Charles F. Longer

Malaria among United States troops in Somalia

PURPOSE United States military personnel deployed to Somalia were at risk for malaria, including chloroquine-resistant Plasmodium falciparum malaria. This report details laboratory, clinical, preventive, and therapeutic aspects of malaria in this cohort. PATIENTS AND METHODS The study took place in US military field hospitals in Somalia, with US troops deployed to Somalia between December 1992 and May 1993. Centralized clinical care and country-wide disease surveillance facilitated standardized laboratory diagnosis, clinical records, epidemiologic studies, and assessment of chemoprophylactic efficacy. RESULTS Forty-eight cases of malaria occurred among US troops while in Somalia; 41 of these cases were P falciparum. Risk factors associated with malaria included: noncompliance with recommended chemoprophylaxis (odds ratio [OR] 2.4); failure to use bed nets (OR 2.6); and failure to keep sleeves rolled down (OR 2.2). Some patients developed malaria in spite of mefloquine (n = 8) or doxycycline (n = 5) levels of compatible with chemoprophylactic compliance. Five mefloquine failures had both serum levels > or = 650 ng/mL and metabolite:mefloquine ratios over 2, indicating chemoprophylactic failure. All cases were successfully treated, including 1 patient who developed cerebral malaria. CONCLUSIONS P falciparum malaria attack rates were substantial in the first several weeks of Operation Restore Hope. While most cases occurred because of noncompliance with personal protective measures or chemoprophylaxis, both mefloquine and doxycycline chemoprophylactic failures occurred. Military or civilian travelers to East Africa must be scrupulous in their attention to both chemoprophylaxis and personal protection measures.

Phylogenetic analysis of hepatitis E virus isolates from Egypt.

Hepatitis E virus (HEV) genome was detected by reverse transcriptase‐polymerase chain reaction (RT‐PCR) in fecal samples of two sporadic cases of hepatitis E in Cairo Egypt. Sequence of the complete putative structural region [open reading frame (ORF)‐2] and complete region of unknown function (ORF‐3) was determined for the two HEV isolates. Phylogenetic analysis of the nucleotide sequences was performed using neighbor joining or maximum parsimony methods of tree reconstruction. Direct correspondence between the HEV evolutionary trees and geographic origin of the HEV isolates was observed. Three genotypes of HEV were identified: genotype I (Asia‐Africa), genotype II (US), and genotype III (Mexico). Genotype I was further divided into two subgenotypes (Asia and Africa). In the Asian subgenotype, three smaller genetic clusters were observed (China‐like sequences, Burma‐like sequences, and sequence from a fulminant case of HEV). The segregation of all these genetic clusters was supported by the high level of bootstrap probabilities. Four regions of the HEV genome were used for phylogenetic analysis. In all four regions, Egyptian HEV isolates were grouped in a separate African clade. J. Med. Virol. 57:68–74, 1999.

Rates of Hepatitis E Virus Infection and Disease among Adolescents and Adults in Kathmandu, Nepal

To determine hepatitis E virus (HEV) infection and disease rates in the Kathmandu Valley of Nepal, serum was collected from 757 healthy Nepalese (ages 12-48 years) during March and September 1992 and September 1993. At each visit, reports of interval illness were obtained. Sera were examined for IgG to HEV, using a commercially available kit. Seroconversion was used as a marker for HEV infection, and an episode of hepatitis E was defined as a history of jaundice with seroconversion. Seroprevalence ranged from 16% to 31% and increased with age, whereas both infection and disease rates decreased with age. Infection and disease rates were as high as 99/1000 and 45/1000 person-years, respectively. These results highlight the importance of sporadic hepatitis E as a public health problem among adolescents and young adults in this region.

Characterization of hepatitis E virus (HEV) from Algeria and Chad by partial genome sequence

The purpose of this study was to analyze partial nucleotide sequences and derived peptide sequences of hepatitis E virus (HEV) from two outbreaks of hepatitis E in Africa (Chad 1983–1984; Algeria 1978–1980). A portion of ORF3 and the major portion of ORF2 were amplified by Reverse Transcriptase‐Polymerase Chain Reaction (RT‐PCR). The PCR products were sequenced directly or after cloning into the pCRII vector. Sequences were then compared to the corresponding regions of reported full length HEV sequences. In the ORF2 and ORF3 regions, the homology between the Algerian and the Chad isolates at the nucleic acid level was 92 and 95%, respectively. At the peptide level the homology was 98% in both regions. In these regions, both strains are more related to Asian strains at the nucleic acid level (89 to 95%) and at the amino acid level (95 to 100%) than to the Mexico strain. At the peptide level the differences are less apparent. Both African isolates have amino acid changes in common with some reference strains although the Chad isolate has three unique changes. These African strains of HEV, based on the ORF2 and ORF3 phylogenetic trees, appear to be a distinct phylogenetic group, separate from the Mexican and Asian strains. J. Med. Virol. 53:340–347, 1997.

Quantitation of Immunoglobulin to Hepatitis E Virus by Enzyme Immunoassay

ABSTRACT We developed a quantitative enzyme immunoassay (EIA) for antibody to hepatitis E virus (HEV) by using truncated HEV capsid protein expressed in the baculovirus system to improve seroepidemiology, to contribute to hepatitis E diagnosis, and to enable vaccine evaluations. Five antigen lots were characterized; we used a reference antiserum to standardize antigen potency. We defined Walter Reed antibody units (WR U) with a reference antiserum by using the four-parameter logistic model, established other reference pools as assay standards, and determined the conversion factor: 1 WR U/ml = 0.125 World Health Organization unit (WHO U) per ml. The EIA performed consistently; median intra- and intertest coefficients of variation were 9 and 12%, respectively. The accurate minimum detection limit with serum diluted 1:1,000 was 5.6 WR U/ml; the test could detect reliably a fourfold antibody change. In six people followed from health to onset of hepatitis E, the geometric mean antibody level rose from 7.1 WR U/ml to 1,924.6 WR U/ml. We used the presence of 56- and 180-kDa bands by Western blotting as a confirmatory test and to define true-negative and -positive serum specimens. A receiver-operating characteristics plot identified 30 WR U/ml as an optimum cut-point (sensitivity, 86%; specificity, 89%). The EIA detected antibody more sensitively than a commercially available test. The EIA was transferred to another laboratory, where four operators matched reference laboratory results for a panel of unknowns. Quantitation of antibody to HEV and confirmation of its specificity by Western blotting make HEV serology more meaningful.

Hepatitis E virus: complete genome sequence and phylogenetic analysis of a Nepali isolate

The complete nucleotide sequence of the Nepali strain TK15/92 of hepatitis E (HEV) was determined. It showed the highest sequence homology with the Burmese B1 strain, but closer evolutionary relatedness to the Indian strains. Difficulties in reverse-transcribing and amplifying the hypervariable region in ORF1 suggested that strong secondary structures might be intrinsically responsible for the high mutational rate observed in this region of the HEV genome.

Epidemiology of Hepatitis E Virus (HEV): A Cohort Study in Kathmandu, Nepal

Hepatitis E is endemic to Kathmandu. To determine age- and sex-specific prevalence and risk factors for antibody to HEV (anti-HEV), blood and historic information was obtained from 4486 volunteers in March 1992 and again in September. Sera were assayed for anti-HEV IgG by enzyme immunoassay. The anti-HEV prevalence (10.2%) was greater in males than in females, and was shown to increase with age in males but not females. Anti-HEV prevalence was greater in subjects with a recent history of jaundice, but most anti-HEV positive subjects had no such history. We concluded that anti-HEV prevalence in female and male children was similar, prevalence in males increased during the young adult years, prevalence was greater in subjects with a recent history of jaundice, and most HEV infection was subclinical.

Experimental African HEV infection in cynomolgus macaques (Macaca fascicularis)

Experimental infection with hepatitis E virus (HEV) from Africa has not been investigated. Our purpose was to study hepatitis E produced by HEV from Chad (North Africa) and to analyze the genetic sequence of the HEV obtained after animal passage. An HEV‐containing fecal sample from Chad was intravenously inoculated in four cynomolgus macaques. When serum Alanine Amino Transferase (ALT) levels rose, open liver biopsy and bile aspiration were performed. In all the monkeys, an ALT rise occured 25 to 32 days after inoculation and new anti‐HEV was detected by Enzyme Immuno Assay (EIA). Hepatic histopathology was consistent with acute viral hepatitis. HEV was detected by polymerase chain reaction (PCR) in bile (3/4 animals) and feces (2/4 animals) and by imunoelectron microscopy (IEM) in the inoculum and one bile specimen. A genetic variant HEV was identified in one monkey. The Chad HEV produced hepatitis E with pathophysiologic and histopathologic findings similar to those observed with HEV from other geographic origins. A genomic variant HEV population was produced after one passage in a macaque. J. Med. Virol. 55:197–202, 1998.