“Of 1551 HIV-1-infected individuals screened for helminth [parasitic worm] infection, 299 were helminth infected. 234 adults were enrolled and underwent randomization and 208 individuals were included in intent-to-treat analyses. Mean CD4 cell count was 557 cells/microl and mean plasma viral load was 4.75 log10 copies/ml at enrollment. Albendazole therapy resulted in significantly higher CD4 cell counts among individuals with Ascaris lumbricoides infection after 12 weeks of follow-up and a trend for 0.54 log10 lower HIV-1 RNA levels. These effects were not seen with treatment of other species of soil-transmitted helminths. [implying that at least one species of helminth reduces CD4 counts, and that part of what is known as ‘viral load’ may be due to these parasites]”

“Data from 1686 women were analyzed: 1203 (71.4%) were categorized as non-users, 429 (25.4%) as intermittent users, and 54 (3.2%) as persistent users of crack [cocaine]…Throughout most of the study period, those reporting persistent crack use had higher viral load and poorer immune function, whereas those reporting no use had the lowest HIV-1 RNA levels and best immune health, with intermittent crack users falling in between…We found that the median reduction in HIV-1 RNA level was highest in non-users, at 1.7 log10 copies/µl, compared with 1.4 log10 copies/µl in inactive crack users and 1.0 log10 copies/µl in active users. The median CD4 cell count increase was highest in non-users, at 161 cells/µl, compared with 123 cells/µl in inactive users, and 100 cells/µl in active users.”

“More research is needed to determine the degree to which the viral load in blood predicts the risk of HIV transmission and to determine the association between the viral load in blood and the viral load in semen and vaginal secretions”

“HIV-1 RNA was not prognostic for mortality”

“It is advised by the manufacturing companies that viral load tests should not be used as a diagnostic assay, but my belief is that the reason they have done that is there haven’t been studies done to validate them as diagnostic tests. In practice, however, many people [doctors] do use a viral load test as a test to help confirm the diagnosis of HIV infection. it is just that manufacturers don’t recommend it.”

“[Lawyer] Turner says 'HIV experts acknowledge that there are problems measuring the actual viral load. Different laboratories and different PCR tests obtain markedly different results for the same viral load of the identical specimens'. Do you agree with that?

[Gallo]Yes, I can't testify to the court that it is markedly. There are also variations depending on how they purchase the assay and exactly what you look for and what your primers for the PCR are. We are getting technical but you can look for this part of the genome or you can look for that part of the genome and sometimes these don't exactly correlate one to one. That is true…

[Lawyer] Do you agree that the nucleic acid tests, that is the PCR tests, cannot be used to prove HIV infection?

[Gallo] I mean of course you can use it as a component of evidence. You couldn't use it to prove necessarily a virus.”

“[This article is not about HIV, but it illustrates the dangers of relying on PCR-based tests for any disease] Dr. Brooke Herndon, an internist at Dartmouth-Hitchcock Medical Center, could not stop coughing. For two weeks starting in mid-April last year, she coughed, seemingly nonstop, followed by another week when she coughed sporadically, annoying, she said, everyone who worked with her. Before long, Dr. Kathryn Kirkland, an infectious disease specialist at Dartmouth, had a chilling thought: Could she be seeing the start of a whooping cough epidemic? By late April, other health care workers at the hospital were coughing, and severe, intractable coughing is a whooping cough hallmark…It was the start of a bizarre episode at the medical center: the story of the epidemic that wasn’t. For months, nearly everyone involved thought the medical center had had a huge whooping cough outbreak, with extensive ramifications. Nearly 1,000 health care workers at the hospital in Lebanon, N.H., were given a preliminary test and furloughed from work until their results were in; 142 people, including Dr. Herndon, were told they appeared to have the disease; and thousands were given antibiotics and a vaccine for protection. Hospital beds were taken out of commission, including some in intensive care. Then, about eight months later, health care workers were dumbfounded to receive an e-mail message from the hospital administration informing them that the whole thing was a false alarm. Not a single case of whooping cough was confirmed with the definitive test, growing the bacterium, Bordetella pertussis, in the laboratory. Instead, it appears the health care workers probably were afflicted with ordinary respiratory diseases like the common cold. Now, as they look back on the episode, epidemiologists and infectious disease specialists say the problem was that they placed too much faith in a quick and highly sensitive molecular test that led them astray…At Dartmouth the decision was to use a test, P.C.R., for polymerase chain reaction…At Dartmouth, when the first suspect pertussis cases emerged and the P.C.R. test showed pertussis, doctors believed it. The results seem completely consistent with the patients’ symptoms…Then the doctors decided to test people who did not have severe coughing…“That’s how we ended up with 134 suspect cases,” Dr. Kirkland said. And that, she added, was why 1,445 health care workers ended up taking antibiotics and 4,524 health care workers at the hospital, or 72 percent of all the health care workers there, were immunized against whooping cough in a matter of days. “If we had stopped there, I think we all would have agreed that we had had an outbreak of pertussis and that we had controlled it,” Dr. Kirkland said…The Dartmouth doctors sent samples from 27 patients they thought had pertussis to the state health departments and the Centers for Disease Control…There was no pertussis in any of the samples…when the Centers for Disease Control tested those 39 samples [for antibodies], its scientists reported that only one showed increases in antibody levels indicative of pertussis…Dr. Cathy A. Petti, an infectious disease specialist at the University of Utah, said the story had one clear lesson. “The big message is that every lab is vulnerable to having false positives,” Dr. Petti said. “No single test result is absolute and that is even more important with a test result based on P.C.R.””

“We studied the response to nevirapine-based antiretroviral treatment among women and infants who had previously been randomly assigned to a single, peripartum dose of nevirapine or placebo in a trial in Botswana…All women were treated with antenatal zidovudine [AZT]. The primary end point for mothers and infants was virologic failure by the 6-month visit after initiation of antiretroviral treatment…By the 6-month visit after the initiation of antiretroviral treatment, 5.0% of the women who had received placebo had virologic failure, as compared with 18.4% of those who had received a single dose of nevirapine…Among 60 women starting antiretroviral treatment within 6 months after receiving placebo or a single dose of nevirapine, no women in the placebo group and 41.7% in the nevirapine group had virologic failure. In contrast, virologic failure rates did not differ [statistically] significantly between the placebo group and the nevirapine group among 158 women starting antiretroviral treatment 6 months or more post partum (7.8% and 12.0%, respectively). Thirty infants also began antiretroviral treatment (15 in the placebo group and 15 in the nevirapine group). Virologic failure by the 6-month visit occurred in significantly more infants who had received a single dose of nevirapine than in infants who had received placebo.”

“A Reactive result indicates that HIV-1 RNA was detected. For a specimen that is repeatedly reactive on an HIV-1 antibody test and reactive in the APTIMA HIV-1 RNA Qualitative Assay, the individual is considered confirmed infected with HIV-1…A specimen that is reactive in the APTIMA HIV-1 RNA Qualitative Assay but that has not been tested in an HIV-1 antibody test should be further tested using a licensed test for HIV-1 antibodies.”

“Presenting HIV RNA level predicts the rate of CD4 cell decline only minimally in untreated persons. Other factors, as yet undefined, likely drive CD4 cell losses in HIV infection. [Theoretically, HIV RNA represents the amount of virus, and CD4 cells are the type that HIV likes to kill. So there should be a correlation]”

“Clinicians treating patients with HIV encounter some patients with low plasma viral levels who experience rapid progression. What mechanism is responsible for their profound and quick CD4 cell loss? On the other end of the spectrum are those patients with high level HIV viremia who respond clinically like sooty mangabeys infected with simian immunodeficiency virus (SIV), which can tolerate high levels of SIV replication without disease progression. Are such patients statistical extremes in an otherwise simple and uncontested paradigm, or are clinicians and researchers missing something? [like, um, missing something really, really big?]…Rodríguez et al have taken the perspective of the individual patient in attempting to quantify how much of the variability of the individual CD4 cell loss is explained by the baseline plasma HIV RNA viral load…The provocative main finding from their study was that the presenting plasma HIV RNA load predicted no more than 10% of the observed CD4 cell loss in patients with chronic untreated HIV infection.”

“A malnourished person usually has a high viral load because they have few resources in their body to combat the virus. People with higher viral loads are much more infectious, thus making their partners far more susceptible to getting HIV. Malaria, bilharzia and intestinal worms also drive up a person’s viral load.”

“Paradoxically, baseline serum cholesterol, but not atorvastatin, influenced viral rebound at week 4.”

“A total of 2,937 sera samples, from [Canadian] individuals who were newly diagnosed between 1984 to March 31, 2004 [were examined]…Viral RNA was successfully amplified from 2,152 (73.2%) of the sera samples [More than 25% antibody-positive people were negative for viral RNA using the incredibly sensitive PCR test!]”

“HCV [Hepatitis C Virus] has been detected in breast milk and colostrum, and in one study the transmission rate was higher in babies exposed to HCV RNA positive breast milk. However, in most studies no association has been observed between transmission and mode of infant feeding. Of the seven breastfed children in our study with late first positive PCR tests, two were last PCR negative after 4 weeks of age, suggesting possible transmission through breast feeding. However, although we cannot exclude the possibility of postnatal transmission, the occurrence of late last negative [i.e. negative well after birth] and late first positive PCR tests in five of the non-breastfed children as well as in these two breastfed children suggests that these viraemia patterns may not necessarily indicate timing of transmission. We would echo the need to be cautious in inferring the timing of infection simply from early PCR results.”

“Quantitative PCR should not be used as a diagnostic test for HIV because false positives and false negatives can occur in these circumstances”

“several independent contemporary studies have shown that the levels of CSF [cerebral spinal fluid] HIV RNA appear to be significantly lower in untreated subjects with HIV-D [HIV-associated Dementia] than those examined in the era before HAART [indicating that levels of HIV RNA in the CSF is an unreliable surrogate marker]”

“Persistent virological response, defined as achieving and maintaining plasma HIV-1 RNA levels at or below the limit of assay quantification (calculated through weeks, 24, 48, and 60), was used as the primary efficacy end point”

“Test Name: HIV-1 RNA U.Quant/Plasma (>100000 copy/ml). THIS TEST IS NOT FDA APPROVED. FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC PROCEDURES.”

“the cumulative proportion of persistent undetectable HIV viral load below 500 copies/ml was significantly higher in the antiretroviral naive patients [those who had never taken AIDS drugs before] than in the non-naive ones…In multivariate analysis, being naive of antiretroviral treatment and having a low viral load, at the time of HAART introduction, were significantly correlated with a sustained undetectable HIV viral load.”

“Survival related significantly to height growth velocity but not to weight growth velocity or head circumference growth velocity. Viral load was not significantly associated with changes in weight-, height-, or head circumference-for-age z scores in children <30 months of age or with changes in weight- or height-for-age z scores in older children.”

“The median viral load…at the time of diagnosis [of PHI-Primary HIV Infection, a flu-like illness sometimes associated with initial seroconversion] was greater than 750,000 copies/mL (range, 8493-90,397,351 copies/mL [A range of more than 10,000 times!])”

“20 HIV-infected subjects were studied…A detectable viral load was present in 15 of 17 plasma samples, 10 of 19 BALF [broncho-alveolar lavage fluid] samples, 3 of 20 BF [bronchial fluid] samples, and 1 of 20 saliva samples…42% of subjects who were smokers had detectable viral loads found in BALF samples, [compared to] 71% of nonsmokers…Thus, the immunosuppressive properties of smoking may actually inhibit viral replication [more importantly, this study illustrates that the difference between a positive and negative viral load could depend upon the type of sample used to measure it]”

“The VERSANT HIV-1 RNA 3.0 Assay (bDNA) is not intended for use as a screening assay for HIV infection or as a diagnostic test to confirm the diagnosis of HIV infection.”

“From April 1996 through December 2000, a total of 501 antiretroviral-naive [never taken AIDS drugs] HIV-seropositive patients who initiated HAART were recruited…at the Hospital Ramón y Cajal [Madrid, Spain]…After 24 months of follow-up, 42 (16.5%) of 255 patients were considered to have a discordant immune response [low CD4 cell counts with low viral load or high CD4 cell counts with high viral load]…Clinical progression of HIV disease was uncommon among the patients included in the analysis. Overall, 4 patients (1.6%) died of HIV infection-related complications, and 44 patients (17.3%) developed HIV infection-related clinical events…Most events (29 [65%] of 44 events) occurred within the first year after initiation of HAART. Overall, clinical events were not more frequent among patients with a discordant immune response than among patients with a good immunologic response.”

“We report in this work that HIV-1 and HIV-2 patients having a similar degree of CD4 depletion displayed similar levels of T cell hyperactivation and similar numbers of cycling cells in the peripheral blood despite great differences in the plasma viral load. These results and other recent reports call for re-evaluation of different hypotheses about causal relationships among virus concentration, CD4 depletion, and activation and turnover of T lymphocytes.”

“56 clinically asymptomatic HIV-1-infected individuals [from Ethiopia], 31 (55%) of whom were also infected with helminths [intestinal worms], were studied…At baseline, HIV plasma VL [viral load] was strongly correlated to the number of eggs excreted and was higher in individuals infected with more than one helminth. After treatment of helminths, the 6-month change in HIV plasma VL was significantly different between the successfully treated group and the persistently helminth-positive group”

“The 2 articles recently published in JAMA are probably among the most important HIV-related papers of the year and have tremendous implications for decision-making. They question the rather arbitrary viral load threshold used in current antiretroviral guidelines and suggest that the main factor that should influence when to start therapy -- and maybe the only one to consider -- should be the CD4+ cell count. The only contributions that viral load can make to this decision are (a) to determine how quickly a given individual is likely to reach the CD4+ cell count threshold at which therapy is indicated, and (b) perhaps to determine the frequency of CD4+ cell-count monitoring. Monitoring viral load only becomes critical when therapy is started.”

“The three tests for HIV-1 RNA were 100% sensitive [in this sample of 40] for preseroconversion PHI [i.e. no people diagnosed with Primary HIV Infection symptoms by other methods did not have detectable RNA]. However, bDNA [branched DNA] testing for PHI had a 5% false-positive rate, PCR testing had a 3% false-positive rate and the transcription-mediated amplification assay had a 2% false-positive rate. The false-positive results on the bDNA test ranged from 584 to 2058 copies/ml. False-positive PCR tests ranged from 58 to 103 copies/ml.”

“Prior to the advent of polymerase chain reaction (PCR) technology, physicians relied on HIV culture and p24 antigen testing. Antigen testing is neither sensitive nor specific enough for diagnosis. HIV culture, although accurate, is time-consuming and expensive.Therefore, physicians have quickly embraced PCR technology for testing HIV-exposed infants…DNA is preferred [over RNA] for diagnostic purposes because of its sensitivity even in the face of HAART.”

“Several potential mechanisms could explain the transient suppression of HIV replication early during the course of measles. Measles virus infection results in lymphopenia, with reductions in CD4+ T lymphocyte percentage and number. The nadir [low point] of the lymphopenia precedes the onset of rash, and values usually return to normal by 1 month after the onset of the rash. This early reduction in CD4+ T lymphocytes could decrease the number of target cells for HIV replication. An alternative explanation is that measles virus infection stimulates the production of soluble factor(s) capable of suppressing HIV replication.”

“The effect of highly active antiretroviral therapy (HAART) in 85 children infected with human immunodeficiency virus type 1 (HIV-1) was compared retrospectively among Centers for Disease Control and Prevention (CDC) immunologic groups 1-3...HAART resulted in the suppression of HIV-1 below detectable levels in 34.8%, 25%, and 32% of patients in the 3 CDC groups, respectively, and in a frequent switch from syncytium-inducing to nonsyncytium-inducing virus. The results support...the use of other markers of disease progression, in addition to virus load.”

“The Spearman's rho correlation between viral load in semen and plasma was not significant…When restricted to participants with detectable virus in semen and plasma, the correlation remained nonsignificant…There was also poor concordance between undetectable viral loads in plasma and semen; 53% of men with undetectable virus in plasma had detectable viral loads in semen and 31% of men with undetectable virus in semen had detectable plasma viral loads…Men with equal or greater virus in their plasma than their semen compared with men with greater virus in the semen than plasma showed no differences in age, ethnicity, sexual orientation, years since testing HIV positive, HIV symptoms experienced, CD4+ cell counts, current HIV treatment status, or current HIV treatment adherence…[The only correlation found was] that men with greater concentration of HIV in their semen relative to concentrations of HIV in plasma reported significantly greater rates of unprotected vaginal intercourse and total number of unprotected sexual intercourse occasions as the insertive partner…Results failed to find differences in STI [sexually transmitted infections] occurrences between men with relatively higher and lower semen viral loads.”

“[Table 2 shows that the results for identical material sent to multiple laboratories provided viral load results varying from 3,849 to 1,291,635 (Roche Amplicor HIV-1 Monitor), from 63,750 to 205,500 (Bayer HIV-1 3.0 RNA) and from 89,000 to 360,000 (Organon Teknika NucliSens)]”

“The NucliSens HIV-1 QT assay is not intended to be used as a screening test for HIV-1 nor is it to be used as a diagnostic test to confirm the presence of HIV-1 infection.”

“Autopsy samples from 13 HIV-infected subjects were examined for HIV-1 viral RNA (vRNA), and viral reverse transcriptase (RT) genotype was determined...The median HIV-1 vRNA level in brain samples from [2] subjects with moderate dementia was 7.79 log(10) copies/g...compared with 5.44 log(10) copies/g for [3] mildly demented subjects and 4.87 log(10) copies/g for those obtained from [7] nondemented individuals...[but] some demented subjects had relatively low levels of HIV-1 vRNA, and paradoxically some nondemented subjects had high vRNA brain levels”

“HIV-1 RNA was…weakly detected in 3 uninfected infants who were negative by DNA PCR at the same age and who ultimately appeared to be RNA undetectable and HIV-1 antibody seronegative [this should be impossible as, if there is no HIV integrated in cells (as DNA) there can't be any virus particles (containing RNA)]”

“That a clinical benefit may not have been achieved with multi-drug rescue therapy calls into question the current wisdom of deeming an undetectable viral load the goal of therapy in the heavily pre-treated population. Even if it can be accepted that an undetectable viral load is an appropriate surrogate marker for clinically relevant outcomes in treatment-inexperienced patients who are initiating combination therapy, it cannot necessarily be accepted without proof that it is a useful surrogate in heavily pre-treated patients...Therefore, until controlled trials are able to prove the utility of an undetectable viral load as a surrogate marker for clinically relevant outcomes in heavily pre-treated patients, we believe that clinicians should show caution before striving for complete viral suppression at any cost”

“The difference in HIV RNA between children on ZDV [AZT] and placebo was relatively small. After two years of ZDV, the proportion of IMM children [assigned to AZT at the beginning of the trial, rather than deferred until AIDS developed] with high level (>50%) resistance to ZDV was quite low (38%), and 13% had no ZDV associated mutations [meaning that viral resistance to therapy is not an explanation for anti-HIV therapy to reduce the amount of HIV in the body]”

“When it came time to write up the data [on the AIDS 'therapeutic vaccine' called Remune] for publication, Kahn, Lagakos, and others on the team concurred that the analyses showed no benefit from the drug. But scientists from Immune Response performed their own analysis of blood tests on a sample of 250 patients in whom, the company argued, some benefit could be seen - not in longer survival, but in having lower levels of virus ['viral load'] in their blood…[Lead researcher] Lagakos said: ''The company did not want our original analysis to go forward. We were put in a position where we had to agree to terms that were unacceptable to us. We decided to go forward with what we had.''”

“Because of the RNA assay's 1.9% to 3.0% false-positive rate, results must be carefully interpreted and compared to HIV-1 viral load levels seen during proven HIV-1 seroconversion. We report the case of a sexually active woman with symptoms suggestive of ARS [Acute Retroviral Syndrome] who had a false-positive HIV-1 RNA assay result”

“Because of the RNA assay's 1.9% to 3.0% false-positive rate, results must be carefully interpreted and compared to HIV-1 viral load levels seen during proven HIV-1 seroconversion. We report the case of a sexually active woman with symptoms suggestive of ARS who had a false-positive HIV-1 RNA assay result.”

“After adjustment, complete nonresponders [CD4 cell counts did not rise, and viral load did not fall after treatment with HAART] and those with only a virologic response had significant higher risks for clinical progression at 6 months [i.e. a reduction in viral load did not reduce the risk for illness]”

“In a small proportion of non-[HIV Type]B samples (about 14%), the HIV RNA copy number, as determined by NASBA, was substantially lower or undetectable compared to results obtained with the other two assays. This phenomenon has also been observed during paediatric diagnosis of HIV…In 8 infected African infants and their mothers, although proviral DNA was detected by the Roche Amplicor assay, viral RNA was either not detected or was detected only at a low copy number by NASBA. When infant specimens were tested by the RT-PCR assay, however, HIV RNA was detected and at a higher concentration…It has been demonstrated that in some cases although HIV viral load may be undetectable by NASBA, the RT-PCR assay subsequently reports a substantial HIV RNA copy number.”

“there was no association [of adherence] with viral load, although only 13 person (31%) met all four adherence goals.”

“Because viral RNA levels [viral load] are quantified as copies of RNA per milliliter, it is possible that nonviable, but persisting, HIV-1 residues are being detected by such testing. This raises an even more fundamental question as to the validity of viral RNA monitoring in general.”

“Maternal plasma HIV-1 RNA level [‘viral load’]…was not associated with perinatal transmission [in this group of women with moderately advanced HIV-1 infection and ZDV/AZT drug resistance]”

“Negative results (<50 copies/mL), were obtained in 30/32 (94%) [bDNA 3.0] assays [meaning that false-positive results were obtained in 2/32 or 6% of cases]”

“In contrast to previous reports...the viral load in the majority of the [long-term survivors] tested was detectable and, in some [long-term survivors], quite high...and variable over time.”

“The AMPLICOR HIV-1 MONITOR Test is an in vitro nucleic acid amplification test for the quantitation of Human Immunodeficiency Virus Type 1 (HIV-1) RNA in human plasma...[It] is not intended to be used as a screening test for HIV or as a diagnostic test to confirm the presence of HIV infection…Quantitative culture has limited utility for monitoring virus levels in infected individuals since only a small fraction of virus particles is infectious in vitro. Infectious virus is often undetectable in asymptomatic individuals…The clinical specificity of the…test was determined by analysis of 495 anti-HIV-1 negative blood donors. None of these specimens was reactive…Assuming[!] a zero prevalence of HIV-1 infection in the seronegative blood donors, the specificity of the test was 100%”

“Participants with baseline RNA levels below the level defining virological response were excluded from the analysis [which creates an artificial downward trend in the viral load, by eliminating people whose viral load might well go up on therapy]...In the ZDV-naive population [had not taken AZT before this trial] the maximum median fall in HIV RNA occurred by 4 weeks for all three [treatment] arms [AZT, AZT+ddI, AZT+ddC]. Thereafter, the median HIV RNA increased towards baseline levels [so you take these drugs for a lifetime for a theoretical benefit that lasts a month!]... The proportions responding (i.e. achieving HIV RNA [viral load] less than 800 copies/ml) in the first [year] in Delta 1 were 61% on ZDV-ddI and 45% on ZDV-ddC [and 10% on ZDV alone]. In Delta 2 [had previously taken AZT], the proportion of responders [whose viral load dropped] was lower; 23% on ZDV-ddI, and 30% on ZDV-ddC [and 2% on ZDV alone]”

“The results obtained for patients with a broad range of plasma viral loads before and after antiretroviral therapy reveal a constant mean viral (v)RNA copy number (3.6 log10 copies) per infected cell, regardless of plasma virus load or treatment status.”

“Compared with patients who maintained undetectable viral load, the adjusted relative hazard of AIDS or death was 1·00 [i.e. no different] for patients with viral rebound [had a viral load that dropped, but later became higher], and 2·40 for patients who failed to reach undetectable concentrations.”

“18 subjects had 1 or 2 positive results with v.2.0 and an undetectable confirmatory test for a false positive rate of 4.4%. The rate is similar at baseline (9/183 subjects = 4.9%), wk. 4 (7/162= 4.3%) and wk. 26 (2/44 = 4.5%). Of the 18 pos. specimens, 9 tested pos. once and 9 twice. With version 3.0, 11 of 67 samples tested were pos. (16.4%). 6 were pos. once and 5 twice. The range of false pos. rates was 9.1% at wk. 4 (total of 22 specimens) to 26.7% at wk. 26 (total of 15 specimens). A week 4 sample with two values of 8,000 copies/ml on v.2.0 was neg. by DNA PCR, p24 antigen and Western Blot. Follow-up testing of this subject at wk. 26 was negative for HIV antibody and RNA. The emotional impact of a false positive screening RNA test in a recently exposed person is significant. With the high false positive rate, we do not advocate the routine use of HIV RNA tests to screen asymptomatic people. The high rate of repeat false positive tests in a given sample (50%) suggests a possible biologic mechanism”

“Plasma viral load tests for HIV-1 were neither developed nor evaluated for the diagnosis of HIV-1 infection; therefore their diagnostic specificity is not well delineated when applied to persons who are negative for HIV antibody. We report two cases of false-positive results obtained by using branched-chain DNA assay…and one case…by using HIV reverse transcriptase polymerase chain reaction (RT-PCR)…These three cases illustrate the potential problems of using HIV-1 plasma viral load tests for diagnosis of HIV infection…Only patients who have a high pre-test probability of a positive result should be evaluated for primary infection by using plasma viral load testing [i.e. preconceptions about risk groups such as gay men makes a positive test more likely]…Their performance in patients who are not infected with HIV is unknown”

“We selected 20 healthy volunteers, all of whom yield negative results for HIV antibodies using different screening tests. Plasma from all of them were analysed by three different currently available HIV viral load tests: branched DNA (bDNA) signal amplification assay (Chiron), nucleic acid sequence-based amplification (NASBA) Nuclisens (Organon Teknika), and Ultradirect reverse transcriptase (RT)-PCR Monitor (Roche)…2 samples [10%] yielded positive results by the bDNA assay…Another 2 specimens [10%] yielded false-positive results by the NASBA Nuclisens…[and] one of the 20 samples [5%] was interpreted as positive by the Ultradirect RT-PCR Monitor assay…using the Monitor test with non-B primers, up to 4 of the 20 samples [20%] yielded positive values…Results were reproduced in more than half of tested specimens for which plasma volumes were enough for repeat testing”

“A reanalysis is presented of 43 cases of apparent transient viremia [one or more positive culture or PCR assays for HIV-1 and the subsequent inability to detect HIV-1 in the specimens on multiple occasions or seroreversion, or both]. 41 cases occurred among 1561 infants in five studies of mother-to-infant HIV-1 transmission...Our negative studies of 43 cases of suspected transient infection indicate that the phenomenon...remains to be proven and that most cases suggestive of transient HIV-1 infection are cases of mislabeling of specimens or their contamination in the laboratory...”

“Specimens were studied from one mother and her child, both with suspected transient viremia. The mother had two and the infant three positive HIV-1 cultures, but subsequently both individuals became negative for HIV-1 by nPCR, standard virus cultures, CD8+-depleted virus cultures, and enzyme-linked immunosorbent assay. HIV-1 RNA and DNA were not detected in two lymph nodes taken from the mother 3 and 4 years after the last virus-positive culture. PCR amplification and DNA sequencing of HIV-1 env sequences from the mother’s two and the child’s three culture supernatants were performed in separate laboratories to eliminate the possibility of cross-contamination. Phylogenetic analysis found that none of the five isolates were genetically linked. Although it is improbable, these five virus isolates appear to have arisen from five separate incidents of specimen contamination or mislabeling. This case remains enigmatic, however, in that both the mother and infant had strong CD8+ cytotoxic T lymphocyte (CTL) responses and lymphocyte proliferation to multiple HIV-1 antigens”

“[This study was a] Prospective follow-up of a cohort of 162 unselected, protease inhibitor-naive, antiretroviral-experienced patients with advanced HIV disease, treated with indinavir combined with two nucleoside analogues…21% of patients exhibited discrepant virological and immunological responses to treatment, of whom one-half failed to exhibit significant increases in CD4 cells despite a virological response to therapy and one-half exhibited increased CD4 cell counts in the absence of significant decrease in plasma viral load. The incidence of AIDS-defining events in the latter group of patients was similar to that of responder patients, whereas their incidence was higher in patients who failed to exhibit a virological and immunological response and those who failed to increase CD4 cells despite a significant decrease in viral load. [calling into question the common use of viral load to estimate the rate of progression to AIDS]”

“Peak levels [of HIV-1 RNA] in the first 120 days were not predictive of disease progression”

“the use of absolute viral levels is problematic for individual patient management because the assessment of the absolute viral RNA level is dependent upon the test method.”

“Unspecific pick-ups during PCR can be due to annealing of primers to target sequences with a low degree of homology”

“Initial testing of plasma by RNA RT-PCR was reported as positive. This result was not confirmed by viral cultures, nested DNA PCR, western blot, or EIA. Additional RNA RT-PCR assays gave positive results from earlier occasions in the vaccine trial. Eventually, testing of all previously reactive samples by RNA RT-PCR was repeated in a quality-controlled laboratory, and confirmed the negative HIV-1 status of the individual. Interpretation This case report exemplifies the difficulties with use of viral-genome measurement as a screening test to diagnose HIV- 1 infection, particularly in individuals who have ever participated in HIV-1 vaccine trials. Monitoring of large numbers of phase III vaccinees by RNA RT-PCR will not be feasible. The design of efficacy trials for new vaccines should be in parallel with development of antibody-based diagnostic tests that are capable of differentiating between immunisation and true HIV-1 infection.”

“[In this study of 78 HIV-positive people with high CD4 cell counts and no symptoms] there were no differences in viral load with regard to time of HIV-1 infection [i.e. amount of virus does not grow over time]…10 patients fulfilled the criteria for LTNP [Long Term Non-Progressors]. 7 of these 10 patients had viral loads above 10,000 RNA copies/ml and 2 above 30,000 RNA copies/ml. The level of viral load of LTNP was not statistically different compared with the other 68 patients”

“We observed that clade A strains [of HIV] were not detected by RT-PCR [Reverse Transcriptase-Polymerase Chain Reaction] and that clade G-strains were not detected by NASBA [Nucleic Acid Sequence-Based Amplification]. However, the copy number detected by RT-PCR in one clade E (CM235) and in one clade F (163.3070) was much lower than the copy number detected by bDNA [branched DNA] and NASBA...for clades B and D (UG270), the HIV-1 RNA levels measured by bDNA were lower than those obtained by RT-PCR and NASBA”

“The copy number obtained with the bDNA assay tended to be lower than that obtained by the reverse-transcription-PCR assay...by a median factor of two.”

“Specimens were tested twice with PCR; only those for which PCR reaction was repeatedly positive were reported as positive [which indicates that false positives occurred at a significant level, although no numbers were reported. Furthermore, if a false positive can occur once, it can occur twice]”

“for 223 specimens for HIV-infected infants [based on two positive cultures], 89% had PCR positivity, 11% had PCR negativity and <1% had indeterminate status.”

“maintenance of plasma HIV RNA levels below 10,000/ml in early HIV disease appears to be associated with decreased risk of progression to AIDS. However, in patients with more advanced disease [low CD4 cell counts], disease progression occurred in up to 30% of patients with fewer than 10,000 HIV RNA copies/ml [In other words, viral load is not terribly well associated with progression to AIDS]...HIV RNA levels should not be measured within a month of acute illnesses or within a month after influenza and pneumococcus immunizations. Increases in HIV RNA levels in blood of as much as 300-fold have been observed within two weeks of routine immunizations against influenza, tetanus, or pneumococcus”

“Our investigation produced two main findings. First, the false-positive and false-negative rates of PCR that we determined are too high to warrant a broader role for PCR in either routine screening or in the confirmation of diagnosis of HIV infection. This conclusion is true even for the results reported from more recent, high-quality studies that used commercially available, standardized PCR assays...We did not find evidence that the performance of PCR improved over time”

“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 majority (83%) of [flu] vaccinated [HIV-positive] individuals experienced a significant increase in plasma HIV-1 RNA levels within 1-2 [weeks after] immunization and returned to their prevaccination levels within 4 weeks after immunization...patients on antiretroviral therapy were not noticeably different from those not in therapy with regard to increases in plasma viremia”

“The sensitivity of the polymerase chain reaction [PCR] technique has been estimated at 30-50% during the first week of life, 70-90% at 1-2 months of age and >95% after 3 months of age…However, the techniques used are not standardized, there is no common quality control programme, few results are available from prospective cohort studies in developing countries and the detection rates in the neonatal period are low”

“the high level of plasma virus observed by Piatak et al, was about 99.9 per cent non-culturable, suggesting that it was either neutralized or defective.”

“96% (105 of 110) of all [HIV-positive] patients tested had quantifiable plasma RNA [although that is high, one wonders how the others can have HIV in their bodies without detectable amounts of HIV RNA (free HIV particles) using PCR, an extraordinarily sensitive method]”

“Primary infection with [HIV] is generally followed by a burst of viraemia [high 'viral load'] with or without clinical symptoms. This in turn is followed by a prolonged period of clinical latency. During this period there is little, if any, detectable viraemia, the numbers of infected cells in the blood are very low, and it is extremely difficult to demonstrate virus expression in these cells…in patient 1 [high CD4 cell count], the frequency of HIV-infected cells is 1/10,000 in the PB [Peripheral Blood], and between 1/100 and 1/1,000 in the LN [Lymph Nodes]. In patient 6 [moderately low CD4 cell count], the frequency is between 1/1,000 and 1/10,000 in the PB, and 1/1000 in the LN. In patient 10 [low CD4 cell count], the frequency is between 1/10 and 1/100 in both PB and LN [but note that Piatak (1993) showed evidence that only 1/60,000 ‘infected’ cells were actually viable, infectious virions, and there is no word in the paper of which, if any, of these 12 people studied were ill]”

“Most CD4+ lymphocytes must be latently infected because we did not detect viral RNA in more than a small fraction of the cells in which HIV DNA was detectable. One in 100-400 cells with HIV DNA had viral RNA as abundant as productively infected cells; and a higher proportion of cells had low levels of viral RNA [up to 6%]”

“Circulating levels of plasma virus determined by QC-PCR also correlated with, but exceeded by an average of nearly 60,000-fold..., titers [amounts] of infectious HIV-1 determined by quantitative endpoint dilution culture of identical portions of plasma.”

“Our study of PBMC [Peripheral Blood Mononuclear Cells] from 56 HIV-1-seropositive patients, using in situ hybridization alone, also [as did other studies] revealed only 1 in 5,000 to 1 in 100,000 cells positive for HIV-1-specific nucleic acids [DNA]...Our finding with the use of in situ PCR that large numbers [if you consider 0.1% to 13.5% of cells a ‘large number’] of PBMC from HIV-1-seropositive patients contain the provirus suggests that direct cytopathic [cell-killing] effects of the virus may be an important but not necessarily the sole cause of depletion of CD4-positive lymphocytes. Our data also argue strongly against the theory that HIV-1 is not the primary etiologic [causative] agent of AIDS [huh?]...we were able to...show a relation between viral load and the stage of HIV-1 clinical infection. Patients in Stage II [HIV+, but no symptoms] had a significantly lower percentage of HIV-1-positive PBMC than those in Stage III and Stages IV-A to IV-C [but the authors neglect to mention that this is only true of the average value, there was considerable overlap between individual values. More importantly, they also do not mention that the average value at Stage III (lymph gland enlargement) was higher than at Stage IV-A to C (AIDS) and much higher than at Stage IV-D (Kaposi’s Sarcoma)]. Patients in Stage IV-D (who had Kaposi’s sarcoma only) had relatively low numbers of HIV-1-infected cells.”

“This proficiency study of PCR detection of HIV-1 DNA in serum identified a disturbingly high rate of nonspecific positivity with a widely employed gag primer pair system. In fact, the overall rate of positivity was not significantly different for serum specimens from seropositive patients and seronegative control donors (26% versus 18%)”

“PCR performed before the age of 1 month identified only 55% of the truly infected infants”

“The [HIV-1 DNA PCR, ‘viral load’] assay did not appear more sensitive than classic serologic tests in the diagnosis of HIV-1 infection [all seronegative individuals were negative on DNA PCR, including those who had had unprotected sex with seropositive people]...it was even less sensitive, because the specimen from one of our seropositive individuals was negative on PCR with the three primer pairs. Our results are consistent with...Jackson, Taylor, and Lifson...However, our results are not consistent with those of other studies: Loche, Wolinski, Imagawa, and Ameisen observed positive PCRs in seronegative at-risk individuals. These discrepancies are difficult to explain.”

“The DNA equivalent of 200 microliters of blood was assayed using the pol, gag, and nef primer pairs in a single PCR, and by performing a nested gag PCR…[on] 11 Western blot anti-p24(gag)-indeterminate blood donors. No positive signal was found for any of these donors.”

“false positive PCR results were unrelated to the child’s age [i.e. occurred at all ages up to 18 months]. PCR was recently proposed as a diagnostic method for HIV-1 infection in neonates in order to initiate early specific therapy. Our findings suggest the need for caution in using this test alone as evidence of true HIV-1 infection. In addition, positive findings with at least two primer pairs in the sample do not avoid the possibility of sample contamination by exogenous DNA and, in our opinion, positive results in at least two different subsequent samples are required.”

“Only 50% of collaborating AIDS Clinical Trial Group laboratories were able to detect 60 copies of HIV DNA in 150,000 cells in a National Institutes of Health PCR standardization programme. 33% of these laboratories had problems with false positives.”

“Among the 8 newborns…born to HIV1-infected mothers, 6 had detectable HIV1 DNA sequences…In group II [babies], 17 of 23 sera were available…HIV1 DNA was detected in PBL [peripheral blood leukocytes] of 14 of 23 babies. Among the 14 babies positive by PCR, 10 had positive or incomplete and 2 had negative Western blots. Among the nine babies negative by PCR, 3 had positive or incomplete and two had negative Western blots. Therefore, 2 of 4 babies, who were seronegative at the time of PCR test, had HIV1 DNA in PBL…8 (including the 4 with symptoms consistent with AIDS) of the 14 HIV1 DNA-positive and 1 of 9 HIV DNA-negative babies had clinical features likely to be related to HIV infection. The results obtained with the three sets of primers markedly differed; indeed 7 of 31 samples yielded positive results with all three sets of primers as compared to 7 of 31 which were positive with two sets of primers but negative with the third, and the remaining six of 31 samples were only positive with one set.”

“We found that in infected but asymptomatic patients with HIV-1, [an average of] 1 in 50,000 PBMC harbored the virus. When such a patient’s condition progressed to AIDS-related complex or AIDS, the viral titer increased significantly, to approximately 1 in 400 PBMC...It is unclear from this study, however, what percentage of the infected cells carry HIV-1 latently and what percentage of the cells express the virus actively. If 1 in 10,000 PBMC...express viral messenger RNA...then 99.6% of the infected mononuclear cells harbor the virus latently and the remaining 0.4 % express it actively [i.e. 1 in 100,000 cells have active virus!]...this information on the quantitation of HIV-1 [“high levels of HIV-1 viremia”] should reduce residual doubts about whether HIV-1 is the true etiologic agent of AIDS”

“17 of 18 DNA samples from seropositive subjects (94%) had positive [PCR] signals…13 of 21 (62%) RNA samples from seropositive subjects were positive for HIV RNA…RNA samples also gave varying intensities of PCR signal.”

“we analyzed samples from individuals known to be at especially high risk of HIV infection-seronegative sexual partners of seropositive individuals...Of 16 seronegative partners tested, 5 were unequivocally positive for HIV DNA. The clinical records of these 5 subjects confirmed that they were seronegative by enzyme-linked immunosorbent assay and western blot and negative for the p24 antigen at the time the blood samples were taken for the DNA assay...the same 5 samples were found to be positive with a second HIV-specific oligonucleotide...The serological [antibody] status was confirmed in each case and each of the 5 individuals was negative for anti-HIV antibodies and p24 antigen 2 and 3 months after the initial detection of HIV DNA”

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

“Detailed characterization of HTLV-III [now called HIV] and serologic [antibody] testing of large numbers of patients with AIDS or AIDS-related complex (ARC) became possible when it was found that the virus could be transmitted to a human T-cell line, H9, that is largely resistant to the cytopathic effects of the virus but is a good virus producer…Sequences of HTLV-III were first detected in DNA of infected H9 cells by Southern blot analysis…There was no evidence of such sequences in uninfected H9 cells…[but] In all, HTLV-III sequences were detected in only 9 out of 65 patients [with AIDS or ARC] evaluated. Roughly the same portion of ARC patients (3 out of 26) as AIDS patients (6 out of 39) was positive for HTLV-III…None of five Kaposi's sarcoma tissue specimens examined was positive for HTLV-III DNA sequences…Southern hybridizations can detect less than one viral DNA copy per 10 cells, [therefore] we interpreted these data to indicate that the three Kaposi's sarcoma lesions did not contain HTLV-III sequences…In most of the patients in which HTLV-III DNA sequences were detected in fresh tissue, the signal intensities were weak…[estimated as] at a level of less than one copyy per 10 cells…Unlike most other retroviruses, HTLV-III appears to persist in both integrated [DNA within the nucleus] and unintegrated [DNA floating around in the cell unattached] forms in chronically infected cells.”