses, and the oncogenic retroviruses.

The situation in the case of EBV, a herpesvirus, is different, as herpesviruses are able to enter into a latent state in host cells. In the latent state the viral genome is maintained as an episome, and only a subset of the viral genes (i.e., those concerned with latency) are expressed. The episomal viral genome is replicated by the cellular DNA replication machinery during the cell cycle S phase, and a viral gene product, EBNA-1, ensures that viral genomes are equally partitioned between the daughter cells. In 1978 George Klein and co-workers were the first to demonstrate episomal EBV DNA in a cell line derived from a Burkitt’s lymphoma biopsy (S. Koliais et al., J. Natl. Cancer. Inst. 60:991, 1978).]

In 1972, while zur Hausen’s attention was focused on EBV and Burkitt’s lymphoma, his research direction took a providential turn that would lead to his most important discoveries. It happened as follows.

Recent seroepidemiological evidence was suggesting a link between herpes simplex virus type 2 (HSV-2), a well known genital infection, and cervical cancer. Since HSV-2, like EBV, is a herpesvirus, and since zur Hausen had already demonstrated that EBV DNA is present in Burkitt’s lymphoma tumor cells, zur Hausen believed he was well positioned to search for HSV-2 DNA in cervical cancer biopsies. However, in this instance, all his repeated attempts failed.

Harald zur Hausen then came across anecdotal reports of genital warts converting to squamous cell carcinomas. Importantly, those genital warts were known to contain typical papillomavirus particles. Taking these two points into account, zur Hausen considered the possibility that papillomaviruses, rather than herpesviruses, might be the cause of cervical carcinomas. Indeed, his initial thought was that the genital wart papillomavirus might also be the etiologic agent for cervical carcinomas.

Thus, Harald zur Hausen began his foray into papillomavirus research. His first experiments found papillomavirus particles in benign plantar (cutaneous) warts. His next experiments demonstrated that there are multiple papillomavirus subtypes. [In brief, zur Hausen used in vitro-transcribed plantar papillomavirus RNA as a hybridization probe against the DNA from various plantar and genital warts. Only some of the plantar wart DNAs, and none of the genital wart DNAs, reacted with his planter wart RNA probe. Restriction endonuclease patterns of a variety of human papillomavirus isolates confirmed that the HPVs comprise a heterogeneous virus family.]

Harald zur Hausen’s next experiments sought to detect papillomavirus DNA in cervical carcinoma biopsies. However, his initial trials in this crucial undertaking were unsuccessful.  He was using DNA from HPV-6 (isolated from a genital wart) as a hybridization probe in those failed attempts. But zur Hausen and co-workers had at hand a number of additional HPV subtypes, from which they prepared other DNA probes. DNA from HPV-11 (from a laryngeal papilloma) indeed detected papillomavirus DNA in cervical carcinomas.

In 1983, two of Zur Hausen’s former students, Mathias Dürst and Michael Boshart, using HPV-11 DNA as a probe, isolated a new HPV subtype, designated HPV-16, from a cervical carcinoma biopsy. In the following year, they isolated HPV-18 from another cervical carcinoma sample. Harald zur Hausen’s group soon determined that HPV-16 is present in about 50% of cervical cancer biopsies, while HPV-18 is present in slightly more than 20%. [The famous HeLa line of cervical cancer cells contains HPV-18 DNA.]

Additional key discoveries took place during the next several years, including the finding that papillomavirus DNA is integrated into the cellular DNA of cervical carcinoma cells. This finding clarified how papillomavirus genes persist in the cancers, and also revealed that the cancers are clonal (see Aside 2, above). Moreover, while the integrated viral genomes often contain deletions, zur Hausen’s group found that two viral genes, E6 and E7, are present and transcribed in all cervical cancer cells. This finding implied that E6 and E7 play a role in initiating and maintaining the oncogenic state. [In 1990 Peter Howley and co-workers demonstrated that the interaction of the E6 gene product with the cellular tumor suppressor protein p53 results in the degradation of p53. In 1992 Ed Harlow and coworkers showed that the E7 gene product blocks the activity of the cellular tumor suppressor protein pRb. Reference 3 details the mechanisms of papillomavirus carcinogenesis.]

The above findings led to widespread acceptance that cervical carcinoma is caused by papillomaviruses. Yet acceptance was not immediate. The prevailing belief, that herpesviruses cause cervical carcinoma, was well-entrenched and slow to fade away. It was based on the observation that many women afflicted with cervical carcinoma also had a history of genital herpes. But, individuals infected with one sexually transmitted pathogen are often infected with others as well. Apropos that, genital warts were long thought to be associated with syphilis, and later with gonorrhea. In any case, in 1995 the World Health Organization officially accepted that HPV-16 and HPV-18 are oncogenic in humans.

Harald zur Hausen was awarded one half of the 2008 Nobel Prize for Medicine or Physiology for proving that cervical cancer is caused by human papillomaviruses. By the time of his award, his findings had led to key insights into the mechanism of HPV-mediated carcinogenesis and, importantly, to the development of a vaccine to prevent cervical cancer. See Aside 3.

[Aside 3: The first generation of Gardasil, made by Merck & Co., helped to prevent cervical cancer by immunizing against HPV types 16 and 18, which are responsible for an estimated 70% of cervical cancers. Moreover it also immunized against HPV types 6 and 11, which are responsible for an estimated 90% of genital warts cases. Apropos genital warts, there are 500,000 to one million new cases of genital warts (also known as condylomata acuminate) diagnosed each year in the United States alone.

The original vaccine was approved by the USFDA on June 8, 2006. An updated version of Gardasil, Gardasil 9, protects against the HPV strains covered by the first generation of the vaccine, as well as five additional HPV strains (HPV-31, HPV-33, HPV-45, HPV-52, and HPV-58), which are responsible for another 20% of cervical cancers. Gardasil 9 was approved by the USFDA in December 2014.]

Harald zur Hausen reviewed the overall contribution of viruses to human cancer in his 2008 Nobel lecture (4). Some of his key points are as follows. HPVs were discussed above with respect to cervical carcinoma. HPVs also are associated with squamous cell carcinomas of the vagina, anus, vulva, and oropharynx. What’s more, 40% of the 26,300 cases of penile cancer reported worldwide in 2002 could be attributed to HPV infection.

Epstein-Barr virus too was discussed above. This member of the herpesvirus virus family causes nasopharyngeal carcinoma, as well as Burkitt’s lymphoma. Another herpesvirus, human herpesvirus 8, causes Kaposi’s sarcoma; the most frequent cancer affecting AIDS patients. Hepatitis B virus (HBV, a hepadnavirus), as well as hepatitis C virus (HCV, a flavivirus), causes hepatocellular carcinoma. The human T-lymphotropic virus 1 (HTLV-1), a retrovirus, induces adult T-cell leukemia. And the recently discovered Merkel cell polyomavirus (MCPyV) is responsible for Merkel cell carcinoma.

Harald zur Hausen estimated that viruses directly cause about 20% of all human cancers, and a similar percentage of all deaths due to cancer! And while 20% might seem to be a remarkably high figure for the extent of viral involvement in human cancer, zur Hausen suggests that it is actually a minimal estimate. That is so because it is difficult to determine that a particular virus is actually the cause of a cancer. Consequently, it is likely that other examples of viral involvement in human cancer will be discovered.

Harald zur Hausen gave two principal reasons for why it is difficult to establish that an infectious agent is the cause of a cancer in humans. First: “… no human cancer arises as the acute consequence of infection. The latency periods between primary infection and cancer development are frequently in the range of 15 to 40 years…” Second: “Most of the infections linked to human cancers are common in human populations; they are ubiquitous… Yet only a small proportion of infected individuals develops the respective cancer type.”

Viruses also contribute to the human cancer burden in an indirect way. For instance, HIV types 1 and 2 play an indirect role in cancer via their immunosuppressive effect, which is the reason for the extraordinarily high prevalence and aggressiveness of Kaposi’s sarcoma in AIDS patients.

Bacterial infections also contribute to the cancer burden. For example, Helicobacter pylori infections may lead to stomach cancer. What’s more, the parasites Schistosoma, Opisthorchis, and Clonorchis have been linked to rectum and bladder cancers in parts of Northern Africa and Southeast Asia, where they are prevalent.

Obviously, but important enough to state anyway, knowing that a particular cancer is caused by a particular infectious agent opens the possibility of developing a rational strategy to prevent that cancer. Gardasil is an exmple. A vaccine against HBV is also available, and one against HCV is under development.

References:

1. Who discovered HIV, Posted on the blog January 23, 2014.

2. MLA style: “Harald zur Hausen – Biographical”. Nobelprize.org. Nobel Media AB 2014. Web. 27 May 2015. <http://www.nobelprize.org/nobel_prizes/medicine/laureates/2008/hausen-bio.html&gt;

3. Norkin, Leonard C. (2010) Virology: Molecular Biology and Pathogenesis. ASM Press, Washington, D.C. See Chapters 15 and 16.

4. Zur Hausen, Harold, The search for infectious causes of human cancers: where and why. Nobel Lecture, December 7, 2008.

Louis Pasteur: One Step Away from Discovering Viruses

Louis Pasteur (1822-1895) is the subject of our first posting of the New Year. Pasteur was history’s greatest microbiologist and, perhaps, its most famous medical scientist. Pasteur was also an early figure in the history of virology for his 1885 discovery of a rabies vaccine; only the second antiviral vaccine and the first attenuated one (see Aside 1). However, the main point of this tale is that Pasteur let pass an especially propitious opportunity to discover that the rabies agent is one of a previously unrecognized class of microbes; a class that is fundamentally different from the already known bacteria. Its members are submicroscopic and grow only inside of a living cell. Pasteur was just one step away from discovering viruses.

Louis Pasteur
Louis Pasteur

[Aside 1: Attenuation is the conversion of a pathogenic microbe into something that is less able to cause disease, yet is still able to induce immunity. Edward Jenner’s 1798 smallpox vaccine, the world’s first vaccine, as well as the first antiviral vaccine, was not based on the principle of attenuation. Instead, it contained live, unmodified cowpox virus. Although hardly understood in Jenner’s day, his smallpox vaccine worked because cowpox, which is not virulent in humans, is immunologically cross-reactive with smallpox. Thus, the relatively benign cowpox virus induced immunity against the related, deadly smallpox virus (1).]

The distinctive nature of viruses would first begin to be revealed in 1887 by a scientist of much less renown than Pasteur; the Russian microbiologist Dmitry Ivanovsky. The virus concept would be further advanced in 1898 by the accomplished Dutch botanist Martinus Beijerinck (2). In any case, to better appreciate how anomalous it was that Pasteur did not discover viruses, we review the greatness of his earlier achievements. After that, we consider the opportune circumstance that he let go by.

Pasteur was a chemist by background. Thus, his first major scientific discovery, at 26 years of age, was as a chemist. It was his 1847 discover of molecular asymmetry; that certain organic molecules exist in two alternative molecular structures, each of which is the mirror image of the other. Additionally, pairs of these asymmetric molecules are chemically indistinguishable from each other, and balanced mixtures of them rotate the plane of polarized light.

Pasteur’s discovery of molecular asymmetry was one of the great discoveries in chemistry. Yet his research would take on a momentous new focus when he began to investigate the chemistry of fermentations. This new course was inspired by the fact that while asymmetric molecules are not generated in the laboratory, they are found in the living world. And, since asymmetric molecules are found among fermentation products, Pasteur hypothesized that fermentation is a biological process, which he proceeded to demonstrate in 1857, basically by showing that fermentation products did not arise in nutrient broth if any microbes that might have been present were either killed by heating or removed by filtration. What’s more, he showed that specific fermentations are caused by specific microorganisms. Additionally, he discovered that fermentation is usually an anaerobic process that actually is impaired by oxygen; a phenomenon known as the “Pasteur effect.” And, he put forward the notion of aerobic versus anaerobic microbes.

Pasteur put his experience studying fermentations to practical use when he came to the rescue of the French wine industry, which was on the verge of collapse because of the wine becoming putrefied. Pasteur showed that the problem was due to bacterial contamination, and then showed that the putrefaction could be prevented by heating the wine to 50 to 60 °C for several minutes; a procedure we now refer to as pasteurization. Wines are seldom pasteurized today because it would kill the organisms responsible for the wines maturing. But, as we know, pasteurization is applied to many contemporary food products, especially milk. Pasteur also aided the beer industry by developing methods for the control of beer fermentation.

Pasteur’s study of fermentations led to an experiment of historic significance for biology in general. In the 1860s, the ancient notion that life can arise spontaneously from nonliving materials, such as mud or water, was still widely believed. The emerging awareness of microbes in the 1860s did not change this belief. Instead, it led to the idea that fermentations and putrefactions result from the spontaneous generation of microbes. In 1862, Pasteur unequivocally dispelled this belief by a simple yet elegant experiment in which he made use of a flask that had a long bending neck that prevented contaminants from reaching the body of the flask. If the broth in the flask was sterilized by boiling, and if the neck remained intact, then the broth remained sterile. But, if the neck of the flask was broken off after the boiling, then the broth became opaque from bacterial contamination.

Taken alone, Pasteur’s achievements that are enumerated above would have been sufficient to have ensured his lasting fame. Nevertheless, Pasteur’s greatest successes were yet to come. In 1867 he put forward the “germ theory of disease.” By this time, the existence of a variety of microorganisms, including bacteria, fungi, and protozoa, was already well established. Pasteur’s new proposal, that microorganisms might produce different kinds of diseases, was inspired by his earlier experimental findings that different microorganisms are associated with different kinds of fermentations, and by his 1865 finding that a microorganism was responsible for a disease in silkworms that was devastating the French silk industry.

After Pasteur proposed his germ theory of disease, Robert Koch (another giant in the history of medical microbiology) established that anthrax in cattle is caused by a specific bacterium, Bacillus anthracis. Koch had taken a sample from diseased cattle and then used his new method for isolating pure bacterial colonies on solid culture media to generate a pure culture of B. anthracis. Next, he inoculated healthy animals with a portion of the pure culture. The healthy animals then developed anthrax. Finally, he re-isolated B. anthracis from the inoculated animals. This sequence of isolation, infection, and re-isolation constitutes Koch’s famous postulates, which provide the experimental basis for establishing that a specific microorganism is responsible for a specific disease.

Even after Pasteur confirmed Koch’s anthrax findings in 1877, some members of the medical establishment still rejected the germ theory of disease, mainly because Pasteur was a chemist by background, rather than a physician. Nevertheless, Joseph Lister, an English surgeon, admired Pasteur’s work on fermentation and was impressed by Pasteur’s disproving of spontaneous generation. Based on Pasteur’s demonstration of the ubiquity of airborne microorganisms (another of his noteworthy achievements), Lister reasoned that infections of open wounds are due to microorganisms in the environment. The aseptic techniques that Lister then introduced were responsible for dramatically reducing infections during surgery.

The following is one of my favorite parts of this story. In 1879, Pasteur made his first important contribution to vaccinology, when he discovered, by accident, that he could attenuate the bacterium responsible for chicken cholera (now known to be a member of genus Pasteurella), and then use the attenuated microbe as a vaccine. It happened as follows. Pasteur instructed his assistant, Charles Chamberland, to experimentally inject chickens with the cholera bacterium so that he, Pasteur, might observe the course of the disease. Then, just before a summer holiday break, Pasteur directed Chamberland to inject the chickens with a fresh culture of the bacteria. Chamberland may have been preoccupied with thoughts of the upcoming holiday, because he forgot to inject the chickens before leaving. When he returned a month later, he carried out Pasteur’s instructions, except that he injected the chickens with the now aged bacteria. What happened next was most important. The chickens that were inoculated with the aged culture developed only a very mild form of the disease. After that, Pasteur had Chamberland inject those same chickens with freshly grown, presumably virulent bacteria. The chickens still did not develop disease.

It is not clear why Pasteur instructed Chamberland to inoculate the freshly grown culture into the chickens that earlier had received the aged culture. Perhaps it was an accident, or perhaps Pasteur saw an opportunity to carry out a possibly interesting experiment. (The chickens had survived a mild infection by the aged culture. Might they now be resistant to freshly grown virulent bacteria?) In any case, Pasteur repeated the entire sequence of inoculating the chickens with an aged culture and then challenging them with a fresh culture. The outcome was the same as before.

Pasteur correctly surmised that the aging process (actually, oxidation by exposure to air) had attenuated the bacteria. And, he learned by experimentation that the virulence of the cholera microbe could be reduced to any desired extent by controlling its exposure to air. Most importantly perhaps, he discovered that the attenuated bacteria could induce resistance to the virulent bacteria and, consequently, could be used as a vaccine. Pasteur’s chicken cholera vaccine was the first vaccine deliberately created in a laboratory. What’s more, it was the first attenuated vaccine. See Aside 2.

[Aside 2: During the years that Pasteur was carrying out his vaccine studies, nothing was known regarding the physiological basis of immunity, or the determinants of virulence, or of mutations, or the underlying mechanism of attenuation that changed a deadly microbe into a harmless one that still could induce immunity. Considering the intellectual milieu in which Pasteur carried out his investigations, it is all the more remarkable that he achieved so much. And while Pasteur’s interpretations for how his attenuated vaccines worked were far from accurate, they are still impressive for their plausibility. Initially, he thought that the attenuated organisms might simply compete with the virulent organisms for a limited availability of nutrients in the host. Later, he thought that the attenuated organisms might release a toxin that blocked growth of the virulent organisms. The notion, that the host might actually initiate its own defense, began to emerge in 1890 when Emil von Behring and Shibasaburo Kitasato discovered that a host factor neutralized the diphtheria toxin. Kitasato then put forward the theory of humoral immunity, proposing that a host serum factor could neutralize a foreign antigen. In 1891 Paul Ehrlich used the term “antibody” for the first time, in reference to those serum factors.]

This account of the cholera vaccine brings to mind Pasteur’s famous remark, “Chance only favors the prepared mind.” Yet in the context of our larger story, it is an ironic statement, considering that Pasteur later missed an auspicious opportunity to discover viruses. But, before getting to that, we briefly note Pasteur’s work on his anthrax vaccine.

In 1879 Pasteur began to develop an anthrax vaccine, which, like the cholera vaccine, would be based on his principle of attenuation. And, as in the case of the cholera vaccine, Pasteur attenuated the anthrax bacillus by exposing it to oxygen. [History records that Pasteur and his assistants developed a second approach to attenuate the anthrax bacillus, based on their discovery that when the bacilli are cultivated at 42 or 43 degrees centigrade, they do not develop the endospores that are necessary to cause a virulent infection.] In 1881 Pasteur carried out a dramatic public demonstration of the effectiveness of his air-oxidized anthrax vaccine in livestock, causing many doubters to accept the validity of his work. See Aside 3 and the end note.

[Aside 3: Currently, the only FDA-licensed anthrax vaccine for use in humans is BioThrax, produced by Emergent BioDefense Operations Lansing Inc. BioThrax is generated from an avirulent, nonencapsulated mutant of B. anthracis. It does not contain any living organisms. As suggested by the name of the manufacturer, BioThrax was produced mainly for the U.S. Department of Defense, for use in case B. Anthracis might be used as a biological weapon. Thus, BioThrax is not available to the public. People who are exposed to B. anthracis are now treated with antibiotics (e.g., ciprofloxacin and doxycycline).]

Pasteur turned his attention to rabies in1880, when the problem of rabid dogs in Paris was getting out of hand. Once again Pasteur sought to develop a vaccine, and once again he wanted to apply the principle of attenuation. But, early on, he found that he could not grow the rabies agent in pure culture. Thus, he was not able to isolate the rabies agent. Moreover, he would need to devise new procedures if he was to grow and attenuate it. His solution was to develop methods for cultivating the rabies agent in the spinal cords of live rabbits. His method for attenuation was then suggested by his assistant, Emile Roux, who had been studying survival of the rabies agent in pieces of rabbit spinal cord that he suspended inside a flask. Following Roux’s example, Pasteur attenuated the rabies agent by air-desiccating spinal cords taken from experimentally infected rabbits that earlier had died of rabies. Each successive day of desiccation resulted in greater attenuation of virulence, such that an extract from a spinal cord aged for 14 days could no longer transmit the disease. What’s more, those extracts could be used as inoculums that prevented rabies in dogs that later were challenged with the virulent microbe.

Pasteur, himself, took saliva samples from rabid dogs. In one such incident, he used a glass tube to suck up a few drops of deadly saliva from the mouth of a mad, squirming bulldog that was held down on a table by two assistants. The assistants wore heavy leather gloves.

Here is another of my favorite parts of this story. In 1885, nine-year-old Joseph Meister was bitten repeatedly by a rabid dog. Young Joseph’s desperate mother then brought her son to Pasteur, hoping that he might help Joseph. But, any attempt by Pasteur to treat the boy was sure to provoke controversy. Pasteur was not a medical doctor. Moreover, his rabies vaccine had never been successfully used in humans. Furthermore, attenuation and vaccination were still new and contentious concepts. For these reasons, Pasteur rejected many earlier requests for help from people in France, and from abroad as well. But, in Joseph’s case, Pasteur relented, convinced that the boy would die if he did not intercede.

Pasteur gave young Joseph a series of 13 injections, one each day, in which each successive injection contained a less-attenuated (stronger) virus. Pasteur dreaded inoculating Joseph with the last shot in the series; a one-day-old vaccine that was strong enough to kill a rabbit. Emile Roux wanted no part in this episode and, in fact, withdrew from the rabies study because of it. But, Joseph never developed rabies, and millions of people subsequently received Pasteur’s anti-rabies treatments. [Pasteur’s attenuated rabies vaccine may not have been entirely safe for humans. Modern rabies vaccines are generally killed virus vaccines, prepared by chemically inactivating tissue culture lysates.] See Asides 4 and 5.

[Aside 4: Post-infection rabies vaccination works and, indeed, is necessary because (for reasons that are still not entirely clear) the human immune response against a natural rabies infection is not able to prevent the virus from reaching the central nervous system, at which point the infection is invariably fatal. Importantly, the incubation period between the time of the bite and the appearance of disease can be more than several months, and is never less than two weeks. Consequently, there is a substantial window of opportunity for the vaccine to cause the virus to be inactivated at the site of the bite.]

[Aside 5: In 1888, Emile Roux, working at the Pasteur Institute (see below), would confirm the existence of the diphtheria toxin by showing that injecting animals with sterile filtrates of liquid cultures of Corynebacterium diphtheriae caused death with a pathology characteristic of actual diphtheria.]

Pasteur worked hard to isolate the rabies agent, but he wrongly presumed that he should be able to grow it in pure culture. Finally, in 1884, he conceded that he had not been able to isolate and cultivate the rabies agent in a laboratory media. So, might that failure alone have been sufficient to cause Pasteur to think of the rabies agent in new terms? Perhaps not, since, at the time, the inability to cultivate a microbial pathogen was assumed to be a laboratory failure, rather than a reason to hypothesize that that the agent was something other than a bacterium. [Even with the eventual awareness of the uniqueness of viruses, the inability of virologists to cultivate viruses outside of an animal would remain a mystery, as well as an obstacle, well into the early 1930s (3).]

Pasteur also got sidetracked while trying to isolate the rabies agent. In 1880 he injected a rabbit with the saliva of a child who died of rabies. He then examined the blood of the rabbit after it too succumbed to rabies. Using his microscope, Pasteur in fact saw a microbe in the rabbit’s blood, which he thought might be the rabies agent. However, he later found the same microbe in the saliva of normal children. Ironically, this microbe, which Pasteur at first thought might be the rabies agent, was actually Pneumococcus pneumoniae, a major bacterial pathogen that was correctly identified several years later by Albert Frankel. Thus, Pasteur missed the opportunity to identify a bacterial pathogen that is much more important in humans than rabies virus. Moreover, and importantly, Pasteur never did see the actual rabies agent under his microscope. Thus, he was aware that the rabies agent might be unusually small in comparison to the usual bacteria.

Here is another bit of irony. The item (apparatus?) that initially played the key role in distinguishing viruses from bacteria was invented in Pasteur’s laboratory. It was the unglazed terra cotta filter, conceived by Charles Chamberland, which he used to provide a good supply of sterile water for Pasteur’s lab. Chamberland allegedly developed these bacterium-proof filters while experimenting with a broken clay pipe that he bought from his tobacconist.

Bearing in mind that Pasteur was never able to grow the rabies agent in pure culture, and that he never saw the rabies agent under his microscope, might he have thought that it might be a submicroscopic infectious agent that is different from bacteria in some fundamental way? I have not come across any definitive answer to that question. But, I feel safe to say that it is unlikely that anyone other than Pasteur might have seriously considered that possibility. Regardless, Pasteur did not take the next logical step, which would have been to see if the rabies agent might pass through Chamberland’s filters. Had he done so, he could have isolated the rabies agent from the rabbit spinal cords, and he would have discovered “filterable viruses” (see below).

That crucial step was taken for the first time in 1887 by the Russian bacteriologist, Dmitry Ivanovsky, who used Chamberland filters in his investigations into the cause of tobacco mosaic disease. Ivanovsky could not propagate the tobacco mosaic agent (later known as the tobacco mosaic virus) in pure culture. However, because of his finding that the agent could actually pass through Chamberland’s filters, Ivanovsky is sometimes credited for discovering viruses. Yet Ivanovsky did not accept his own results. He still presumed that the disease was caused a bacterium, and he thought that the filters were defective or, instead, that the disease was due to a toxin produced by the bacterium.

In 1898, Martinus Beijerinck, unaware of Ivanovsky’s earlier work, also could not see or cultivate the tobacco mosaic agent. In addition, he too found that the agent passed through Chamberland filters. Beijerink expected, and perhaps even hoped that the filters would remove the agent from diseased plant extracts, so that he might prove it to be a bacterium. But despite his possible disappointment, Beijerinck went one major step further. He demonstrated that the filtered sap from a diseased plant did not lose its ability to cause disease after being diluted by repeated passage through new healthy plants. Consequently, the filterable agent was replicating in the plant tissue and, thus, could not be a toxin.

Little is recorded about Ivanovsky, aside from his four-page report on the tobacco mosaic disease (see Aside 6). In contrast, Beijerinck was a major scientist, who made numerous important contributions, including the discovery of nitrogen-fixing bacteria and bacterial sulfate reduction (4). Yet even Beijerinck found it difficult to conceive that the filterable, incredibly small, submicroscopic tobacco mosaic agent might be particulate in nature. Instead, he famously described it as a “contagious living fluid.” Nonetheless, Beijerinck, a botanist by background, is often considered to be the first virologist.

[Aside 6: Ivanovsky’s four-page paper would be unremarkable if it were not for the single sentence, “Yet I have found that the sap of leaves attacked by the mosaic disease retains its infectious qualities even after filtration through Chamberland filters.”]

Pasteur was probably unaware of Ivanovsky’s findings, and he did not live long enough to know of Beijerinck’s. So, we do not know what he might have made of their results. Regardless, Pasteur remained one step away from making these discoveries himself.

In 1898, after the announcement of Beijerinck’s findings, Friederich Loeffler and Paul Frosch isolated the foot and mouth disease virus; the first virus isolated from animals. Next, in 1901, in Cuba, U.S. Army doctor Walter Reed isolated yellow fever virus (5); the first pathogenic virus of humans to be isolated. In 1903, Paul Remlinger, working at the Constantinople Imperial Bacteriology Institute, filtered and then isolated rabies virus. Despite these early achievements, it was not until 1938 that the development of the electron microscope made it possible to resolve that viruses are indeed particulate, rather than liquid in nature. See Aside 7.

[Aside 7: The term “virus” indeed appears in the scientific literature of Pasteur’s day. However, at that time “virus” referred to any microbe that might cause disease when inoculated into a susceptible human or animal. By the 1890s, the term “filterable virus” came into use, meaning an infectious agent which, like the tobacco mosaic virus, passed through filters that retained bacteria. But, bearing in mind that there was not even a consensus regarding the identity of the genetic material until the early 1950s, there would be no clear understanding of viruses until then. In fact, the classic, early 1950s blender experiment of Alfred Hershey and Martha Chase, which featured bacteriophage T4, played a key role in establishing DNA as the genetic material, while also elucidating the essentials of virus replication (2).]

In 1887 Louis Pasteur founded the Institute in Paris that bears his name. A minor irony is that the Pasteur Institute was founded as a rabies vaccine center. The Institute has since been the site of numerous major discoveries in infectious diseases. But we underscore here that it was the site where, in 1910, Constantin Levaditi and Karl Landsteiner demonstrated that poliomyelitis is caused by a filterable virus, and where Félix d’Herelle in 1917 discovered bacteriophages. And it was also the site where, in 1983, Luc Montagnier and Françoise Barré-Sinoussi were the first to isolate HIV (6).

In a fitting end to our story, when Joseph Meister grew up, he became the gatekeeper of the Pasteur Institute. Meister was still minding the gate at age sixty four when, in 1940, the Nazis invaded Paris. Legend has it that when Nazi soldiers arrived at the Institute and ordered Meister to open Pasteur’s crypt, rather than surrendering Pasteur’s resting place to the Nazis, Meister shot himself (7).

Pasteur Institute: Museum and Crypt
Pasteur Institute: Museum and Crypt

End note:

Science historian, Gerald L. Geison, wrote a controversial revisionist account of Pasteur’s achievements, that was based on Geison’s reading of Pasteur’s laboratory notes (8). Geison undermines Pasteur’s integrity and discredits some of his major accomplishments. For example, Geison asserts that Pasteur surreptitiously used the oxidation procedure of French veterinary surgeon, Henry Toussaint, when preparing his own widely acclaimed anthrax vaccine for its public demonstration.

Max Perutz, who shared the 1962 Nobel Prize for Chemistry with John Kendrew for their studies of the structures of hemoglobin and myoglobin, reviewed Geison’s book for The New York Review of Books (December 21, 1995). Perutz’s review, entitled The Pioneer Defended, contains a vigorous rebuttal of Geison’s claims. Geison responded to Perutz’s review in the April 4, 1996 issue of The New York Review of Books. Perutz’s counter-response immediately follows.

I make note of all this because Geison’s uncertain assertions are reported as unqualified fact in some accounts of Pasteur’s work. And, while Perutz’s representations are not entirely accurate, the review, the response, and the counter-response make a very interesting read.

References:

(1) Edward Jenner and the Smallpox Vaccine, Posted on the blog September 16, 2014.

(2) Norkin, L. C. Virology: Molecular Biology and Pathognesis, ASM Press, 2010. Chapters 1 and 2 review key developments towards the understanding of viruses.

(3) Ernest Goodpasture and the Egg in the Flu Vaccine, Posted on the blog November 26, 2014.

(4) Chun, K.-T., and D. H. Ferris,  Martinus Willem Beijerinck (1851-1931) Pioneer of general microbiology, ASM News 62, 539-543, 1996.

(5) The Struggle against Yellow Fever: Featuring Walter Reed and Max Theiler, Posted on the blog May 13, 2014.

(6) Who Discovered HIV?, Posted on the blog January 23, 2014.

(7) Dufour, H. D., and S. B. Carroll, (2013), History: Great myths die hard, Nature 502, 32–33. This note contains an update on the myth.

(8) Geisen, G. L., The Private Science of Louis Pasteur, Princeton University Press, 1996.

Thabo Mbeki and the South African AIDS Epidemic

I recently watched the fact-based 2009 movie Endgame, which depicts the final days of apartheid in South Africa. The movie focused on how a young Thabo Mbeki facilitated the late 1980s secret talks between Afrikaner leaders and the about-to-be freed Nelson Mandela’s African National Congress. The purpose of these talks was to arrange negotiations between de Klerk’s apartheid government and the ANC, in order to facilitate the stable transition of South Africa from apartheid.

imbeki, mandelaThabo Mbeki with Nelson Mandela

Mbeki is portrayed in the film as a wise, compassionate, and respected political activist. Moreover, all commentary on the movie that I’ve read is consistent with it having accurately depicted people and events. Here then is Mbeki in a different guise, when dealing with the South African AIDS epidemic, after succeeding Mandela as the nation’s president. As usual, we begin with some background.

The South African AIDS outbreak may well have been the most devastating of all the world’s AIDS epidemics. Consider the following statistics. By 2007, more than one in five South African adults (approximately 5 million people) were living with HIV, and the disease claimed nearly 1,000 lives daily. Shockingly, 71% of all deaths among individuals between 15 and 49 years of age were due to AIDS. Moreover, a 15-year old South African had a 50:50 chance of dying of AIDS before his or her 30th birthday.

Yet despite these statistics, South Africa escaped the initial HIV/AIDS epidemic of the 1980’s. Why then did HIV/AIDS get so out of hand in South Africa in the early 21st century? One reason is that it was a time when the government was preoccupied with the nation’s poverty and its transition from apartheid. Moreover, and importantly, any rational efforts to deal with the South African HIV/AIDS epidemic were severely compromised when, in 2001, President Mbeki embraced the “denialist” view that AIDS is not caused by HIV. [See the addendum on the AIDS denialists at the end of this posting.]

Mbeki’s “denialist” position on AIDS dates back to 2001, when his minister of health convened a 36-member international panel of supposed AIDS experts to advise Mbeki on how best to confront the nation’s AIDS crisis. Regrettably, the14 known HIV/AIDS denialists, who were invited by the health minister to join Mbeki’s AIDS Advisory Panel, convinced Mbeki to adopt their denialist point of view.

Afterwards, Mbeki and his administration were repeatedly accused of improperly dealing with their nation’s AIDS epidemic. Noting that AIDS patients succumb to the opportunistic infections that appear following the HIV-caused breakdown of their immune systems, South African AIDS patients, who used the public health system, were able to get treatment for their opportunistic infections. However, the Mbeki government prevented those patients from receiving antiretroviral therapy, which might have prevented the breakdown of their immune systems in the first place.

Governmental restrictions against the use of antiretroviral drugs remained in place until August 2003, when the South African Cabinet overruled Mbeki, and declared as Cabinet policy that HIV is the cause of AIDS. Moreover, the cabinet promised to formulate a national treatment plan that would include antiretroviral therapy. Yet despite the efforts of the cabinet to make antiretroviral drugs the mainstay of the country’s treatment plan, the health minister continued to promote nutritional approaches to treating AIDS, while also proclaiming the toxicity of the antiretroviral drugs.

The continued unwillingness of the Mbeki administration, to acknowledge that HIV is the cause of AIDS, prevented the cabinet’s more enlightened AIDS policy from being fully realized. By the end of 2007, only about 28% of South Africa’s AIDS patients were able to obtain antiretroviral therapy. And, since many HIV-infected pregnant women were not getting the antiretroviral drugs that might have prevented transmission of the virus to their babies, HIV remained tragically common among South African children. In this regard, in 2007, UNAIDS reported that there were about 280,000 children under 15 years of age who were living with HIV in South Africa. Moreover, AIDS-related adult deaths resulted in 1.4 million South African children becoming orphans in 2007; a rise from the 780,000 new orphans reported in 2003.

A 2008 report from the Harvard School of Public Health estimated that Mbeki’s denialist stance caused an estimated 330,000 or more preventable AIDS-related deaths in South Africa. That estimate was based on comparing the effect of antiretroviral therapy in neighboring Botswana and Namibia, to the state of affairs in South Africa, during the period from 2000 to 2005.

Mbeki resigned the presidency in September 2008, after losing the support of his party. His resignation was not related to his AIDS denialism. Rather, it involved his alleged interfering with the country’s National Prosecuting Authority in a political matter.

Following Mbeki’s resignation, his successor, Kgalema Motlanthe, fired health minister Manto Tshabalala-Msimang, who, like Mbeki, was an AIDS denialist. South Africa then initiated the largest antiretroviral therapy roll-out program in the world, resulting in a 5-year jump in life expectancy to a current 36 years-of age. Moreover, in 2011, there were 100,000 fewer AIDS-related deaths in South Africa than occurred in 2001, and over 300,000 fewer than occurred in 2006. Still, about 5.6 million South Africans were estimated to be infected with HIV in 2011; the highest number of infected people in any country! Almost one-in-three women aged 25-29, and over a quarter of men aged 30-34, were HIV-positive

Considering the prevalence of HIV in South Africa, it may surprise some readers that there is widespread prejudice in the country against those living with AIDS. With that in mind, we note Nelson Mandela’s action when his son died of AIDS in 2005. Mandela deliberately revealed the cause of his son’s death to the public. His purpose was to countermand the stigma associated with being infected with HIV and, also, as a “political statement designed to… force the President [Mbeki] out of his denial.” Earlier, in 2002, South African politicians, who were loyal to Mbeki, attacked Mandela for questioning the government’s AIDS policy.

Bearing in mind that Mbeki was a respected and intelligent leader, how might we explain his rejection of the orthodox view, based on indisputable evidence, that HIV is the cause of AIDS? Could he simply have been taken in by the AIDS denialists? Or, might there be more? A likely possibility is as follows. Mbeki was known to harbor strong anti-colonialist and anti-West sentiments, born of having come of age during South Africa’s apartheid era. Perhaps his earlier experiences caused him to see his country’s AIDS crisis as a means by which the West sought to exploit his nation’s people.

In the same vein, Mbeki was acutely conscious of South Africa’s poverty and, consequently, was likely affected by the huge expense of antiretroviral drugs. In that regard, his health minister advised him that the cost of treating the 5 million or so HIV-infected South Africans with AZT would exceed the annual health department budget by a factor of ten. And, given Mbeki’s anti-West sentiments, he was particularly sensitive to the fact that antiretroviral drugs were made and sold by powerful Western pharmaceutical companies. Additionally, Mbeki may have been skeptical of the efficacy of those drugs; an attitude reflecting the fact that the health policies of the colonial and apartheid governments in South Africa indeed were often self-serving and manipulative.

Mbeki indeed accepted that AIDS results from the collapse of the immune system. Nevertheless, he believed (or at least claimed) that the cause of that collapse was poor nutrition and poor health resulting from poverty, rather than from a virus. Thus, he argued that he needed to attend to poverty in general, rather than to AIDS in particular.

Treating HIV/AIDS has indeed been expensive for South Africa. Additionally, the more than $1 billion that the nation now spends annually on its HIV/AIDS program has been largely financed from its own domestic resources. Yet, South Africa is still desperately trying to emerge from the poverty of its former oppression. So, if you were in Mbeki’s position as the President in 2001, would you have chosen to support AIDS therapy, which might have exceeded the limits of your financial resources for the foreseeable future or, instead, would you invest in water systems, housing, schools, and hospitals? And what of other crucial social and medical problems that still abound in South Africa, such as malaria, tuberculosis, and violence against women? You cannot have it all. And even if Mbeki and his government had believed in and fully supported antiretroviral therapy, impediments to its effectiveness would still have remained. These comment are not meant to defend Mbeki, but, instead, to point up the complexity of the AIDS problem in a poverty-stricken nation emerging from apartheid.

Addendum: The AIDS denialsts: While some AIDS denialists had legitimate scientific credentials, none, except Peter Duesberg, was an expert retrovirologist. Indeed, before Duesberg emerged as an AIDS denialist, he made important contributions to the retrovirus field, including his finding that Rous sarcoma virus contains an oncogene (1). For this contribution, and others, Duesberg was elected to the prestigious U.S. National Academy of Sciences. However, once Duesberg broke ranks with other scientists over AIDS, he became a scientific outcast, no longer receiving invitations to scientific conferences and no longer able to obtain research grants.

Duesberg advocated the belief that AIDS results from drug abuse, parasitic infections, and malnutrition, rather than from a retrovirus, and that HIV itself is just another opportunistic infection. He stated that if he discovered that he was HIV antibody-positive, he would take that as an encouraging sign that his immune system was working.

Yet Duesberg was never able to offer any plausible evidence that might substantiate his dissident views. Moreover, he repeatedly ignored the numerous rebuttals of his claims that appeared in the scientific literature. [Kary Mullis, who was awarded the Nobel Prize for inventing the polymerase chain reaction (PCR), is another AIDS denialist with legitimate, indeed prestigious scientific credentials. But, unlike Duesberg, Mullis has no expertise that might be relevant to HIV/AIDS.]

In an interesting aside, in 2009, Duesberg published a paper in Medical Hypotheses, which defended Mbeki and disputed the 2008 study which reported that hundreds of thousands of lives were lost in South Africa because antiretroviral drugs were not available to AIDS patients there. Prominent AIDS researchers, including Nobel laureate Francoise Barre-Sinoussi (2) then complained to the journal’s publisher, Elsevier, asking that the paper (which had not yet been printed) be withdrawn.

Since Medical Hypotheses was the only Elsevier journal to have a policy against peer review, Elsevier then asked the editors of another of its journals, The Lancet, to oversee review of Duesberg’s paper. After The Lancet reviewers found that the paper contained numerous errors and misinterpretations, Elsevier permanently withdrew it. What’s more, Elsevier then forced Medical Hypotheses to introduce peer review. [On the one hand, the editorial policy of Medical Hypotheses, to “consider radical, speculative and non-mainstream scientific ideas,” may well have provided a means for airing new, potentially important premises. On the other hand, publication of Duesberg’s denialist notions, in what is nominally a scientific journal, would have given those notions an air of credibility, potentially impairing worldwide efforts against AIDS.]

Far from conceding his position, Duesberg claimed that Elsevier’s measures are the latest example of “censorship” imposed by the “AIDS establishment.” He then published a revised version of the paper in the Italian Journal of Anatomy and Embryology, causing further controversy.

So, taking into account Duesberg’s very real expertise as a retrovirologist, can he possibly have been right about HIV and AIDS, and is his alternative view helpful in the fight against the disease? It would make for a fascinating story if the answers were yes, or just even maybe. But, the evidence that HIV causes AIDS is, without exaggeration, overwhelming. Consider just the following. Data from matched groups of homosexual men and hemophiliacs show that only those who are infected with HIV ever develop AIDS. Moreover, in every known instance where an AIDS patient was examined for HIV infection, there was evidence for the presence of the virus. These data have been available for years, and Duesberg should have been well aware of them. What is more, there has been the enormous success of antiretroviral therapy in changing AIDS from a nearly invariably fatal disease, into a manageable one, for many HIV-infected individuals.

It is not known how many people might have been infected with HIV, or might not have benefited from effective antiretroviral therapy, because they heeded the arguments of the AIDS denialists. These individuals continue to tout their notions, to the detriment of the millions of HIV-infected individuals who listen to them.

1. The relevance of retroviral oncogenes is discussed in: The Politics of Science: Vignettes Featuring Nobel Laureate Harold Varmus during his Tenure as Director of the NIH, posted on the blog, June 2, 2014.

2. see: Who Discovered HIV, posted on the blog, January 24, 2014

Who Discovered HIV?

The Nobel Committee rewarded Luc Montagnier and Françoise Barré-Sinoussi for the discovery, but passed over Robert Gallo, who did much of the basic research that made the discovery possible.

Our story mainly involves two research groups; Robert Gallo and his colleagues at the U.S. National Institutes of Health, and Luc Montagnier and his colleagues at the Pasteur Institute in Paris. But first, we begin with a few tangential personal recollections, followed by relevant background to provide the setting for our tale.

The herpes simplex viruses and Epstein-Barr virus were the viruses that most excited the interest of my students in the 1970s, almost certainly because of their association with genital infections and infectious mononucleosis (the “kissing disease”), respectively. But, the major interest that these herpesviruses held for my students changed suddenly and dramatically in 1983 with the discovery of HIV as the cause of AIDS. In the more straitlaced early 1980s, excitement over HIV/AIDS was at least in part due to its association with human sexuality in all its forms.

HIV remains a hot topic. Nevertheless, the attention that HIV initially garnered has to some extent diminished as new emerging viruses, such as West Nile Virus, the SARS coronavirus, and the avian and swine influenza viruses arrived to replace the already familiar HIV as the most interesting of viruses. Another development which somewhat diminished concern over HIV is that while a positive HIV diagnosis in the 1980’s was essentially a “death sentence,” the development of new antiretroviral drugs has since turned AIDS into a manageable chronic infection for many HIV-infected individuals. And, in our less prudish times, the association of AIDS with sex may now rouse less interest than it did in earlier times.

The somewhat ephemeral nature of what is trendy in science, as illustrated here by the declining interest in HIV/AIDS, perhaps reflects the short-lived nature of what is hot in contemporary culture in general. This may help to explain my experience of only a few years ago when I began to excitedly recount for my virology class, comprised mostly of microbiology majors, how Robert Gallo and Luke Montagnier vied to be recognized for the discovery of HIV. I was most surprised to realize that not any of my students had ever heard of Gallo and Montagnier. This disquieting experience compels me to tell this story here. It is rich in human, scientific, political and, perhaps now, historic interest, and needs to be told.

Before beginning the story of the discovery per se, it would be good to recount again the extent of human suffering wrought by the AIDS epidemic. It was indeed enormous, especially in its early years when there were no therapies to treat what was then an almost invariably fatal infection. Indeed, the emergence of HIV/AIDS was by some criteria the worst outbreak of an infectious disease in history. Approximately 65 million people in the world were infected by the fall of 2007, and the rate of new infections remains at several million per year. Of these, 50,000 HIV infections still occur annually in the United States, and these disproportionally involve African-Americans and other minorities.

In view of the above, the unearthing of HIV as the cause of AIDS can well be regarded as one of the great discoveries of medical science. First, it led to the development of sensitive tests for HIV infection, which made it possible to asses the effectiveness of world-wide prevention efforts. And, by identifying those who might be infected, the tests significantly slowed the spread of the infection. What’s more, the tests also made the world’s blood supply safe from the virus. Second, and critically, the identification of a retrovirus as the cause of AIDS opened up the use of antiretroviral therapy to treat AIDS patients, thereby dramatically reducing morbidity and death. In fact, current antiretroviral regimens can lower viral levels in some HIV-positive patients to the point where even the risk of transmission is negligible.  And, pre-exposure prophylaxis (PrEP) may soon be available, in the form of a single pill (e.g., Truvada, a combination of the antiretrovirals, tenofovir disoproxil fumarate and emtricitabine). [Nevertheless, bear in mind that even now, with the availability of effective antiretroviral drugs, the virus is still present and ready to multiply if treatment is interrupted. Furthermore, many patients, particularly in the developing world, do not have access to these therapies. And, an HIV vaccine remains problematical. The reasons for the latter are discussed in Virology: Molecular Biology and Pathogenesis.]

Precious little was known about the underlying basis of AIDS before HIV was isolated and confirmed as its cause. Consequently, many wrongheaded hypotheses were put forward to explain the origin of the disease. For example, since AIDS first appeared among cohorts of gay men, some researchers proposed that the disease might be caused by sperm in the male bowel. And, since AIDS is characterized by a severe immunodeficiency, others suggested that it might be caused by excess stress that some individuals placed on their immune systems. Yet some investigators did suggest that AIDS might be caused by a virus. Thus, cytomegalovirus, Epstein-Barr virus, hepatitis B virus, and the herpes simplex viruses were all investigated as the possible cause of AIDS.

Interestingly, very few biomedical scientists thought that AIDS might be caused by an as yet unknown infectious agent (based on the conceit that all infectious agents had already been identified), much less a retrovirus. Indeed, prior to the discovery of HIV, it was generally thought that there are no human retroviruses; a view based on previous failed attempts to find retroviruses in human cancers. In this regard, it had been the hope of many an ambitious retrovirologist to find an oncogenic human retrovirus.

As it was, the two research groups featured here were among the very few that persisted in the search for retroviruses in human cancers. And, it was fortunate that their searches focused on leukemias in one laboratory, and T lymphocyte cultures from breast cancer patients in the other. As a consequence of their ongoing efforts, when the first patients with AIDS were identified in 1981, one of these groups was able to provide the conceptual and technical tools to isolate the AIDS virus, which the other group used to actually isolate the virus.

The stage is now set for Robert Gallo to play a key part in our story. In 1980, just before the first patients with AIDS were recognized, Gallo and his associates discovered the first known human retroviruses. These were two closely related viruses, isolated from patients with an unusual adult T-cell leukemia. Accordingly, Gallo originally named them “human T-cell leukemia virus I and –II” (HTLV-I and –II). These viruses are also known as the human T-lymphotropic virus I and –II. [See below regarding the origin of the second meaning of “L” in the acronym.]

Bearing in mind that all earlier efforts to isolate a human retrovirus were unsuccessful, why was Gallo able to isolate HTLV-I and -II in 1980? Part of the answer is as follows. During the previous 15 years, Gallo had been studying other mammalian leukemogenic retroviruses. To facilitate those studies, Gallo’s group developed methods for growing T lymphocytes in culture for extended periods. This advancement depended on the earlier discovery by Doris Morgan in Gallo’s laboratory of the T-cell growth factor, now known as IL-2.1 Importantly, the ability to grow T lymphocytes in cell culture, which enabled Gallo to grow the HTLVs, would be a critical breakthrough with regard to isolating HIV, since HIV specifically targets CD4 helper T lymphocytes in vivo. Moreover, it would be a key to developing the blood tests that detected the virus.

Other investigators also made significant advances. One of these was the discovery only ten years earlier of reverse transcriptase by Howard Temin and David Baltimore. 2 The availability of reverse transcriptase made it possible to develop highly sensitive PCR-based assays for detecting a retrovirus. These developments, taken together, enabled Gallo’s group to isolate HTLV-I and –II in 1980. And, consequently, when AIDS emerged, tools were already in place to search for a retrovirus as its causative agent.

When AIDS then suddenly appeared on the scene, Gallo saw several clues which hinted to him that its etiologic agent might be a retrovirus similar to the HTLVs.3 First, AIDS is characterized by a severe loss of CD4 CD4 helper T lymphocytes, and HTLV was already known to target T cells. Second, HTLV was known to be transmitted via blood and sexual activity, and from mother to infant; the very modes by which AIDS was proving to be transmitted. Third, a high incidence of AIDS was being reported in Haiti, a region in which HTLV is endemic. Thus, Gallo’s premise was that AIDS is caused by a variant of HTLV. That premise would prove to be incorrect, but Gallo was indeed correct in hypothesizing that it is caused by a retrovirus.

Now we turn to Luc Montagnier, a retrovirus researcher at the Pasteur Institute, who was at the time of the AIDS outbreak investigating the possible involvement of retroviruses in human breast cancers. Toward that end, Montagnier was cultivating T cells from breast cancer patients, and assaying the culture medium for reverse transcriptase activity.

In 1982, influenced by Gallo’s arguments, Montagnier set out to isolate a retrovirus as the possible etiologic agent of AIDS. As Montagnier noted, “At that time there were only a few cases in France, but they attracted the interest of a group of young clinicians and immunologists. They were looking for virologists, especially retro-virologists, as a likely hypothesis was that HTLV – the only human retrovirus known so far, recently described by R. C. Gallo – could be involved.” 4

Before Montagnier began his search for the AIDS agent, a group of French physicians and scientists suggested to him that the best chance to find and isolate it might be at the start of the disease, before the patient’s T cells had severely declined. Their reasoning was that if a virus were found at this early stage of the disease, then it would more likely be its cause, rather than merely a consequence of the immune depression. So, Montagnier and co-workers looked for a retrovirus in a lymph-node biopsy from a patient with persistent lymphadenopathy (swollen lymph glands), an early sign in patients progressing towards AIDS, but with little sign yet of the impending severe immunodeficiency. [In their later joint report, Gallo and Montagnier noted: “The idea that the causative agent of AIDS should be sought in swollen lymph nodes was partly right, since we now know that lymph nodes are the main site where the virus hides during the presymptomatic phase.” 3] Cells from this patient were cultivated in the presence of IL-2, as per Gallo’s earlier finding, as well as anti-interferon antiserum. The latter was an innovation of Montagnier, based on the earlier finding in Paris that interferon repressed the replication of retroviruses in cell culture. Two weeks later, in early January 1983, Montagnier’s research group detected the first evidence of reverse transcriptase activity in the cell culture medium.

Contrary to expectations, the new retrovirus detected in Montagnier’s laboratory was not an HTLV. This was initially shown by the fact that it did not react with anti-HTLV antibodies that were provided by Gallo. Moreover, when Montagnier’s isolate was viewed by electron microscopy, its morphology was clearly different from that of an HTLV. The difference between these viruses was further confirmed by sequence analysis. However, and crucially important, antibodies against Montagnier’s new virus were later found to be present in serum from most AIDS patients, and the virus was shown to have a tropism for CD4 T cells.

Since Montagnier’s new virus came from an AIDS patient with lymphadenopathy, he dubbed it “lymphadenopathy-associated virus” or LAV. This particular isolate of LAV was named “Bru.” Interestingly, Montagnier later obtained a biopsy from another AIDS patient who was infected with HTLV, as well as with the virus that he called LAV. If this had been the first patient sample, results might have been very confusing indeed.

Returning now to Gallo, concurrently and independently of Montagnier, he too was attempting to isolate a retrovirus from biopsies of AIDS patients. The sequence of events which then transpired was truly bizarre, beginning with the fact that while Gallo was seeking to isolate an AIDS retrovirus, he received a sample of Bru from Montagnier. Shortly afterwards, Gallo announced that he had isolated a retrovirus from an AIDS patient pool in his laboratory. 5 Moreover, Gallo’s isolate had somewhat different properties from those earlier ascribed to Bru. For example, unlike Bru, which grew only in fresh T cell cultures, Gallo’s isolate also grew in permanent T-cell lines. Bearing in mind Gallo’s premise that AIDS is caused by an HTLV variant, he reported that he had isolated a second type of AIDS retrovirus, which he named HTLV-III.

Here now is the crucial part of our story. When the nucleotide sequence of HTLV-III was determined afterwards, it turned out to be essentially identical to that of another LAV sample that had been isolated earlier in Montagnier’s laboratory. This finding was remarkable since HIV has an extraordinarily high mutation rate. And, since an untreated HIV-infected individual can produce between 108 and 1010 new virus particles each day, it would be extremely improbable to obtain virtually identical isolates from two different patient samples.

Considering the enormous importance of the discovery of the virus responsible for AIDS, and the resultant accolades that would surely go to its discoverer, the fact that Gallo’s HTLV-III was identical to a LAV isolate from Montagnier’s laboratory resulted in accusations flying back and forth between the two men. And, in part because of the sensational nature of AIDS itself, the competing claims of Gallo and Montagnier led to likewise sensational accounts of their controversy in the media of the day.

Now might be a good time to comment on the fact that Montagnier and Gallo gave different names to their AIDS isolates; LAV and HTLV-III, respectively. Importantly, the discoverer of a new virus is generally accorded the privilege of naming the virus. So bearing in mind the competing claims of Gallo and Montagnier, if the scientific community were to designate the AIDS virus as either LAV or HTLV-III, it would have been tantamount to recognizing Montagnier or Gallo, respectively, as its discoverer.

Harold Varmus, as chairman of the Retrovirus Study Group of the International Committee on Taxonomy of Viruses (ICTV), was mainly responsible for arriving at an outcome to the naming dispute that was acceptable to both protagonists, settling on “human immunodeficiency virus,” or HIV, as the AIDS virus is now universally known. [The story of how the naming issue was resolved will soon be covered in a separate posting.]

The fact that HTLV-III was identical to LAV, taken together with subsequent events, ultimately resulted in Gallo’s integrity being questioned and his reputation being compromised. We begin this part of our tale at a September 1983 Cold Spring Harbor meeting, ostensibly organized to discuss retroviruses in human leukemias. At this meeting, Montagnier reported isolating LAV from three AIDS patients; a homosexual, a hemophiliac, and a Haitian. Moreover, Montagnier also pointed up key differences between LAV and HTLV-I and -II. As for Gallo’s response to Montagnier’s presentation, some conference attendees described it as scornful and arrogant. In addition, in the introduction to the conference proceedings, which Gallo wrote, he brought up HTLV-III, although he never actually spoke about HTLV-III at the meeting. And, apropos the two meanings of “L” in the HTLV acronym noted above, it was in Gallo’s introduction to these proceedings that he subtly changed the meaning of “L” from “leukemia” to “lymphotropic.”

Next, in April 1984, Margaret Heckler, President Ronald Reagan’s Health and Human Services Secretary, hastily called a press conference to publicly announce that Gallo had discovered the AIDS virus. Heckler then introduced Gallo, who confirmed the discovery, while neglecting to mention that Montagnier had isolated the same virus. Gallo also managed to avoid questions from reporters who were primed to raise this issue. [In an interesting sidelight, Heckler confidently announced to the press that an AIDS vaccine would be available within two-years-time, leaving every scientist in the room aghast. Her rash optimism regarding an AIDS vaccine may well have been based on the earlier successes of Jonas Salk and Albert Sabin, who developed the killed and attenuated polio vaccines, respectively. (Salk and Sabin, like Gallo and Montagnier, were also bitter rivals; the topic of a future post.) Some have suggested that Heckler’s prediction of the vaccine was meant to distract attention from Reagan’s earlier silence on AIDS. The U.S. government indeed appeared to be indifferent to what was perceived by the public as a gay disease.6]

On the same day that Gallo announced his discovery of the AIDS virus at the press conference noted above, he also filed a U.S. patent application for a blood test that would detect signs of the virus in people. Gallo’s patent application became another sore point in his controversy with Montagnier, since the latter charged that Gallo’s blood test made use of a virus that was isolated at the Pasteur Institute. And, considering that the patent was estimated to be worth about $100 million per year, even the governments of the United States and France weighed in on the dispute. In fact, to end the disagreement over patent rights to the blood test, and so enable the U.S. and France to share proceeds from the patent equally, U.S. President Ronald Reagan and French Prime Minister Jacques Chirac signed a declaration that Gallo and Monatagnier were co-discoverers of the virus.

Here now is an example of politics intruding on science. As a condition of the agreement signed by Reagan and Chirac, Gallo and Montagnier had to write a history of the discovery of HIV that was in the accord with the agreement. What’s more, Gallo and Montagnier were forbidden from later publishing any statement that might undermine the agreement. Nevertheless, and irrespective of the political settlement of the patent rights to the blood test, the patent dispute also worked to undermine Gallo’s standing, not only because of persisting questions regarding the origins of the virus on which the test was based, but also because some of Gallo’s critics contend that his patent claim delayed use of the blood test for a year.

Considering that the undermining of Gallo’s standing began with the finding that his HTLV-III was virtually identical to a virus isolated in Montagnier’s laboratory, how did it happen that isolates of a highly mutable virus, from laboratories more than 3,000 miles apart, were virtually identical? Here is what many believe to have been the likely scenario. After Montagnier isolated Bru in Paris, he then isolated HIV from biopsies of several other AIDS patients. One of these isolates, called “Lai,” replicated much more rapidly than Bru, as well as other HIV isolates, in cell culture. Unbeknownst to Montagnier, Lai then contaminated and overgrew stocks of Bru in his laboratory. Then, Montagnier sent a sample of Bru to Gallo that unknowingly was contaminated with Lai. Next, Lai contaminated the culture that Gallo’s research group thought contained a virus that originated in their pool of AIDS patient biopsies. 7

Apropos the above, such mix-ups are not uncommon (a warning to beginning researchers). In fact, Montagnier also sent Lai-contaminated LAV samples to several other laboratories, and Lai likewise contaminated cell cultures in those laboratories. But, before entirely absolving Gallo of any culpability, we well might ask why he did not compare his HTLV-III to the sample that Montagnier had sent him, before announcing that he had discovered a new virus.

The virtual certainty, that a Nobel Prize would go to the scientist recognized as the discoverer of the AIDS virus, was for sure a major factor behind the bitter rivalry between Gallo and Montagnier. As it was, in 2008, 25 years after the first article describing HIV and its causal link to AIDS, 8 the Nobel Prize for Physiology or Medicine was awarded to Luc Montagnier and his co-worker, Francoise Barre-Sinoussi, for discovering the AIDS virus. Harold Zur Hausen shared in the award for his work identifying human papilloma viruses as the cause of cervical carcinoma. Gallo was not included in the award.

Was the decision of the Nobel Committee to exclude Gallo from the award correct? The Committee stated that Barre-Sinoussu and Montagnier “made the most important contributions to the discovery.” The Committee did acknowledge Gallo’s “detection of a novel…virus from a vast number of patients with AIDS or pre-AIDS in 1984…[which] showed considerable similarity with LAV-1.” Those findings of Gallo, taken alone, may not have justified a Nobel award. Importantly, however, the Nobel Committee did not acknowledge that Gallo’s group had been responsible for much of the basic research that made the discovery of HIV achievable. As noted above, Gallo’s group discovered IL-2, which made it possible to grow T cells in culture and, consequently, HIV as well. [One source I came across claimed that Gallo himself was unimpressed by Doris Morgan’s discovery of IL-2, and discouraged her from working on it, and that Gallo did not see any value in growing T cells.] Moreover, these breakthroughs enabled Gallo’s group to also isolate HTLV-1 and HTLV-II, thereby demonstrating the existence of human retroviruses and, what’s more, the feasibility of isolating them. And it was Gallo who first suggested that AIDS might be caused by a retrovirus. Furthermore, Gallo’s group was also the first to grow HIV in an established T-cell line, which was crucial to the development of the blood test for HIV. Additionally, Gallo’s group provided the more definitive evidence that HIV is indeed the etiologic agent of AIDS, as based on their repeated isolation of HIV from patients with AIDS and, subsequently, by means of the blood test.  Also note that Montagnier was quick to acknowledge that Gallo deserved the award as much as he and Barre-Sinoussi.

Gallo said it was a “disappointment” not to be included in the Nobel award, but he affirmed that all three of the recipients deserved the honor. Jay Levy, at the University of California, San Francisco (UCSF), is also recognized as a co-discoverer of HIV, which he originally termed the AIDS-associated retrovirus, or ARV. Levy is not as well known as Gallo and Montagnier, in part because he was not involved in their controversy. Levy reacted to being passed over by the Nobel Committee with the gracious comment: “In the end, what they (the Nobel Committee) did was quite, quite fair…And I congratulate them (Montagnier, Barre-Sinoussi, and Zur Hausen).”

Considering the importance of Gallo’s ground-breaking work, what might really have been behind the Nobel committee’s decision to exclude him from the award? Even if the Nobel committee did not regard Gallo’s contributions as equal to those of Montagnier and Barre-Sinoussi, weren’t they still worthy of the Nobel Prize?

Questions concerning Gallo’s integrity may have worked against him in the eyes of the Nobel committee; most importantly those arising from the virtual identity of Gallo’s HTLV-III and Montagnier’s earlier LAV-1 isolate. Other researchers also had concerns regarding Gallo’s ethics. Consequently, in 1990, to get to the bottom of the origin of HTLV-III, the Office of Scientific Integrity at the National Institutes of Health authorized a group at Hoffmann-La Roche to analyze HIV samples isolated in the laboratories of Gallo and Montagnier between 1983 and 1985. The conclusion of the Roche group, published in Nature in 1993, was that Gallo’s HTLV-III indeed had originated in Montagnier’s laboratory. 9 But, the group also concluded that the initial mix-up, Lai in place of Bru, occurred in Montagnier’s laboratory. The Lai-contaminated sample that Montagnier then sent to Gallo subsequently may have contaminated the culture that Gallo was working with in his laboratory.

In the end, the Roche investigating team dropped all charges against Gallo. However, before publication of their findings, Gallo’s group was found guilty of “minor misconduct” by the Office of Scientific Integrity in 1991. Thus, while the Roche team cleared Gallo of all charges of misconduct, his reputation had already been tarnished by the accusations against him. Moreover, questions still remained, particularly those pertaining to Gallo having grown Montagnier’s LAV in his own laboratory, before he reported isolating HTLV-III. What’s more, there is evidence that a micrograph published by Gallo that is said to show HTLV-III, actually depicts Montagnier’s LAV.

A second reason suggested for the Nobel committee’s slight to Gallo concerns his ego and fiercely competitive nature. Still, while Gallo’s personality may not have endeared him to the Nobel committee, it most certainly should not have precluded his contributions from being recognized by them.

Here then is another possible take on the Nobel committee’s decision to leave Gallo out of the award, as noted at the time by Anthony S. Fauci, director of the National Institute of Allergy and Infectious Diseases. “The committee has a long history of awarding the prize to the person or group that makes the first seminal observation or discovery, and they did that in this case.” Hence, in the end, it may simply have come down to who the committee considered to be the actual discoverer of HIV.

So, who actually discovered the AIDS virus? The answer is that HIV was first isolated by Françoise Barré-Sinoussi in Montagnier’s laboratory at the Pasteur Institute, in collaboration with other French clinicians and researchers, including Jean-Claude Chermann, Willy Rozenbaum, David Klatzmann and, of course, Montagnier. They published their findings in Science, in May 2003; about a year before anyone else. 8 Jean Claude Chermann, the second author of the Science paper, is considered by many to be equally deserving of the Nobel Award. Chermann supervised Barre-Sinoussi in Montagnier’s laboratory, and had the idea of focusing efforts to isolate HIV on patients with lymphadenopathy.

This remarkable photograph was taken in the park of the Institut Pasteur Annex in Garches, near Paris, during a break of a "100 guards meeting" in 1987. From left to right: Jonas Salk, Jean- Claude Gluckman, Jean-Claude Chermann, Luc Montagnier, Robert Gallo, Françoise Barré-Sinoussi, Willy Rozenbaum, Charles Mérieux.
This remarkable photograph was taken in the park of the Institut Pasteur Annex in Garches, near Paris, during a break of a “100 guards meeting” in 1987. From left to right: Jonas Salk, Jean- Claude Gluckman, Jean-Claude Chermann, Luc Montagnier, Robert Gallo, Françoise Barré-Sinoussi, Willy Rozenbaum, and Charles Mérieux.

Also note that, in 1985, Montagnier’s research group, in collaboration with physicians in Lisbon and virologists from Hopital Claude Bernard in Paris, also discovered HIV-2 (which they initially dubbed LAV-II) in West African patients with AIDS. Concurrent with the above efforts, Jay Levy and colleagues at UCSF demonstrated that HIV is present in AIDS patients and in healthy carriers as well.

Still, consider the following. First, the issue of which research group was the first to isolate HIV was resolved by the early 1990s. Second, Howard Temin and David Baltimore had to wait a mere five years after announcing their discovery of reverse transcriptase before receiving their Nobel Prizes. 2 So, bearing in mind the enormous significance of the discovery of HIV, why did 25 years elapse before the Nobel committee rewarded that discovery? Can it be that it was concerned with, and needed to resolve some of the ethical issues noted above? Or, was it simply that the Nobel committee tends to steer clear of controversies? Since the Nobel committee’s deliberations are shrouded in secrecy, we can only speculate on the reason for the 25-year hiatus, and why Gallo was excluded from the prize.

Next, bearing in mind Gallo’s extensive experience as a retrovirologist and that only his group had ever isolated a human retrovirus, as well as all of the resources available to him at the NIH, why didn’t he succeed in isolating the AIDS virus ahead of Montagnier?  Was it because he was fixed on the notion that AIDS is caused by a virus closely related to HTLV-I and –II? Indeed, until May 1983, Gallo was looking only for, and reporting only isolates that were like the HTLVs. So, perhaps there is the irony that if Gallo’s group had not discovered HTLV-I and -II, it might well have been the first to discover HIV.

The controversy between Gallo and Montagnier has long since subsided (although some sources state that the animosity between them remains), and they appear to be in agreement on all major issues. For his part, Gallo has stated that he never claimed to have discovered HIV, but rather claims credit for demonstrating that it is the cause of AIDS. Montagnier concedes that Gallo’s evidence in that regard was more convincing than his own. Regardless, efforts of these two individuals resulted in the identification of a new retrovirus as the cause of AIDS, and made it possible to grow large enough amounts of the virus to enable further studies. Moreover, their discovery quickly resulted in a blood test for HIV, and opened up the development of anti-retroviral drug therapies for HIV-positive individuals.

One fundamental lesson learned from these experiences was well stated in a report jointly written by Gallo and Montagnier: “Our experience with AIDS underscores the importance of basic research, which gave us the technical and conceptual tools to find the cause less than three years after the disease was first described.” 3

1 Doris Morgan, working in Gallo’s laboratory, discovered a T-cell growth factor that enabled her to grow T lymphocytes in culture for extended periods. Kendall Smith, at Dartmouth, followed up Morgan’s observations, isolating interleukin-2 (IL-2) as the T-cell growth factor that Morgan detected.

2 Howard Temin: In from the Cold (on the blog)

3 Gallo, R C., and L. Montagnier, (2003) The Discovery of HIV as the Cause of AIDS, N. Engl. J. Med. 349:2283-2285.

 4 Luc Montagnier-Biographical, at Nobelprize.org, The official web site of the Nobel Prize.

5 Mikulas Popovic, in Gallo’s laboratory, proposed isolating the virus from a pool of 10 different AIDS patient biopsies. His reason was that the pool should yield the most viable virus, by a process akin to natural selection.

6 Even the media shied away from covering the AIDS epidemic during its early years. When news stories about AIDS did appear in newspapers, they tended to be buried in the back pages, and AIDS stories were seldom reported on television. This was largely because it was difficult for the media of the day to talk openly and honestly about sex; particularly gay sex. The watershed event that changed this state of affairs was actually the July 1985 disclosure by movie star Rock Hudson that he was suffering from AIDS. Hudson was the first major public figure to reveal that he had AIDS, and his celebrity status put AIDS on the front page. The press now covered AIDS with gusto, and they had photos of Hudson to add pizzazz to their stories.

Younger readers may get an accurate glimpse of the homophobia and public attitudes towards AIDS during the early years of the AIDS epidemic from the 1993 movie Philadelphia, which often appears on TV. The main character, played by Tom Hanks, is a brilliant lawyer who is set up to be fired from his prestigious Philadelphia law firm when it discovers that he has AIDS. He then sues the firm, basing his case on the Americans with Disabilities Act of 1990, which prohibits discrimination against any individual with a disability, including those living with HIV/AIDS.

7 Slow-growing HIV isolates like Bru tend to be present at early stages of HIV infection, whereas rapidly growing viruses like Lai are seen in late stage infection. Also, the slow-growing isolates like Bru are not readily transmissible to permanent T cell lines, whereas fast-growing isolates like Lai are. This was important in the current context, since only some viral isolates from patients with fully developed AIDS could be grown in permanent T cell lines, as soon would be learned. The fast-growing strains also induce the formation of large syncytia. For details on the relevance of these points to infection in vivo, see Virology: Molecular Biology and Pathogenesis.

8 Barré-Sinoussi, F., J.C. Chermann, F. Rey, M.T., Nugeyre, S., Chamaret, J., Gruest, C Dauguet, C. Axler-Blin, F. Vézinet-Brun, C. Rouzioux, W. Rozenbaum, and L. Montagnier, (1983) Isolation of a T-Lymphotropic Retrovirus from a Patient at Risk for Acquired Immune Deficiency Syndrome (AIDS), Science 220: 868-871.

9 Chang, S-Y. P., B. H. Bowman, J. B. Weiss, R. E. Garcia, andT. J. White, (1993) The origin of HIV-1 isolate HTLV-IIIB, Nature 363: 466–469.