Brian T Foley,
Los Alamos National Lab, Los Alamos, NM 87545
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The Perth group wrote:
The Perth group has repeatedly been told that this is not true; that genes, whether they come from viruses or any other organism, can indeed be cloned without 100% “purification” of the protein or organism which they encode. How, for example, would it be possible to clone, sequence and analyze the human insulin receptor gene, if the silly rules invented by the Perth group were a requirement?
The Perth group has invented some rules which are impossible to carry out, and then they claim that if their rules are not followed then any results are not “proven”. If I claim that I once read a paper that claimed that computer viruses must be found in the boot sector of a 5 and ¼ inch floppy disk, would my claim cast any real doubt on the existence of the existence of the ”Green Caterpillar” computer virus? I am a biologist who knows very little about computer viruses, why would anyone believe me, when they could ask computer virus experts if indeed all computer viruses are required to be found in the boot sector of a floppy disk? It turns out that if you ask virologists from any field of virology, whether it be plant viruses, bird viruses, fish viruses or mammalian viruses, whether or not HIV-1 infectious molecular clones have been properly created and studied, more than 99.9% of them will agree that they have been. Even Peter Duesberg, a respected leader in HIV denialism, agrees that many HIVs and SIVs have been properly isolated.
As far as I know, no virus has ever been put through the series of steps described by the Perth group as being “required” to “prove” the existence of a virus. When I asked the Perth group to name a virus that they thought had met their criteria, they named (June 26, 2003) two retroviruses that were proven by molecular analyses (serology, cloning and sequencing, etc.) to be mixtures of at least 2 viruses. Anyone who knows a bit of history of RNA tumor viruses would have known that a highly oncogenic virus cannot be “purified” by the Perth group method because these viruses have picked up cellular oncogenes by exchange for a portion of the viral genome. These oncoviruses are thus replication defective and require a “helper virus” for propogation.
On June 19, 2003 the Perth group wrote:
I have never said that banding viruses on density gradients is impossible. I have only stated that it is not required that the band of virus be 100% “pure”, free of absolutely all contaminating materials, and composed of viral particles that are all “identical” by appearance in electron micrographs. Density gradient centrifugation is indeed a useful technique, and has indeed been used in many key experiments on HIV-1, HIV-2, SIVs and other lentiviruses. Even if the virus particles were 100% “pure” after retrieval from the 1.16 g/ml band, they could still have many human proteins embedded in them, in addition to viral proteins. For example, the HIV-1 particles are known to have human HLA molecules on their surface if they are grown in human cells, and chimpanzee HLA molecules on their surface if they are grown in chimpanzee cells prior to centrifugation. It is for this reason that cloned viral genomes are useful for production of viral antigens for EIA and western blot reagents. Use of pure HIV proteins with no cellular protein contamination is beneficial.
The Perth group has quite convincingly shown that they are not interested in learning anything about virology or molecular genetics. When it is convenient for them, they claim that they trust the experts. For example, they claim to have absolute faith that Francoise Barre-Sinoussi and Jean-Claude Chermann were correct when they spoke at some meeting in 1972. But when these same virologists worked with HIV-1 M group subtype B virus from AIDS patients 11 years later the Perth group dismisses their work because it is not convenient for their theory that oxidative stress is the real cause of AIDS. Likewise, if the Perth group thinks they can confuse people about lentiviral genomic variability by claiming that some “expert” said that no RNA virus can vary by more than 1%, they do so. But if thousands of virologists sign a document stating that they have studied HIV and that it has indeed been properly isolated, cloned and studied the Perth group feels free to dismiss it, because it is not convenient to their theory.
HIV-1 and all other lentiviruses are asymmetrical, with a cone-shaped core. Thus even 100% “pure” virus particles do not all appear “identical” by electron microscopy. Some are viewed end-on and appear round, others are viewed from the side and are more oval in cross-section. Conversely, many other retroviruses appear to be identical when they are in fact mixtures of two or more very different species.
The Perth group finds it convenient to remain ignorant of how it is that scientists know what they have cloned. When they think that they have cloned the complete gene or cDNA for any protein-encoding gene, such as the gene encoding human insulin receptor or the gene encoding the HIV-1 M group subtype B Gag protein, there are many different ways to determine if they have indeed cloned the right piece of DNA. One method is to show that antibodies raised against the original protein react to the cloned product. Another is to show that the cloned product produces the same effect in cells as the original protein.
Infectious molecular clones of lentiviruses do indeed meet all the requirements needed to prove that they are indeed clones of lentiviruses. When they are transfected into permissive cells they produce infectious viral particles. Those particles are composed of many proteins that bind to antibodies from people or animals that were infected with identical or similar viruses but not to antibodies from hosts that were infected with more distantly related viruses.
The Perth group is very clever with empty rhetoric. They have excellent skills for arguing. If they would put just 5% of those skills to use in formulating and testing hypotheses about HIV vs. oxidative stress as the cause of AIDS, they could settle this all very quickly. For example, if clones of HIV-1 are not truly derived from the exogenous lentivirus that has been found in AIDS patients (and pre-AIDS HIV-seropositive healthy people), then the molecular epidemiology of the viral sequences would not match up with the conventional epidemiology. For example, if HIV is some sort of endogenous virus that is released by oxidative stress, then the viral sequences would agree with human lineages rather than with viral lineages. A Haitian man for example would not be able to pass his virus along to a chain of Swedish women, and Europeans who visited prostitutes in Africa or Thailand would not pick up African or Thailand strains of HIV-1. If the HIV proteins used in EIA or western blot test kits are not of pure viral origin, then the results of those tests might not correlate well with AIDS or with HIV gene sequences obtained from the individuals identified as seropositive. Instead of formulating and testing such hypotheses, the Perth group entered the political arena with an attempt to convince the government of South Africa that all of AIDS research might be bogus, because perhaps the virus was not isolated.
It has been many years since I studied the papers of Gallo and Montagnier about the isolation and cloning of the very first HIV-1 genomes, so I need to dig them out and re-study them before I can comment on whether or not clearly show that they used polyadenylated RNA obtained from viral particles, banded at 1.16 g/ml in sucrose density gradients. I am sure that some researchers have used density gradient banded viruses to make their infectious genomes, while others used integrated proviral DNA harvested from infected cells. There are currently many acceptable methods of cloning viruses, and in the future it is quite likely that even better methods will become available.
Competing interests: None declared