Virus discovery in chronic inflammatory demyelinating polyneuropathy.

The events triggering and/or sustaining the auto-immune response underlying chronic inflammatory demyelinating polyneuropathy (CIDP) are unknown. Similar to Guillain-Barré syndrome (GBS), a viral infection might play a role in CIDP. In this study, an virus detection method (VIDISCA-next generation sequencing) capable of detecting known and unknown viruses, was used to analyze the virome in serum of 47 CIDP patients at different time points of the disease and, when available, in cerebrospinal fluid (CSF) samples (N: 17). Serum samples of GBS patients (N:24) and healthy controls (N:114) were used for comparisons. In 5/47 (10.6%; 95% CI: 4-23) CIDP samples, 10/24 (42%; 95% CI: 22-63) GBS samples and 32/114 (28.1%; 95% CI: 20-37) healthy controls samples, anelloviruses were detected, generally regarded as a non-pathogenic species. Parvovirus B19 and GB virus C were found in two CIDP samples (4%). Parvovirus B19, HIV-1 and GB virus C were found in three GBS samples (13%). In 2/17 CIDP CSF samples, an anellovirus and polyomavirus were detected, probably due to contamination during lumbar puncture. No sequences of other viruses were detected in serum or CSF. A (persistent) viral infection sustaining the auto-immune response in CIDP seems therefore unlikely.

Diagnostic accuracy of VIDISCA-NGS in patients with suspected central nervous system infections.

OBJECTIVES: Confirming the diagnosis in viral central nervous system (CNS) infections can be difficult with the currently available diagnostic tools. Virus discovery cDNA-amplified fragment length polymorphism next-generation sequencing (VIDISCA-NGS) is a promising viral metagenomic technique that enables the detection of all viruses in a single assay. We performed a retrospective study on the diagnostic accuracy of VIDISCA-NGS in cerebrospinal fluid (CSF) of individuals with suspected CNS infections. METHODS: Consecutive adult patients presenting to the Emergency Department or inpatients, who underwent a lumbar puncture for the suspicion of a CNS infection, were included if they were diagnosed with a viral CNS infection, or if a viral CNS infection was initially suspected but eventually a different diagnosis was made. A quantitative PCR panel of the most common causative viruses was performed on CSF of these patients as reference standard and compared with the results of VIDISCA-NGS, the index test. RESULTS: We included 38 individuals with viral CNS infections and 35 presenting with suspected CNS infection for whom an alternative aetiology was finally established. Overall sensitivity and specificity were 52% (95% CI 31%-73%) and 100% (95% CI 91%-100%), respectively. One enterovirus, detected by VIDISCA-NGS, was only identified by quantitative PCR upon retesting. Additional viruses identified by VIDISCA-NGS consisted of GB virus C, human papillomavirus, human mastadenovirus C, Merkel cell polyoma virus and anelloviruses. CONCLUSION: In patients for whom routine diagnostics do not yield a causative pathogen, VIDISCA-NGS can be of additional value as it can detect a broader range of viruses, but it does not perform well enough to replace quantitativePCR.

Are low and high utilization related to the way GPs manage their practices? An observational study.

BACKGROUND: General practice care plays a key role in keeping healthcare effective and cost-efficient. However, variation in the utilization rates of practices may reveal variation in practice performance. Our research goal is to investigate whether the socio-demographic profile of the patients' area of residence and practice organization characteristics influence the low or high utilization of general practice care. METHODS: Data on the utilization of general practice care were derived from the electronic health records of 232 general practices participating in the NIVEL Primary Care Database for the year 2013. Census data for the year 2013 were matched with the postal code of the patients. A small area estimation (SAE) technique was used to calculate the estimated utilization rate for general practice care per practice based on the socio-demographic profile of the patients' area of residence. Subsequently, the actual utilization rates were compared to the estimated rates per practice. Linear regression analysis was used to link the differences between the actual and estimated utilization rates to practice organization characteristics. RESULTS: The socio-demographic profile of the patients' area of residence accounted for 25.7% of the estimated utilization rates per practice. Practice organization characteristics accounted for 19.3% of the difference between the actual utilization rates and the estimated rates. Practices had higher utilization rates than estimated when a practice was a dual practice, when it employed female GPs, when it employed other healthcare providers and/or when it offered more services related to a disease management programme. CONCLUSION: We found that utilization rates of general practice care can be partially explained by the socio-demographic profile of the patients' area of residence, but also by practice organization characteristics. Insight into these factors provides both GPs and the other stakeholders involved in the organization of general practice care with information to help reflect on the utilization of care.

Changes in microbiota during experimental human Rhinovirus infection.

BACKGROUND: Human Rhinovirus (HRV) is responsible for the majority of common colds and is frequently accompanied by secondary bacterial infections through poorly understood mechanisms. We investigated the effects of experimental human HRV serotype 16 infection on the upper respiratory tract microbiota. METHODS: Six healthy volunteers were infected with HRV16. We performed 16S ribosomal RNA-targeted pyrosequencing on throat swabs taken prior, during and after infection. We compared overall community diversity, phylogenetic structure of the ecosystem and relative abundances of the different bacteria between time points. RESULTS: During acute infection strong trends towards increases in the relative abundances of Haemophilus parainfluenzae and Neisseria subflava were observed, as well as a weaker trend towards increases of Staphylococcus aureus. No major differences were observed between day-1 and day 60, whereas differences between subjects were very high. CONCLUSIONS: HRV16 infection is associated with the increase of three genera known to be associated with secondary infections following HRV infections. The observed changes of upper respiratory tract microbiota could help explain why HRV infection predisposes to bacterial otitis media, sinusitis and pneumonia.

Short- and long-term effects of real-time medication monitoring with short message service (SMS) reminders for missed doses on the refill adherence of people with Type 2 diabetes: evidence from a randomized controlled trial.

AIMS: To investigate short- and long-term effects of real-time monitoring medication use combined with short message service (SMS) reminders for missed doses on refill adherence to oral anti-diabetic medication. METHODS: A randomized controlled trial with two intervention groups and one control group involving 161 participants with Type 2 diabetes with suboptimal adherence. For 6 months, participants in the SMS group (n = 56) were monitored and received SMS reminders if they missed their medication. Participants in the non-SMS group (n = 48) were only monitored. The control group (n = 57) was not exposed to any intervention. Primary outcome measure was refill adherence to oral anti-diabetic medication. Multi-level regression analyses were performed to examine intervention effects on adherence between and within groups after 1 and 2 years of follow-up. RESULTS: At baseline, mean refill adherence was comparable between the groups. After 1 year, adherence in the SMS group was significantly higher than in the control group (79.5% vs. 64.5%; P < 0.001) and showed a significant improvement from baseline (+16.3%; P < 0.001). Mean adherence in the non-SMS group reached 73.1% (+7.3%; P < 0.05), but did not differ from the control group (P = 0.06). After 2 years, the improved adherence in the SMS group persisted and remained significantly higher than in the control group (80.4% vs. 68.4%; P < .01), contrary to the non-SMS group whose adherence approached baseline level again (65.5%). CONCLUSIONS: This study shows the long-term effectiveness of real-time medication monitoring combined with SMS reminders in improving refill adherence. This new reminder system can strengthen the self-management of people with diabetes.

Specific serology for emerging human coronaviruses by protein microarray.

We present a serological assay for the specific detection of IgM and IgG antibodies against the emerging human coronavirus hCoV-EMC and the SARS-CoV based on protein microarray technology. The assay uses the S1 receptor-binding subunit of the spike protein of hCoV-EMC and SARS-CoV as antigens. The assay has been validated extensively using putative cross-reacting sera of patient cohorts exposed to the four common hCoVs and sera from convalescent patients infected with hCoV-EMC or SARS-CoV.

Comprehensive full-length sequence analyses of human parechoviruses: diversity and recombination.

Human parechoviruses (HPeVs) are highly prevalent pathogens among very young children. Although originally classified into two serologically distinct types, HPeV1 and -2, recent analyses of variants collected worldwide have revealed the existence of 12 further types classified genetically by sequence comparisons of complete genome sequences or the capsid (VP1) gene. To investigate the nature of HPeV evolution, its population dynamics and recombination breakpoints, this study generated 18 full-length genomic sequences of the most commonly circulating genotypes, HPeV1 and -3, collected over a time span of 14 years from The Netherlands. By inclusion of previously published full-length sequences, 35 sequences were analysed in total. Analysis of contemporary strains of HPeV1 and those most similar to the prototype strain (Harris) showed that HPeV1 variants fall into two genetically distinct clusters that are much more divergent from each other than those observed within other HPeV types. Future classification criteria for HPeVs may require modification to accommodate the occurrence of variants with intermediate degrees of diversity within types. Recombination was frequently observed among HPeV1, -4, -5 and -6, but was much more restricted among HPeV3 strains. Favoured sites for recombination were found to flank the capsid region, and further sites were found within the non-structural region, P2. In contrast to other HPeV types, the majority of the HPeV3 sequences remained monophyletic across the genome, a possible reflection of its lower diversity and potentially more recent emergence than other HPeV types, or biological and/or epidemiological constraints that limit opportunities for co-infections with potential recombination partners.

Clinical manifestations of human coronavirus NL63 infection in children in Taiwan.

Human coronavirus NL63 (HCoV-NL63) is a global respiratory tract pathogen; however, the epidemiology of this virus in subtropical area is not well known. To evaluate the epidemics and disease spectrum of HCoV-NL63 infection in children in Taiwan, we prospectively screened children admitted to the hospital with respiratory tract infection from May 2004 to April 2005. Every enrolled child had a nasopharyngeal aspirate (NPA) sample taken. Quantitative RT-PCR was used to detect 1b gene of HCoV-NL63. A total of 539 NPAs were collected. Seven (1.3%) were positive for HCoV-NL63. All cases were boys younger than 3 years of age and most cases occurred in autumn. Co-infection with other pathogens was observed in three cases. The most common symptoms/signs of HCoV-NL63 infection were cough, fever, and inspiratory stridor. HCoV-NL63 was the most common pathogen (14.7%) in children with croup and was the cause of three cases of croup in October. The odds ratio of croup in children infected with HCoV-NL63 was 43.4 (95% CI 8.1 approximately 233.1). In conclusion, HCoV-NL63 is an important respiratory tract pathogen as the main cause in children admitted to the hospital in Taiwan.

Antiviral strategies against human coronaviruses.

Since the mid 60's the human coronaviruses (HCoV), represented by HCoV-OC43 and HCoV-229E, were generally considered relatively harmless viruses. This status changed dramatically with the emergence of SARS-CoV in 2002/2003. The SARS-CoV pandemic took 774 lives around the globe and infected more than 8000 people in 29 countries. SARS-CoV is believed to be of zoonotic origin, transmitted from its natural reservoir in bats through several animal species (e.g., civet cats, raccoon dogs sold for human consumption in markets in southern China). The epidemic was halted in 2003 by a highly effective global public health response, and SARS-CoV is currently not circulating in humans. The outbreak of SARS-CoV and the danger of its re-introduction into the human population, as well as the danger of the emergence of other zoonotic coronaviral infections triggered an intense survey for an efficient treatment that resulted in the evaluation of several anticoronaviral compounds. HCoV-NL63 and HCoV-HKU1 were identified shortly after the SARS-CoV outbreak. The 4 human coronaviruses HCoV-229E, HCoV-OC43, HCoV-NL63 and HCoV-HKU1 cause mild respiratory illnesses when compared to SARS, but these infections are involved in 10 - 20 % of hospitalizations of young children and immunocompromised adults with respiratory tract illness. Therefore, there is an urgent need for a successful therapy to prevent disease induction or a vaccine to prevent new infections. This review summarizes the current status of anticoronaviral strategies.

Detection of human coronavirus NL63, human metapneumovirus and respiratory syncytial virus in children with respiratory tract infections in south-west Sweden.

Two recently detected viruses, human metapneumovirus (hMPV) and coronavirus NL63 (HCoV-NL63), have been associated with acute respiratory tract infections, particularly in young children. This study investigated the frequency of hMPV and HCoV-NL63 infections in Swedish children by screening 221 nasopharyngeal aspirates, collected between November 2003 and May 2005, from 212 children attending the paediatric department of a county hospital in Sweden or submitted from local general practitioners. The samples were originally submitted to be tested for respiratory syncytial virus (RSV), and were examined retrospectively for hMPV and HCoV-NL63 by RT-PCR. Of the 212 patients, 101 were positive for RSV (48%), 22 (10%) were positive for hMPV, and 12 (6%) were positive for HCoV-NL63. The frequency of HCoV-NL63 infection increased from 1% in 2003-2004 to 10% in 2004-2005. Sequence analysis of parts of the coronavirus genomes showed considerable similarity to the HCoV-NL63 prototype sequence. The study demonstrated that HCoV-NL63 and hMPV occur in south-west Sweden with essentially the same frequency, seasonal distribution and clinical characteristics as have been reported in other countries.

Carrier rate of zidovudine-resistant HIV-1: the impact of failing therapy on transmission of resistant strains.

OBJECTIVE: Because maintenance of treatment success in HIV-1 infection requires viruses to remain therapy sensitive in drug-naive seropositive persons, we looked at the primary infections caused by drug-resistant HIV-1 over time. Furthermore, to study the coverage rate of therapy and therapy failure in relation to the transmission of resistant viruses a mathematical model was developed. DESIGN: The reverse transcriptase and protease genes of viruses were analysed in newly infected people in the period 1990-1998 in the Amsterdam Cohort Study on HIV infection and AIDS in homosexual men. METHODS: The mathematical model was based on the coverage of drug regimens selecting zidovudine (ZDV) resistance, the lag time in which resistance is gained or lost, the death rate of people infected with resistant virus, and the replacement of resistance-selecting regimens by more potent treatments that substantially reduce viral load and mortality. RESULTS: Of 43 individuals with a primary HIV-infection, three (7%) harboured ZDV-resistant viruses. The first of the ZDV-resistant strains was transmitted in 1995, the last two in 1996. The build-up of ZDV resistance was described by the mathematical model indicating that the equilibrium level of resistance due to treatment depends only on the treatment rate and the outflow rate of patients with resistance virus. CONCLUSIONS: Our model indicates that the frequency of viral resistance in a population is determined largely by the number of individuals on insufficient or failing therapy and is influenced only modestly by secondary transmission of ZDV-resistant strains.

Establishment of new transmissible and drug-sensitive human immunodeficiency virus type 1 wild types due to transmission of nucleoside analogue-resistant virus.

Sequence analysis of human immunodeficiency virus type 1 (HIV-1) from 74 persons with acute infections identified eight strains with mutations in the reverse transcriptase (RT) gene at positions 41, 67, 68, 70, 215, and 219 associated with resistance to the nucleoside analogue zidovudine (AZT). Follow-up of the fate of these resistant HIV-1 strains in four newly infected individuals revealed that they were readily replaced by sensitive strains. The RT of the resistant viruses changed at amino acid 215 from tyrosine (Y) to aspartic acid (D) or serine (S), with asparagine (N) as a transient intermediate, indicating the establishment of new wild types. When we introduced these mutations and the original threonine (T)-containing wild type into infectious molecular clones and assessed their competitive advantage in vitro, the order of fitness was in accord with the in vivo observations: 215Y < 215D = 215S = 215T. As detected by real-time nucleic acid sequence-based amplification with two molecular beacons, the addition of AZT or stavudine (d4T) to the viral cultures favored the 215Y mutant in a dose-dependent manner. Our results illustrate that infection with nucleoside analogue-resistant HIV leads in newly infected individuals to mutants that are sensitive to nucleoside analogues, but only a single mutation removed from drug-resistant HIV. Such mutants were shown to be transmissible, stable, and prone to rapid selection for resistance to AZT or d4T as soon as antiretroviral therapy was administered. Monitoring of patients for the presence of new HIV-1 wild types with D, S, or N residues at position 215 may be warranted in order to estimate the threat to long-term efficacy of regimens including nucleoside analogues.

Slower decline of plasma HIV-1 RNA following highly suppressive antiretroviral therapy in primary compared with chronic infection.

OBJECTIVES: To study the effect of highly suppressive antiretroviral therapy on the slopes of HIV-1 RNA decline in primary compared with chronic HIV-1 infection. METHODS: Slopes of HIV-1 RNA decline in plasma were compared before and after the start of highly suppressive antiretroviral therapy from five acutely infected patients who started treatment 2 to 5 weeks following the onset of clinical symptoms. Slopes of decline after the initiation of therapy were also compared with those found in 12 chronically infected individuals on the same therapy. Numbers and percentages of activated CD4 and CD8 T cells at baseline were compared as well. RESULTS: The pre-treatment slopes of HIV-1 RNA decline in the acutely infected individuals increased significantly (P = 0.0001) after the start of anti-retroviral therapy. However, these post-treatment slopes were lower than those found in the chronically infected individuals (P= 0.012). Slopes were inversely correlated (P= 0.012) with baseline HIV-1 RNA. Although the number of CD38+HLA-DR+ CD4 cells was higher in primary infection (P= 0.02), the percentage did not differ between primary and chronic infection. CONCLUSIONS: These findings indicate that antiretroviral therapy contributes significantly to the clearance of HIV-1 during primary infection. Based on the mathematical model the less steep RNA slope following the start of treatment in primary infection can be predicted to be the result of lower clearance of productively infected cells and higher burst size per cell per unit time. This may indicate a growing immune response to HIV-1 in this very early stage of infection.

Human immunodeficiency virus type 1 in faeces and serum: evidence against independently evolving subpopulations.

It is not known whether independent tissue-specific evolution accounts for the differences between human immunodeficiency virus type 1 (HIV-1) subpopulations in intestinal tissue and blood. To study this, sequential serum samples from three persons were analysed for the presence of HIV-1 V3 genotypes which were detected exclusively in faeces at a specific time-point. For two persons the faeces genotype was found in serum samples collected before the time of faeces collection: 7 months for one person and 32 months for the other person. In the third person, serum collected 1 month after faeces collection contained the faeces genotype in abundance. These data indicate that a difference between intestinal tissue and blood HIV-1 subpopulations is not the result of complete compartmentalization and independent HIV-1 evolution in intestinal tissue, but that it reflects an unequal distribution of HIV-1 in different tissues.

Genetic differences between human immunodeficiency virus type 1 subpopulations in faeces and serum.

To study human immunodeficiency virus type 1 (HIV-1) compartmentalization between intestine and blood, paired faecal and serum samples were collected from 204 HIV-1-infected persons. Direct sequencing of the gp120 V3 region obtained from 33 persons showed that faecal and serum sequences could be nearly homologous (0.3% different) or very dissimilar (11.3% different). Individual clones were obtained and sequenced from the faecal and serum samples of 13 persons. In 6 persons the HIV-1 subpopulations in faeces and serum were similar, whereas in 7 persons, distribution of V3 genotypes showed a marked difference. Genetic characterization of the HIV-1 subpopulations showed less heterogeneity in faecal subpopulations than in serum subpopulations in 5 of the 7 subjects. Furthermore, faecal and serum subpopulations differed predominantly by nonsynonymous nucleotide substitutions (in 6 of 7 persons). Comparison of the HIV-1 subpopulations in faeces and serum of these 7 persons, using resampling techniques, revealed a significant difference between faecal and serum subpopulations at an N-linked glycosylation site, C-terminal of the V3 loop (amino acids 331-333). Sequences from faecal subpopulations of all 7 persons contained a glycosylation site at amino acid position 331-333. Four of these 7 harboured serum variants lacking a glycosylation site at this position. The faecal subpopulations in these 4 persons showed limited nonsynonymous substitutions compared to synonymous substitutions, indicating that purifying selection is operational on these subpopulations.

Prevalence of Campylobacter-associated diarrhea among patients infected with human immunodeficiency virus.

We performed a cross-sectional study at an outpatient AIDS clinic to assess the prevalence of Campylobacter species in stool specimens from 201 consecutive patients infected with human immunodeficiency virus (HIV). We characterized campylobacters phenotypically and genetically by using primers for the group of common species (i.e., C. jejuni, C. coli, C. lari, and C. upsaliensis) and for most individual uncommon species. We performed cultures with use of a membrane filter technique on nonselective blood agar and found that Campylobacter species were the most frequent enteropathogenic bacteria: the organisms were recovered from 7 (16%) of 43 patients with diarrhea and 5 (3%) of 158 patients without diarrhea (P = .001). We isolated only one campylobacter with use of conventional culture techniques on selective media. Phenotypic characterization of 10 campylobacter strains resulted in the misidentification of four isolates. C. upsaliensis was the most frequently isolated species, followed by C. jejuni and C. coli. Two strains could not be identified with the available primers. Two of 12 Campylobacter strains were resistant to erythromycin, and two were resistant to ciprofloxacin. We conclude that Campylobacter species other than C. jejuni can frequently be detected in the stools of HIV-infected patients and that these organisms could be associated with diarrhea.

Human immunodeficiency virus type 1 RNA populations in faeces with higher homology to intestinal populations than to blood populations.

To determine whether human immunodeficiency virus type 1 (HIV-1) in faeces is representative of the HIV-1 population in intestinal tissue, we studied HIV-1 V3 variation in faeces, intestinal biopsies and serum from two individuals. Phylogenic analysis of HIV-1 V3-coding RNA in faeces from one individual showed three distinct genotypes. Viruses belonging to all three genotypes were also present in sigmoidal tissue and in serum. Jejunal tissue contained two of these three genotypes. Analysis of the V3-coding RNA in faeces of the other individual showed five distinct genotypes. One of these genotypes was present in all specimens from this individual. Besides this shared genotype, jejunal tissue and serum contained sequences belonging to one other genotype. In addition, one of the other three V3 variants was detected in sigmoidal tissue. For both persons the shared HIV-1 RNA genotypes in faeces and serum displayed a distinctly different frequency distribution. In one individual, the genotype which was detected in a majority of the clones in faeces (59%) and as a minority in serum (11%), was the most abundant genotype in jejunal and sigmoidal tissue (61% and 80%, respectively). For the other individual the genotype that was present in faeces in a significant number of clones (43%) was detected in serum as a minority (8%), whereas this genotype composed 47% of the clones isolated from jejunal tissue. Taken together these data suggest that faeces contain HIV-1 sequences that are derived from local HIV-1 replication in intestinal tissue.

Syncytium-inducing (SI) phenotype suppression at seroconversion after intramuscular inoculation of a non-syncytium-inducing/SI phenotypically mixed human immunodeficiency virus population.

Two distinct biological phenotypes of human immunodeficiency virus (HIV) have been described: the non-syncytium-inducing (NSI) phenotype, best characterized by the inability to infect MT-2 cells, and the syncytium-inducing (SI) phenotype, with the ability to infect MT-2 cells. The earliest virus population observed following HIV transmission is generally of the NSI phenotype, even after exposure to inocula of mixed NSI/SI phenotype. In this study, the issue of intrapatient selection of virus phenotype following transmission was addressed by studying two cases of accidental transmission. A comparison of the sequences of the V1-V2 and the V3 coding regions of the envelope gene and the p17 region of the gag gene showed that the donor-recipient pairs were tightly clustered in all gene segments, but away from local and published transmission controls. The intrasample variation of the p17 sequence was greater in the recipients and smaller in the donors than that of the V3 region sequence, indicating selection of V3 at transmission. In these transmission cases, the effects of an intravenous inoculation of a small quantity of blood containing predominantly SI V3 sequences (6 of 8 clonal sequences) were compared with those of an intramuscular inoculation of a large quantity of blood containing predominantly NSI viruses (14 of 16 clonal sequences). Both SI and NSI V3 regions were demonstrated to be phenotypic expressions of genetically related viral strains. The inoculation of the predominantly SI virus population resulted in the persistence of an SI virus population in the recipient and a rapid CD4+ T-cell decline. The inoculation of the predominantly NSI population resulted in a selective amplification of SI viruses before seroconversion, followed by a suppression of SI viruses at seroconversion and a rapid decline of CD4+ T-cell numbers. These data suggest that the suppression of SI viruses can be accomplished following the development of HIV-specific immunity and that the ability to suppress SI viruses does not prevent the development of immunodeficiency.

Isolation of human immunodeficiency virus type 1 (HIV-1) RNA from feces by a simple method and difference between HIV-1 subpopulations in feces and serum.

A simple method for the isolation and subsequent detection of human immunodeficiency virus type 1 (HIV-1) RNA from feces is described. Viral RNA was isolated by the method developed by Boom et al. (R. Boom, C.J.A. Sol, M.M.M. Salimans, C.L. Jansen, P.M.E. Wertheim-van Dillen, and J. van der Noordaa, J. Clin. Microbiol. 28:495-503, 1990), which was adapted for feces. HIV-1 RNA was detected by reverse transcription (RT) followed by a nested PCR encompassing the V3 region. Reconstruction experiments revealed that the efficiencies of the extraction technique and the subsequent RT-PCR were not considerably affected by the varied composition of feces. The method was applied on fecal specimens from 18 HIV-1-infected individuals, among which were samples that had been stored for 9 years. It appeared that HIV-1 RNA was detectable in the feces of 12 persons (67%). Viral RNA was present in the feces of persons who fulfilled the criteria for CDC class II and CDC class III HIV infection as well as in patients who were diagnosed with AIDS (CDC class IV). Direct sequencing of amplimers obtained from paired fecal and serum specimens showed that differences in sequence heterogeneity existed. In one patient a remarkable difference in the HIV-1 sequences between isolates from feces and serum was observed. In conclusion, HIV-1 RNA is frequently present in the feces of HIV-1-infected individuals, and in some cases the HIV-1 subpopulation in feces differs from the HIV-1 subpopulation in serum.