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Genetic variation between geographically and temporally distinct isolates of dengue-1 (DEN-l) and dengue-4 (DEN-4) viruses was investigated. The nucleotide sequences of a fragment of the envelope protein gene encoding amino acids 28 to 87 of 35 DEN-1 isolates and 28 DEN-4 isolates were determined. Maximum nucleotide sequence variation was 6-9 % and 4-9% for DEN-1 and DEN-4 viruses, respectively. Taking a divergence of 6 % between the nucleotide sequences as the cut-off value, three genotype groups were defined for DEN-1 viruses, whereas only one was observed for DEN-4 viruses. Molecular analysis of isolates from the South Pacific permits the classification of the recent strains of DEN-1 (1988DEN-1 ( -1989 into a genotype distinct from the genotype which comprises earlier strains. This observation suggests that the recent epidemics were due to the introduction of a new genotype rather than to the re-emergence of the earlier strain.Dengue viruses occur as four distinct serotypes (DEN-1, DEN-2, DEN-3 and DEN-4), which can be differentiated serologically. Infection with any of the serotypes may result in a mild febrile disease (dengue fever, DF) or the more severe dengue haemorrhagic fever (DHF) and dengue shock syndrome. The four dengue viruses are transmitted by Aedes mosquitoes, and are responsible for a growing health problem in the tropical world, infecting millions of people annually (Gubler, 1991;Monath, 1994). Because of air travel, dengue is often seen in nonendemic areas as a disease of travellers (Monath, 1994).Dengue viruses are characterized by a positive-sense ssRNA genome of about 11 kb in length. The genome consists of a single open reading frame which encodes a precursor polyprotein (Rice et al., 1986). Proteolytic cleavages of the polyprotein result in the formation of the core (C), membrane (M) and envelope (E) proteins and the non-structural proteins NS 1, NS2A, NS2B, NS3, NS4A, NS4B and NS5. Studies of the molecular evolution and epidemiology of DEN-l, DEN-2 and DEN-3 viruses using genome sequence relatedness have demonstrated the occurrence of genotype groupings among these viruses (Rico-Hesse, 1990;Deubel et al., 1993;Chungue et al., 1993b, Lewis et al., 1993Lanciotti et al., 1994).In this paper, we have analysed 35 DEN-1 and 28 * Author for correspondence. Fax +689 43 15 90. e-mail chungue@pasteur.fr DEN-4 virus strains isolated over periods of 50 years and 33 years, respectively, from humans and mosquitoes. Included in this study were the isolates of DEN-4 from humans and mosquitoes obtained from 1979 to 1988 in Tahiti where only DEN-4 virus was transmitted continuously (Chungue et al., 1993c). Gene fragments corresponding to the 5' terminus of the E protein gene were amplified by RT-PCR and sequenced directly. This method was valuable for epidemiological analysis and classification of DEN-2 and DEN-3 viruses Chungue et al., 1993b) and was therefore applied to the genetic study of DEN-1 and DEN-4 viruses.The characteristics of each strain used in this study are depicted in Tables 1 and 2. Vi...

We examined the cause of death during a 12-month period (1995196) in all consecutive patients admitted to hospital with leptospiral infection in Seychelles (Indian Ocean), where the disease is endemic. Leptospirosis was diagnosed by use of the microscopic agglutination test and a specific polymerase chain reaction assay on serum samples. Seventy-five cases were diagnosed and 6 patients died, a case fatality of 8%. All 6 patients died within 9 days of onset of symptoms and within 2 days of admission for 5 of them (5 days for the 6th). On autopsy, diffuse bilateral pulmonary haemorrhage (PH) was found in all fatalities. Renal, cardiac, digestive and cerebral haemorrhages were also found in 5,3,3 and 1 case(s), respectively. Incidentally, haemoptysis and lung infiltrate on chest radiographs, which suggest PH, were found in 8 of the 69 non-fatal cases. Dengue and hantavirus infections were ruled out. In conclusion, PH appeared to be a main cause of death in leptospirosis in this population, although haemorrhage in other organs may also have contributed to fatal outcomes. This cause of death contrasts with the findings generally reported in endemic settings.

The nucleotide sequences of a short fragment of the envelope protein gene encoding amino acids 25 to 89 of 27 dengue 3 viruses were determined by direct sequencing of PCR-amplified products, and the viruses were compared regarding their time of isolation and geographic distribution. Four distinct genotypic groups were discerned at 6 % divergence between nucleotide sequences. The first group contained isolates from the South Pacific (1988 to 1992), Singapore (

A dual viremia resulting from naturally acquired dengue 1 and dengue 3 infections in six patients experiencing a dengue-like syndrome during the epidemic in New Caledonia in 1989 is reported. Serotype identification was first based on virus isolation in mosquito cells and immunofluorescence using type-specific monoclonal antibodies. The double infection was confirmed directly in blood samples by the polymerase chain reaction (PCR) on genomic RNA and hybridization of the amplified cDNA fragments with type-specific DNA probes.