“To date [2001], attempts at viral isolation from C135 and C49 [two people who have been HIV+ at least since a blood transfusion between 1981 and 1984] have all been negative”

“Virus was obtained by coculturing one million peripheral blood mononuclear cells (PBMCs) from these individuals with one million uninfected phytohemagglutinin (PHA)- stimulated PBMCs [peripheral blood mononuclear cells]. Cells were maintained at one million per ml by either addition of medium or addition of one million phytohemagglutinin-stimulated PBMCs per ml in fresh medium. Cultures were expanded and maintained in this manner and then tested for HIV-1 gag(p24) after about 2 weeks in culture or when the cell number did not increase. Fresh medium was added to positive cultures, and they were harvested several days later, typically after a total of 3 to 4 weeks in culture. Cultures were spun at 1,000 rpm for 5 min in a Beckman GPR centrifuge, and the supernatant was collected, passed through a 0.2-mm syringe filter [in other words, the end result is clearly not pure virus, it is not even clear that there is any virus in the resulting mixture]”

“fewer than 50% of patients with CD4 [immune cell] counts greater than 200 cells/microliter had positive plasma cultures”

“the expression of HERVs [Human Endogenous Retroviruses] has been linked with disruption of immune function, particularly associated with autoimmune disease. Reports of retrovirus-like particles in Sjogren’s syndrome and sequences in Graves disease and multiple sclerosis suggest that their expression may have considerable impact on the immune system...We detected a 6 kb mRNA and lower levels of a 4.5 kb RNA...in PHA-stimulated T cells...the transcripts are absent or undetectable in unstimulated T cells [which, by extension, could mean that HIV is an endogenous (built-in) retrovirus that is inactive until stimulated by PHA using culture techniques]”

“Testing for HIV by PCR or culture also may be helpful in determining HIV status; however, neither test is licensed for diagnosis of HIV infection.”

“15 patients had no detectable virus from plasma culture at entry; 8 of these patients remained consistently culture negative, while the other 7 had virus isolated from plasma at some point during the 12 weeks of study…During the study, 9 [patients who did not have a culture obtained at study entry] had consistently negative cultures, 1 had consistently positive cultures, and 10 had intermittently positive cultures.”

“HIV could not be isolated from the plasma of subjects with long-term nonprogressive HIV infection, but it could be isolated from lymph-node mononuclear cells (in seven patients) after coculture with phytohemagglutinin- activated mononuclear cells from an HIV-negative donor (data not shown).”

“The first report [on HIV], by Barré-Sinoussi et al gave no details of the suspension medium and the presence of virus was monitored by the use of reverse transcriptase, an indirect and relatively insensitive assay…[in our studies] The presence of infectious virus was determined by examining viral cultures every 3 to 4 days for a cytopathic effect [cell death]…Replicating virus was measured by determining the level of p24 core antigen every 7 to 10 days. [ooh, now that's better than using reverse transcriptase]”

“Although serologic assays are capable of identifying prior exposure to human immunodeficiency virus (HIV), they cannot alone demonstrate whether an individual is currently harboring the virus. The first method used to ascertain if a blood specimen contained HIV was co-cultivation with stimulated primary human lymphocytes or continuous human T cell lines and monitoring the culture supernatants for the presence of reverse transcriptase. Although viral isolation has proved to be a poor diagnostic tool because of its relative insensitivity [about half of HIV-antibody-positive people are culture negative], high costs, and lengthy time requirements, culture has served as the standard by which all other diagnostic tests have been judged and established. Furthermore, virus culture remains the steadfast route by which new variants are identified, isolated and initially characterised.”

“Data from 11 laboratories, referred to as the reference laboratories, that consistently were able to isolate virus in a pilot QA program were compared with information provided by 8 laboratories less successful in their attempts to isolate virus. Eleven other laboratories whose abilities to isolate virus were unknown were also invited to submit answers to the questionnaire.”

“After 4 days of incubation the percentage of HIV antigen containing cells was determined by immunofluorescence.”

“Virus has been repeatedly isolated [cultured] in an additional small proportion of children (2.5%) who lost maternal antibody and have remained clinically and immunologically normal [culture is considered to be the closest thing available to direct detection of the virus, so it is hard to explain why these children are not considered infected]…virus isolation [culture] has been attempted in 163 [children who lost antibody]; in 10 (6.1%), positive results were obtained at least once…In 4 children the initial positive culture was confirmed on subsequent samples by repeat virus culture and/or polymerase chain reaction (PCR)”

“Overall, plasma viremia [as measured by signals from a culture] developed during follow-up in 17 of 27 patients (63 percent) who initially did not have viremia, and was sustained in 35 (88 percent) of the 40 patients who initially had viremia”

“HIV was isolated from 49 of 156 immunoblot[ELISA]-positive [blood] donors (31%)…Virus was detected in cultures from 56 of 131 seropositive donors (43%) at the CDC”

“The main difference in the [retroviral and retrotransposon life] cycles is the presence of infectious, extracellular, membrane-enveloped particles in the retroviruses [i.e. the presence of reverse transcription cannot be taken to indicate the presence of any retrovirus, let alone a specific retrovirus]”

“225 cultures of peripheral-blood lymphocytes from 133 seronegative men were performed, and HIV-1 was isolated [actually, detected indirectly using non-specific markers] in cultures from 31 men (23%). Of these men, four have seroconverted after being seronegative for 11 to 17 months after the initial isolation of the virus. The virus was not always isolated at every visit after the first successful isolation”

“there is no strict correlation between HIV isolation [actually merely co-culturing] from CSF [cerebro-spinal fluid] and the stage of HIV infection or the presence of neurologic complications [but, if HIV is the cause of these complications, it should be found roughly in proportion to the severity of the symptoms!]...additional mechanisms [unspecified] may account for the overt appearance of neurologic symptoms”

“virus culture is time-consuming, expensive and does not directly measure virus titre”

“Testing for HIV by PCR or culture also may be helpful in determining HIV status; however, neither test is licensed for diagnosis of HIV infection”

“The study population consisted of...patients with AIDS and patients with AIDS-related complex...The criteria for eligibility also included...an absolute number of [CD4 cells] less than 500 per cubic millimeter..., serum positive for antibody to HIV...HIV was isolated [by culture] at entry in 57% of the AZT group and 58% of the placebo group [i.e. almost half of patients with AIDS and pre-AIDS were negative by HIV by culturing techniques]”

“The protein fraction [of HIV] is obtained by disruption of sucrose gradient purified HTLV-III [HIV] [without any proof that the material obtained represents a retrovirus, let alone HIV]”

“Continuous production of HTLV-III [HIV] is obtained after repeated exposure of parental HT cells to concentrated [not purified] culture fluids containing HTLV-III harvested from short term cultured T-cells (grown with TCGF [T-Cell Growth Factor]) which originated from patients with pre-AIDS or AIDS. The concentrated fluids were first shown to contain particle-associated reverse transcriptase (RT) [i.e. no direct evidence of the presence of a retrovirus]...Samples exhibiting more than one of the following were considered positive [for an HTLV-family virus]: repeated detection of a Mg++ dependent reverse transcriptase activity in supernatant fluids [but reverse transcription occurs in cells even without retroviruses present]; virus observed by electron microscopy [but merely looking at particles cannot prove that they are retrovirus particles]; intracellular expression of virus-related antigens detected with antibodies from sero-positive donors or with hyperimmune serum [but, without purification of HIV, it is not possible to say with certainty what virus-related antigens are, and even if it was, antibody cross-reactions are still possible]; or transmission of particles, detected by reverse transcriptase assays or by electron microscopic observation, to fresh human cord blood, bone marrow, or peripheral blood T-lymphocytes [but the ability to stimulate a cellculture to produce particles is not proof that an infectious agent is present, and certainly not that it is a specific retrovirus]. All isolates not classified as either HTLV-I or HTLV-II by immunological or nucleic acid analysis were classified as HTLV-III [HIV]”

“Samples exhibiting more than one of the following were considered positive: repeated detection of a Mg++ dependent reverse transcriptase activity in supernatant fluids; virus observed by electron microscopy; intracellular expression of virus-related antigens detected with antibodies from sero-positive donors or with hyperimmune serum; or transmission of particles, detected by reverse transcriptase assays or by electron microscopic observation, to fresh human cord blood, bone marrow or peripheral blood T-lymphocytes. All isolates not classified as either HTLV-I or HTLV-II by immunological or nucleic acid analysis were classified as HTLV-III [HIV]. The cells in the HTLV-III producing cell cultures, characterized using established immunological procedures, are predominantly T-lymphocytes [the cells that HIV is supposed to kill in vivo]”

“In an early set of experiments, HTLV-III [HIV] was cultured from 48 subjects, including 18 or 21 patients with ARC [AIDS-related Complex, also known as pre-AIDS], 3 of 4 clinically normal mothers of children with AIDS, and [only] 26 of 72 adults and children with AIDS...Of interest is the finding, that while antibody to HTLV-III was more often associated with advanced disease, HTLV-III itself was more frequently cultured in ARC or newly diagnosed AIDS patients”

“The AIDS-associated retrovirus (ARV) was isolated [cultured] from vaginal and/or cervical secretions from 4 out of 8 women whose sera contained antibodies to the virus…Culture supernatants were assayed every 3 days, after day 8, for particle-associated reverse transcriptase (RT) activity. ARV [HIV] was further isolated [sic] by transfer of RT-positive fluids to normal PHA-stimulated PMC obtained from healthy volunteers…All but 1 of the 8 women also had replicating ARV in their cultured PMC [i.e. peripheral blood mononuclear cells that resulted in the detection of RT when cultured]. In general, the ARV was detected at low levels in these cells but on passage [transfer of fluids to another culture] replicated to high titre…In this small sample, the recovery of virus from the vaginal and endocervical canals did not seem to correlate with the degree of virus production by the cultured PMC.”

“14 women seropositive for HTLV-III/LAV [HIV using ELISA antibody test] gave informed consent to culture of genital secretions and venous blood…from September, 1985, to December, 1985…[HIV] was cultured from cervical secretions in 4 of 14 women…[and] was detectable in blood in 7 of 13 women tested.”

“The production of HTLV-III [HIV] was monitored as release of extracellular reverse-transcriptase activity [not unique to retroviruses, let alone any specific retrovirus], and by electron microscopic examination [which can only show that particles are compatible in size and shape with retroviruses, not that the particles are retroviruses], measurement of intracellular expression of viral proteins [probably p24, found in about half the people who are HIV antibody positive], and transmission to fresh human T lymphocytes or established T-cell lines [wherein the same non-specific phenomena will be used to declare the presence of HIV]”

“the AIDS virus has been isolated [actually, cultured, which is not equivalent to isolation] from the blood of 85% or more of seropositive high-risk individuals with AIDS or blood donors implicated in the transmission of AIDS through transfusion [but, what about the other 15%?]”

“In the last few years, the view that reverse transcription is solely a retroviral mechanism has been disproven [even though this assumption is the basis of much culture evidence]”

“Evidence for the presence of HTLV-III [HIV] included: (i) viral reverse transcriptase (RT) activity in supernatant fluids; (ii) transmission of virus by coculturing T cells…; (iii) observation of virus by electron microscopy; and (iv) the expression of viral antigens…using serum from a patient positive for antibodies to HTLV-III…or antisera prepared against purified, whole disrupted HTLV-III…[using this methodology] we found HTLV-III in…13 of 43 of adult AIDS patients with Kaposi’s sarcoma, and 10 of 21 adult AIDS patients with opportunistic infections”

“In answer to your letter of March 9, I would like to address some of the points you made. First, there is no evidence that the situation with HTLV is similar to EBV. On the contrary, the epidemiologic evidence shows a close association between disease and HTLV infection. EBV is ubiquitous. Second, I don’t understand why there is a problem with one virus causing “clonal inducer T-cell malignancies” and immunosuppressive disorders. In the cat system It’s been accepted for years (at least 10) that FeLV more often induces an immunosuppressive state than leukemia. The age of initial infection, route of exposure and whether there is repeat exposure are all apparent factors in the disease outcome of FeLV infection. If the T4 cells are the target of HTLV and this infection abrogates their function (as shown by M. Popovic, B. Dupont, A. Fauci and myself), then I can easily see that infection could lead to immunosuppression. Third, I’m not surprised that you have not found p19 expression on fresh cells of “AIDS” patients. It’s extremely rare to find fresh cells expressing the virus. As in the bovine system, cell culture seems to be necessary to induce virus. This is probably due to removal of inhibiting factors present in the patient. The antigens p24 and p19 are almost always detected simultaneously. Finally, we know now there are many variants of HTLV-I. We believe the cause of AIDS is a more highly cytopathic variant.”

“a growing constellation of eukaryotic genetic elements – various pseudogenes and repetitive sequences – appear to depend upon reverse transcriptases of unknown provenance for their existence or amplification [yet, in an HIV-culture, the presence of reverse transcription is considered unambiguous evidence for the existence of HIV]”