Dharam V. Ablashi is an American biomedical researcher born in India. He is best known for his co-discovery of Human herpesvirus 6 (HHV-6), an immunosuppressive and neurotropic virus that can cause encephalitis and seizures during a primary infection or when reactivated from latency in immunosuppressed patients. He spent 23 years working at the NIH while additionally serving as an adjunct professor at Georgetown University School of Medicine. Ablashi’s research has also helped clarify the pathogenic role of HHV6-A and HHV6-B in neurological disorders such as multiple sclerosis, epilepsy, ME/CFS, and in tumors such as Hodgkin’s disease lymphomas and other brain tumors. He has co-authored 96 articles on HHV-6 and co-edited three books on HHV-6. He is also internationally known for his research on Human and Simian herpesviruses and has also been a major contributor to research on HIV, Epstein Barr Virus (EBV) and the field of immunovirology. He has served as the Scientific Director of the HHV-6 Foundation since 2004.

How did you discover HHV6? What led you to begin studying the virus?

It was late 1984 when I met Zaki Salahuddin and Dr. Robert Gallo at the National Cancer Institute (NCI). At that time, I was a member of another branch of NCI working in the same building where Dr. Gallo’s laboratory was located. They had found EBV in two-thirds of B-cell lymphomas of HIV patients. Gallo suspected there must be another herpesvirus causing cancer in the rest of the patients and asked us to look for it.  So, that was the beginning of our discovery.

We cultured the spleen cells and lymphocytes and stimulated with PHA from the B-cell lymphoma AIDS patients, and we saw very large cells (juicy cells-see image) appearing. When we sent these cells to get electron microscopy, we found that there was a herpesvirus. I thought I had isolated an EBV virus, but none of the monoclonal antibodies from other human herpesviruses were positive. At that time, we named it human B lymphotropic virus and determined that it was highly cell associated, spreading from cell to cell.  Of 8 isolates from HIV patients with AIDS lymphomas, all were HHV-6A, not HHV-6B.

NIAID investigator Dr. Paolo Lusso, who also was in Dr. Gallo’s lab as a post-doc, tested these cells and showed that these infected juicy cells were T-cells. That is when we changed the name to HHV-6 in 1987. Then in 2013 HHV-6 was reclassified as HHV-6A, and HHV-6B.

We also discovered HHV-7. However, we decided to characterize it more before publishing and, and Dr. Niza Frankel beat us to publication.  Dr. Gallo was very upset on that one!

What was it like to work with Robert Gallo? Do you have any interesting stories about working in that lab?

My relationship with Dr. Gallo developed quite well.  In fact, I considered him to be my mentor, and he respected my training and knowledge working with herpesviruses.  He was very hard to please but respected those who produced solid data.  He expected us to work very hard and would often call for meetings on weekends.

Ironically Bob Gallo helped kill funding for HHV-6 research in the early 1990s. At that time, HHV-6A was suspected as a possible co-factor in AIDS progression, because HHV-6 also infects CD4 T cells and a co-infection was thought to hasten the depletion of CD4 T cells in AIDS. Then a vocal AIDS activist and publisher of the New York Native started claiming HHV-6A was the primary cause of AIDS and that Gallo was covering up this “truth.”  Gallo became irritated and started proclaiming HHV-6 to be a benign virus, even though he knew it was not true, in order to get them to stop. He admitted to me years later that he feels guilty about this. It is a great example of how uninformed activism can be quite damaging. This individual is still accusing Gallo and myself of a cover-up, threatens protests outside our small scientific conferences, and accuses our speakers of “covering up the true cause of AIDS.”

You earned degrees in India, the UK, and the USA. Do you think this international education had an impact on how you approach research challenges?

My education in India was based on British teaching, so I had my DVM, and then when I came to the US, the education system was completely different.  I did all the work for a Ph.D., but could not pass the language requirement at the time, so gave up. That same institution, University of Rhode Island, recently gave me an honorary doctorate which was very kind of them. Most of my education came from long hours of hard work in the laboratory.

You’ve received many awards for your HHV6 research. Which are you most proud of and why?

In 2006, at the International Conference on HHV-6 and HHV-7 held in Barcelona, Spain, the keynote speaker said he was very proud to present me with the Lifetime Achievement Award on behalf of the HHV-6 Foundation. This award was named the Dharam Ablashi Lifetime Achievement Award, and I was the first recipient.  I am very proud of this award.  There have been many other ones, and I remember one given to me on the Rudy Perpich Memorial Achievement Award for Chronic Fatigue Syndrome (CFS) research.

What has been the greatest challenge you’ve faced in pushing research on HHV6 forward?

One reason was due to the confusion caused by chromosomally integrated HHV-6. Once physicians saw occasional asymptomatic patients with extremely high viral loads (not understanding that this was an inherited condition), they assumed that the virus must be benign. Ironically Bob Gallo also admitted to me that he helped kill interest in HHV-6 research. HHV-6A was suspected as a possible co-factor in AIDS progression (working with HIV to deplete CD4 T cells) but when AIDS activists started claiming it was the primary cause of AIDS, Gallo started saying that HHV-6 was unimportant, just to get them to stop accusing him of covering up the “true” cause of AIDS.

The other challenge is that HHV-6 is very difficult to grow because infected cells will die very quickly. It is much more difficult than other herpesviruses, and as a result, there are very few laboratory scientists currently who understand the right techniques to even grow the virus.

HHV6 infection is connected to a range of chronic conditions that are deemed “autoimmune” or “of unknown cause.” Do you think that’s accurate? Do you think the scientific community needs to take HHV6 infection more seriously in many of these disease states?

Several researchers from diverse fields have found a connection between HHV-6A and Multiple Sclerosis (MS) as well as autoimmune thyroid disease. HHV-6B reactivation occurs in most severe DRESS/DIHS cases, and many of those patients go on to develop autoimmune diseases such as diabetes type 1. HHV-6A has been found in over 80% of Hashimoto’s thyroiditis patients vs 10% of controls, and HHV-6 is found at high levels in the islet cells of patients with diabetes type 1. The mechanisms require further study.

DRESS expert Chia-Yu Chu in Taiwan believes that IP-10 may be the link between HHV-6 reactivation in DRESS and subsequent autoimmunity. A group in Italy has found that HHV-6 infection of thyrocytes and T cells alters the expression of miRNA in a pattern similar to that of autoimmune thyroiditis. The scientific community needs to follow up on these important potential associations.

What research team do you think is currently doing the best research on HHV6, and why?

The Japanese have led the way in molecular biology. Virologist Koichi Yamanishi was the first to associate HHV-6B with roseola in 1998, and his protégé Yasuko Mori has contributed enormously to basic science, including valuable data on cell entry, and identifying CD134 as the receptor for HHV-6B.  Transplant specialists such as Masao Ogata in Japan have also done valuable work in characterizing HHV-6B encephalitis, and Japan is the first country in the world to have a drug approved specifically to treat this condition (foscarnet). Japanese dermatologists have done valuable characterizing of frequent HHV-6B reactivation in drug-induced hypersensitivity syndrome, a phenomenon that has been mostly ignored in the US. Finally, Kazuhiro Kondo has recently published a study tying HHV-6B latency protein to a 12-fold increased risk of depression as well as increased activity of the HPA axis.

In the US, very little research has been done on HHV-6 compared to Europe and Japan. Over the past several decades, only one investigator at NIH has consistently studied HHV-6 in neurological disease (Steve Jacobson, NINDS) who showed that HHV-6A accelerates neuroinflammation in a marmoset model of MS.  The only academic center that has demonstrated a sustained interest in HHV-6 has been at the University of Washington led by Danielle Zerr (Seattle Children’s Hospital) and Michael Boeckh, Keith Jerome, Joshua Hill, and Alex Greninger (Fred Hutch). They have characterized HHV-6B in children, found increased CNS dysfunction in transplant patients with HHV-6 reactivation, revealed increased graft vs host disease in stem cell transplant patients with HHV-6 reactivation, as well as with chromosomally integrated HHV-6.

What are the top mistakes that research teams can make when trying to study HHV6?

Many groups studying HHV-6A/B in chronic conditions want to look in the plasma or cerebrospinal fluid (CSF), and fail to understand that HHV-6 DNA appears in those compartments only briefly and typically only during acute reactivations. These are low copy number viruses that spread cell-to-cell. They also generate potent chemokines and cytokines, even in latency.

There may be low-level DNA detectable by nested PCR in a chronic case, but generally, qPCR, ddPCR, and RNA sequencing techniques are not able to detect low level central nervous system infections of HHV-6A/B in plasma in ME/CFS and MS patients. HHV-6 DNA levels can be extremely high in the brain, liver, or lung, with barely a trace in the plasma. So biopsy analysis is extremely important. Also, HHV-6A/B viral infections have scattered foci, multiple samples per biopsy are necessary to get a true understanding of prevalence in any organ.

Another mistake is to look at DNA in whole blood, where viruses like HHV-6B and EBV can be found in latent form. For HHV-6, this is too simplistic. It is more important to identify organ tissues where there might be active infection, or a smoldering infection – a latent infection that is still throwing off inflammatory cytokines and chemokines.

Finally, many studies fail to rule out inherited chromosomally integrated HHV-6, which occurs in 0.86% of the US population controls and 1.5 – 2% of patients. These patients will always be positive for HHV-6 DNA in the plasma and CSF, even if asymptomatic, so this condition needs to be determined. (This is easy to do by measuring HHV-6 DNA in whole blood by qPCR because there is one genome per nucleated cell, so the viral load is typically in the millions per mL.)

What role do you think HHV6 plays in the illness myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS)?

We don’t think ME/CFS is triggered by a single pathogen; we believe it can be triggered by a number of intracellular pathogens including enterovirus, parvovirus B-19, mycoplasma pneumonia, chlamydia pneumonia, EBV, Cytomegalovirus (CMV), and HHV-6A/B and HHV-7. After the Dan Peterson and Paul Cheney first discovered HHV-6 in patients with the outbreak of ME/CFS in Incline Village Nevada in the 1980s, Dan Peterson, Anthony Komaroff, and Deidra Buchwald came to see Dr. Gallo to discuss whether HHV-6 played a role in ME/CFS.  At their request, we tested the antibodies of sera or plasma from ME/CFS patients.  During this process we found a subset of samples with very high antibody titers to HHV-6.  Buchwald, Komaroff, and associates published a paper in 1992 that found used primary cell culture to determine that 70% of ME/CFS patients but only 20% of controls showed signs of active replication.  Later in 2001, I published another paper that showed an increased level of IgM antibodies to an HHV-6 early antigen protein in ME/CFS patients. This was an assay based on a reagent produced at Gary Pearson’s lab at Georgetown, and unfortunately, this reagent is no longer available. Recently Dr. Bhupesh Prusty in Germany has shown how persistent HHV-6 infection can cause mitochondrial dysfunction, which may play a role in ME/CFS.  Also, Dr. Kondo recently published very interesting data showing that a neurovirulent HHV6 latency protein plays a pathogenic role in both depression and fatigue. He previously determined that HHV-6 builds up in the saliva when the body is fatigued and says this virus finds its way to the nasal passages and then on to the olfactory bulb and brain. A latency protein he calls SITH-1 then causes hyperactivation of the HPA axis and subsequent depression and fatigue.

What are your thoughts on HHV6 and Alzheimer’s disease?

Again, we believe that infections are the most logical explanation for the inflammation neurodegeneration associated with Alzheimer’s, but don’t believe that there is a single pathogen responsible. HHV-6A and HHV-7 were identified as likely culprits in 2018 paper, using RNAseq analysis by Readhead and team. Two subsequent studies did not find an increase in HHV-6/7 using the same data, but at the same time did not find a significant presence of ANY herpesviruses in the brain, which we know is not the case. Dozens of careful studies have identified a high prevalence of low-level HHV-6 and HSV1 DNA in brain tissues. This tells us that RNAseq and ddPCR methods employed to date are not sufficiently sensitive to really answer the question of whether these viruses play a role in Alzheimer’s. Future studies should be done with both extremely sensitive assay as well as multiple samples from each brain region to answer the question of HHV-6A/B prevalence in Alzheimer’s samples vs. controls. In 2002, Ruth Itzhaki’s group looked for HHV-6 and HSV1 by nested PCR and found HHV-6 DNA in 70% of Alzheimer’s brains vs. 40% of controls. They found HSV1 at high levels in both AD patients and controls. Old-fashioned nested PCR is time-consuming and complicated because measures must be taken to prevent contamination, but this old-school technique may still be a better approach than newer techniques (such as RNAseq analyses) that can find little to no herpesvirus in these brains. The other issue is that infections may be early triggering events that disappear over time. It will take a lot of time and a solid investment in research to find any answers.

Are there any key experiments that you think should be conducted on HHV6 that haven’t happened yet?

The NIH has dedicated several hundreds of million dollars each toward the study of CMV, HHV-8 and EBV over the past 20 years, but almost nothing for HHV-6. As a result, we have a long way to go in understanding much of the basic science behind HHV-6 that would allow more sophisticated analysis. For example, a great deal of effort has gone into characterizing EBV early and immediate early proteins, and there are assays that can measure EBV early antigen and EBNA antibodies. We don’t have those tools for HHV-6. This is a complicated virus, and it will require significant funding to unravel disease associations.

One simple clinical study that is at the top of our list is a clinical trial to determine if antiviral treatment would benefit infants with HHV-6B induced febrile status epilepticus (FSE). A large study in 2012 confirmed that 32% of infants with FSE have active HHV-6B, yet for reasons we don’t understand, these infants are not given an antiviral and are treated only with anti-seizure drugs. Around 30% of these infants eventually develop epilepsy, and many of them suffer from cognitive dysfunction.

You describe HHV6 as an immunosuppressive virus. Can you explain why you’ve come to that conclusion and describe some key research findings that led to that understanding?

All three roseoloviruses (HHV-6A, HHV-6B, HHV-7) are immunosuppressive, and animal homologues such as murine and porcine roseolovirus are immunosuppressive as well. HHV-6 preferentially infects CD4 T lymphocytes and is associated with delayed engraftment in transplant patients. HHV-6 reactivation in transplant patients is also associated with bone marrow suppression, graft rejection and increased bacterial and fungal infections. Transplant patients with HHV-6B reactivation are 15X more likely to develop a CMV infection after reactivating with HHV-6. Severe cases of primary HHV-6B infection are tied to neutropenia.

Do you think that HHV6 can act along with other pathogens to drive chronic disease processes? If yes, how might this happen (for example can HHV6 support HIV survival and vice versa)?

Yes, but the mechanism is likely to be indirect, and not due to active replication. HHV-6A/B can generate inflammatory cytokines, chemokines and virokines even in latency. This causes chemoattraction of leukocytes which in turn triggers inflammation. In addition, there appears to be a state of abortive infection wherein some early proteins are expressed in the absence of full replication. HHV-6 is actually likely to reduce HIV patient survival by accelerating the depletion of CD4 and CD8 T cells. HHV-6A was shown to accelerate AIDS progression in macaques.  Macaques co-infected with simian HIV and HHV-6A died quickly and showed a dramatic depletion in both CD4+ and CD8+ T cells, whereas those infected with only simian HIV never progressed to full blown AIDS.

What advice do you have for young researchers starting to learn about and study HHV6?

For decades scientists have avoided HHV-6 research because the NIH was not funding HHV-6 related grants. As a result, there is a lot of opportunity to make important clinical and scientific discoveries, with very little competition. My advice is to be courageous and aim for a major discovery in an emerging field, rather than making a marginal contribution to a well-studied pathogen.