Blenda Böttiger

Increase in viral gastroenteritis outbreaks in Europe and epidemic spread of new norovirus variant

BACKGROUND Highly publicised outbreaks of norovirus gastroenteritis in hospitals in the UK and Ireland and cruise ships in the USA sparked speculation about whether this reported activity was unusual. METHODS We analysed data collected through a collaborative research and surveillance network of viral gastroenteritis in ten European countries (England and Wales were analysed as one region). We compiled data on total number of outbreaks by month, and compared genetic sequences from the isolated viruses. Data were compared with historic data from a systematic retrospective review of surveillance systems and with a central database of viral sequences. FINDINGS Three regions (England and Wales, Germany, and the Netherlands) had sustained epidemiological and viral characterisation data from 1995 to 2002. In all three, we noted a striking increase in norovirus outbreaks in 2002 that coincided with the detection and emergence of a new predominant norovirus variant of genogroup II4, which had a consistent mutation in the polymerase gene. Eight of nine regions had an annual peak in 2002 and the new genogroup II4 variant was detected in nine countries. Also, the detection of the new variant preceded an atypical spring and summer peak of outbreaks in three countries. INTERPRETATION Our data from ten European countries show a striking increase and unusual seasonal pattern of norovirus gastroenteritis in 2002 that occurred concurrently with the emergence of a novel genetic variant. In addition to showing the added value of an international network for viral gastroenteritis outbreaks, these observations raise questions about the biological properties of the variant and the mechanisms for its rapid dissemination.

Etiology of Diarrhea in Young Children in Denmark: a Case-Control Study

ABSTRACT Infectious gastroenteritis is one of the most common diseases in young children. To clarify the infectious etiology of diarrhea in Danish children less than 5 years of age, we conducted a 2-year prospective case-control study. Stools from 424 children with diarrhea and 870 asymptomatic age-matched controls were examined, and their parents were interviewed concerning symptoms. Rotavirus, adenovirus, and astrovirus were detected by enzyme-linked immunosorbent assay, and norovirus and sapovirus were detected by PCR. Salmonella, thermotolerant Campylobacter, Yersinia, Shigella, and Vibrio spp. were detected by standard methods. Shiga toxin-producing (STEC), attaching-and-effacing (A/EEC), enteropathogenic (EPEC), enterotoxigenic, enteroinvasive, and enteroaggregative Escherichia coli were detected by using colony hybridization with virulence gene probes and serotyping. Parasites were detected by microscopy. Overall, a potential pathogen was found in 54% of cases. More cases than controls were infected with rotavirus, Salmonella, norovirus, adenovirus, Campylobacter, sapovirus, STEC, classical EPEC, Yersinia, and Cryptosporidium strains, whereas A/EEC, although common, was not associated with illness. The single most important cause of diarrhea was rotavirus, which points toward the need for a childhood vaccine for this pathogen, but norovirus, adenovirus, and sapovirus were also major etiologies. Salmonella sp. was the most common bacterial pathogen, followed by Campylobacter, STEC, Yersinia, and classical EPEC strains. A/EEC not belonging to the classical EPEC serotypes was not associated with diarrhea, underscoring the importance of serotyping for the definition of EPEC.

A New Human Retrovirus Isolate of West African Origin (SBL-6669) and Its Relationship to HTLV-IV, LAV-II, and HTLV-IIIB

A new human retrovirus of West African origin (SBL-6669) has been isolated from a patient with immunological and clinical signs of immunodeficiency. Using radioimmunoprecipitation assays (RIPA) and Western blot (WB) tests with human sera, the new virus isolate has been compared with HTLV-IV, LAV-II, and the HTLV-IIIB prototype strain of the human immunodeficiency virus (HIV). The West African isolates appeared to be members of the same virus group since their glycoproteins were antigenically indistinguishable. West African sera showed no detectable cross reaction with HTLV-IIIB glycoproteins. The external glycoprotein in the different virus strains only showed minor variations in size. The size of the transmembranous protein was not unambiguously defined. In the West African virus isolates a 30-35 kD protein was seen similar to the protein previously described possibly to represent this component. However, in SBL-6669 a distinct 41 kD protein was also identified. There were interstrain variations in the size of several viral proteins among the West African virus isolates. Only minor differences were seen between SBL-6669 and LAV-II. The variations were most pronounced in two core proteins corresponding to the 19 kD and 24 kD proteins of HTLV-IIIB. In addition, West African human retroviruses appear to differ in pathogenicity. LAV-II and SBL-6669 are associated with immunodeficiency, whereas HTLV-IV was isolated from healthy individuals. Since further spread of these viruses to other parts of the world is imminent, it is necessary to consider their antigenic and immunogenic properties in serodiagnosis of HIV infections and in planning for immunoprophylactic interventions.

Immunological changes in primary HIV-1 infection

Homosexual men with symptomatic primary HIV-1 infection displayed a pronounced lymphopaenia with significantly depressed numbers of CD3+, CD4+ and CD8+ cells and B cells during the first week of illness. Subsequently, the CD8+ cell counts rose in parallel with numbers of CD3+ cells, atypical lymphocytes and activated (CD38+ and HLA-Dr+) cells to attain maximal levels about a month following onset of illness. In contrast CD4+ and B cell numbers remained low for an extended period of time. Early signs of a host response included a transient appearance of interferon-alpha in the blood and raised levels of neopterin and beta 2-microglobulin (beta 2-M). Neither CD4+/CD8+ cell ratio nor beta 2-M resumed completely normal values during a follow-up period of 2 years. These findings shed some light on pathogenetic events during early HIV-1 infection and suggest that the infection, following the acute symptomatic stage, usually enters a stage of chronic active rather than latent infection.

Host Genetic Resistance to Symptomatic Norovirus (GGII.4) Infections in Denmark

ABSTRACT A total of 61 individuals involved in five norovirus outbreaks in Denmark were genotyped at nucleotides 428 and 571 of the FUT2 gene, determining secretor status, i.e., the presence of ABH antigens in secretions and on mucosa. A strong correlation (P = 0.003) was found between the secretor phenotype and symptomatic disease, extending previous knowledge and confirming that nonsense mutations in the FUT2 gene provide protection against symptomatic norovirus (GGII.4) infections.

Characterization of Rotavirus Strains in a Danish Population: High Frequency of Mixed Infections and Diversity within the VP4 Gene of P[8] Strains

ABSTRACT We characterized the G and P types from 162 rotavirus-positive stool specimens collected from 162 persons in Denmark (134 children and 28 adults) with acute diarrhea in 1998, 2000, and 2002. Samples were obtained during outpatient consultations (73%) and from hospitalized patients (27%). Although more than 20 different G-P combinations were identified, only 52% represented the globally most common types G1P[8], G2P[4], and G4P[8]. The G9 genotype, which is emerging worldwide, was identified in 12% of all samples. Twenty-one percent of the samples were of mixed genotypic origin, which is the highest frequency reported in any European population. The standard reverse transcription-PCR methods initially failed to identify a considerable fraction of the rotavirus P strains due to mutations at the VP4 primer-binding sites of P[8] strains. The application of a degenerate P[8] primer resulted in typing of most VP4 strains. There was considerable year-to-year variation among the circulating G-P types, and whereas G1P[8] was predominant in 1998 (42% of samples) and 2002 (26%), G2P[4] was the strain that was most frequently detected in 2000 (26% of samples). Our findings might implicate challenges for rotavirus vaccine implementation in a European population and underscore the importance of extensive strain surveillance prior to, during, and after introduction of any vaccine candidate.

Risk factors for diarrhea among children in an industrialized country.

Background: Risk factors for childhood diarrhea in industrialized countries are not well characterized, although diarrhea remains an important cause of morbidity. Methods: We conducted a case–control study of 422 cases and 866 controls over 22 months in Denmark. We selected cases among children under 5 years of age with diarrhea. Age-matched healthy controls were selected from the background population using a population register. Parents were interviewed about possible exposures and underlying conditions. In addition, stool samples from both cases and controls were analyzed for viruses, parasites, and bacteria. We analyzed risk factors for diarrhea in general and for diarrhea of a viral, bacterial, or “unknown” etiology using logistic regression. Results: The following factors were independently associated with an increased risk of diarrhea: recent foreign travel, contact with symptomatic persons (particularly in daycare centers), hospitalization, contact with a dog with diarrhea, private daycare, consumption of products containing formula milk, unemployment and low educational status of parents, and prior diagnosis of several types of atopic diseases. In a pathogenic-specific analysis of diarrhea of bacterial (73 patients), viral (88), or “unknown” (222) etiology, the major risk factor for viral diarrhea was contact with symptomatic persons. For bacterial diarrhea, foreign travel and socioeconomic factors were the main risk factors. Conclusions: Viral diarrhea appears to be transmitted predominantly from person to person, whereas bacterial diarrhea appears to be primarily foodborne. A substantial portion of the diarrheal episodes may be of noninfectious etiology. Limiting child-to-child transmission of disease in daycare centers may substantially reduce the disease burden.

Replicative capacity of HIV-2, like HIV-1, correlates with severity of immunodeficiency.

We have obtained 15 HIV-2 isolates from the peripheral blood mononuclear cells (PBMCs) of 24 HIV-2-infected west African people. The frequency of virus isolation correlated with the severity of HIV-2 infection; only three isolates were obtained from 11 asymptomatic individuals, whereas virus was isolated from nearly all (12 of 13) individuals with symptoms. The HIV-2 isolates showed distinct replicative and cytopathic characteristics and, similarly to HIV-1 isolates, could be divided into two major groups: rapid/high and slow/low. Rapid/high isolates, i.e. isolates with the ability to replicate in tumour cell lines, were obtained from individuals with symptomatic HIV-2 infection and CD4+ lymphocyte counts less than 360/microliters blood; these isolates induced syncytia in PBMC cultures. HIV-2 isolates unable to replicate continuously in tumour cell lines (slow/low isolates) induced small syncytia, cell death, or no cytopathic effect at all. All HIV-2 isolates obtained from asymptomatic individuals showed a slow/low replication pattern.

Clinical signs and laboratory markers in predicting progression to AIDS in HIV–1 infected patients

In a prospective longitudinal study 89 men with HIV-1 infection were observed for a mean time of 51 months with regard to clinical signs and laboratory findings predictive of progression to AIDS/opportunistic infection (OI). In a bivariate regression analysis the clinical signs showing a significant relation to AIDS development were: dermatitis of the face, yellow toe nail changes, hairly leukoplakia and oral candidiasis. The laboratory findings significantly associated with progression to AIDS were: decrease of the relative and absolute number of CD4 lymphocytes, decrease of the CD4/CD8 ratio, HIV p24 antigenaemia, lack of anti-HIV p24, elevated erythrocyte sedimentation rate, anaemia and elevated serum-beta-2-microglobulin. The relative number (%) of CD4 cells was found superior to the absolute number and the CD4/CD8 ratio. In a multivariate regression analysis decrease of CD4 lymphocytes and lack of anti-HIV p24 were independently associated with subsequent AIDS/OI development.

Transmission of HIV Infection to Heterosexual Partners but Not to Household Contacts of Seropositive Haemophiliacs

The prevalence of HIV antibodies in 44 heterosexual female partners and 56 nonsexual family household contacts of 61 HIV seropositive haemophiliacs (41 adults and 20 children or adolescents) was determined to evaluate the risk of transmission of HIV infection. HIV antibodies were determined by enzyme-linked immunosorbent assay and positive reactions were confirmed by Western blotting. HIV antibodies were demonstrated in 4/40 (10%) regular heterosexual partners of 40 seropositive patients with haemophilia A. Four temporary heterosexual partners of one additional seropositive haemophiliac were seronegative. 56 nonsexual household contacts including 30 parents, 13 siblings and 13 children of 29 seropositive haemophiliacs were all negative for HIV antibodies. Thus transmission of HIV occurred from seropositive haemophiliacs to their heterosexual partners but not to nonsexual household contacts.