Re: Re: Where is the proof for HIV purification by any method? 27 June 2003
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Brian T Foley,
HIV Researcher
Los Alamos National Lab, Los Alamos NM 87545

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Re: Re: Re: Where is the proof for HIV purification by any method?

The Perth group stated:
"Barre-Sinoussi and Chermann, the principal and the 2nd author of the 1983 Montagnier et al paper who, in 1973, used it to purify the murine sarcoma virus and to obtain material which contained nothing else but particles with “No apparent differences in physical appearance” (1)"

In fact, this paper states on 240 in Results:
"the particles isolated under conditions of rate sedimentation considered [sic] mainly of virus particles and very little amounts of contaminating material."

Furthermore, in the Discussion on page 243, they correctly note that what they have purified or enriched is not a single virus, but a mixture of at least two species of viruses:
"It is known that stocks of MSV consist of at least two different viral particles, muringe leukemia viruses (MLV) and defective murine sarcoma viruses (MSV) (12)."

This is because the Moloney Murine Sarcoma Virus has had its envelope gene replaced by the c-mos protooncogene which encodes a serine/threonine kinase. Without an env gene, the Mo-MSV is defective and cannot infect cells unless it is packaged into virions in which the env protein is provided by a helper virus which in this case is the murine leukemia virus.

Rodent genomes contain dozens of copies of endogenous retroviruses, some of which are nearly identical to the Moloney Murine Leukemia Virus. In addition, there are dozens of exogenous rodent retroviruses such as the Friend, Kirsten, SL3 and Abelson murine leukemia viruses. Whether or not these are all truly "different" viruses, or just different isolates of the same virus, is an open question. The point I want to make is that Sinoussi et al. did get viral particles that looked identical to each other by electron microscopy, but without molecular clones, sequences, or serological profiles they could only take the word of the people they obtained their 78A rat fibroblasts from, that these particles were derived from the Moloney strain of MSV and not the Kirsten strain or some endogenous mouse or rat retrovirus that had become activated. All of these retroviruses look alike.

In the 1980s, when cloning and sequencing and other techniques became available, many of these exogenous and endogenous, defective and competent viruses were cloned and sequenced. With this new genomic information, it became clear why the Sarcoma viruses cause various sarcomas and require a helper virus. For example, the Moloney Murine Sarcoma virus has no env gene, it has been replaced by the c-mos gene (which is called the v-mos gene when it is in the virus genome).

The other example of 100% pure virus that the Perth group cites is a paper by Crawford and Crawford on Rous Sarcoma virus (now known to be one of several strains of the Avian Leukosis Virus
Bieth E, Darlix JL. Complete nucleotide sequence of a highly infectious avian leukosis virus. Nucleic Acids Res. 1992 Jan 25;20(2):367. PMID: 1311072 )
These authors used a culture of "high titer strain" of RSV given to them by Dr. H Rubin. Again, they had to take the word of the donor that this was indeed the high titer strain of a virus charachterized by Peyton Rous in the 1908 to 1915 time period and further passaged by Bryan and Moloney in the 1950s to create a "high titer strain". By electron microscopy, it looks like a retrovirus but there is no way to tell if it is RSV or some other avian retrovirus. Again, in this paper, the authors do not state that the virus preparations are 100% pure. In fact they state on page 229 "The third preparation was RSV from a tumor wich has been purified by density gradient centrifugation. Such preparations contain much nonviral material, and counts of physical particles are therefore less accurate." and "the preparation is homogeneous and little nonviral material is evedent". This is similar to preparations of HIV-1 or HIV-2 which have been electron micrographed: some preparations contain less cellular debris than others but a little cellular debris is inevitable in this type of preparation.

The Rous Sarcoma Virus paper also states "DNA was barely detectable in the preparation; it would be equivalent to 2 x 10^5 molecular weight units per particle if it were actually present in the virus." again showing that this was not 100% "pure" virus and nothing else.

I fail to see why these two papers which describe centrifugation of retroviruses are accepted by the Perth group as meeting all of their criteria for isolation and purification of a virus, when similar papers describing the same procedures carried out with HIV-1 or HIV-2 are dismissed. It is especially irritating to me that they like the Moloney Murine Sarcoma Virus paper even when the authors state that the prepared virus is a mixture of at least two different viruses; the Moloney MSV and its MLV helper virus.

Competing interests:   None declared