High prevalence of Helicobacter pylori in the Alaska native population and association with low serum ferritin levels in young adults.

Iron deficiency anemia is a common public health problem in the Alaska Native population. Yet, a clear etiology has eluded researchers for decades. Previous studies suggested a link between Helicobacter pylori infection, gastrointestinal blood loss due to hemorrhagic gastritis, and generalized iron deficiency anemia in adult Alaska Natives. Therefore, we examined the association between the prevalence of H. pylori-specific immunoglobulin G (IgG) and serum ferritin levels, a marker of iron deficiency. A random sample of 2,080 serum samples from Alaska Native residents drawn between 1980 and 1986 from residents in 13 regions was selected, and the samples were stratified by age, sex, and region. Overall, 75% were positive for H. pylori-specific IgG. The rate of H. pylori seropositivity increased with age; by age 14 years, 78% of the residents were positive. There were no gender differences in H. pylori seropositivity. However, marked regional differences were observed. Serum ferritin levels of <12 ng/ml were found most commonly among persons <20 years of age and among women of childbearing age. A significant association between low serum ferritin levels and prevalence of H. pylori-specific IgG was found, particularly for people aged less than 20 years. H. pylori may be a factor contributing to the iron deficiency anemia in the Alaska Native population.

Seroprevalence of Helicobacter pylori infection in cystic fibrosis and its cross-reactivity with anti-pseudomonas antibodies.

BACKGROUND: The prevalence of Helicobacter pylori infection and its role in gastroduodenal disease in cystic fibrosis (CF) are controversial. Additionally, serologic determination of infection in this population may be inaccurate because of cross-reactivity with other bacterial species. The seroprevalence of H. pylori in a cohort of patients with CF and its cross-reactivity with Pseudomonas antibodies were investigated. METHODS: A research enzyme-linked immunosorbent assay (ELISA), and three commercial serologic assays (PyloriStat; BioWhittaker, Walkersville, MD, U.S.A.; Flexsure; SmithKline Diagnostics, Inc., San Jose, CA, U.S.A.; and HM-CAP; EPI, Stony Brook, NY, U.S.A.) at three independent laboratories determined the seroprevalence of anti-H. pylori IgG antibodies in 70 patients with CF. Cross-reactivity between solid-phase H. pylori antigens and Pseudomonas antibodies was ascertained by a competitive inhibition assay, preadsorbing sera of patients with CF with whole cell proteins from different Pseudomonas species, and serum reanalysis by each assay. Western blot analysis before and after adsorption was performed to identify potential cross-reactive antigens. RESULTS: The research ELISA, Flexsure, Pyloristat, and HM-CAP initially showed H. pylori seropositivity of 47%, 28%, 24%, and 37%, respectively. Postadsorption seropositivity declined to 8%, 0%, 0%, and 15%, respectively. All patients with research ELISA true-positive results were confirmed endoscopically to have H. pylori infection. Western blot analysis showed a 31-kDa H. pylori protein with antigenic epitopes common to both bacterial species. CONCLUSIONS: Cross-reactivity between solid-phase H. pylori antigens and anti-Pseudomonas antibodies occurs in patients with CF. A high index of suspicion should be assumed in evaluating results of serologic H. pylori tests in this population. Preadsorption of CF sera with Pseudomonas proteins should be used in serologic testing.

Excretion of hepatitis A virus (HAV) in adults: comparison of immunologic and molecular detection methods and relationship between HAV positivity and infectivity in tamarins.

Fecal excretion of hepatitis A virus (HAV) in 18 patients with HAV infection was evaluated by enzyme immunoassay (EIA) to detect viral antigen and by reverse transcription-PCR amplification followed by ethidium bromide staining (PCR-ETBr) or nucleic acid hybridization (PCR-NA) to detect viral genetic material. A gradation of sensitivity was observed in the detection of virus by the three methods. In persons who had detectable virus, serial stool samples were found to be positive by EIA for up to 24 days after the peak elevation of liver enzymes. Viral genetic material could be detected by PCR-ETBr for up to 34 days and by PCR-NA for up to 54 days after the peak elevation of liver enzymes. After intravenous inoculation of tamarins with stool suspensions categorized as highly reactive for HAV (positive by EIA, PCR-ETBr, and PCR-NA), moderately reactive (positive by PCR-ETBr and PCR-NA), or weakly reactive (positive by PCR-NA), only tamarins infected with highly reactive stool suspensions (EIA positive) developed HAV infection. We conclude that positivity of stool specimens for HAV by PCR-ETBr or PCR-NA indicates a lower potential for infectivity, compared to that of EIA-positive stools.

Use caution with serologic testing for Helicobacter pylori infection in children.

Commercial serologic assays accurately detect adult Helicobacter pylori infection. Their use in children remains controversial. An ELISA to detect H. pylori IgG in children was developed and compared with three commercial assays. ELISA standardization was done with sera from all ages and validation was done with another cohort of sera with known H. pylori status. Three commercial serologic assays were subsequently compared against this pediatric ELISA at independent sites, at which 142 pediatric serum samples from different countries were evaluated. The pediatric ELISA was 91.4% sensitive. Assay 3 demonstrated a sensitivity of 78%. Less sensitivity was observed for assay 1 (70%) and assay 2 (63%). Accuracy of commercial assays was greatly reduced when sera from developing countries and younger ages were evaluated. Results of serologic tests used to diagnose H. pylori should be interpreted with caution when evaluating children with abdominal pain. Accurate serologic assays in children may be more important for epidemiologic research than for clinical decision making.

Helicobacter pylori acquisition in infancy after decline of maternal passive immunity.

We evaluated the natural history of Helicobacter pylori infection and the host immune response in 80 infants, and determined seroprevalence of H. pylori infection in their Taiwanese mothers. Decline in passively transferred maternal anti-H. pylori IgG antibodies and subsequent H. pylori infection was assessed in infants over 14 mo. A sensitive and specific, 96-well microtiter ELISA for the detection of H. pylori IgG antibodies was used to evaluate maternal serum (single specimen) and their infants (birth, 1, 2, 3, 6, 12, and 14 mo). Sera were also evaluated by ELISA for the presence of anti-H. pylori IgM antibodies in the infants. Maternal H. pylori IgG seroprevalence was 62.5% [50/80; 95% confidence intervals (CI), 51-73%]. All infants born to the 50 seropositive mothers passively acquired maternal H. pylori IgG. Transplacentally transferred maternal anti-H. pylori IgG lasted until about the 3rd mo of life, and disappeared in nearly all the infants by 6 mo of age. Seven and one-half percent of infants (6/80; 95% CI, 3-16%) acquired H. pylori infection; two were born to H. pylori-negative mothers. Among the six IgG seropositive infants, an IgM response specific for H. pylori antigens was detected and appeared to precede the rise in IgG in five. We conclude that maternal passive transfer of IgG antibodies occurs in the infant and disappears by 6 mo of age. H. pylori infection is acquired in infancy in this population; IgM antibodies against H. pylori are detectable, seem short-lived, and appear to precede IgG antibody development.

Hepatitis A post-viral encephalitis.

We report a seven-year-old girl who developed a hepatitis A viral infection and encephalitis. The patient developed fever, abdominal pains and jaundice. Five days later she became delirious, combative, and did not respond to verbal commands. Laboratory studies showed elevated liver enzymes and elevated serum immunoglobulin M (IgM) and immunoglobulin G (IgG) antibodies to hepatitis A virus. Cerebrospinal fluid contained IgG antibodies to hepatitis A virus but not IgM antibodies. Polymerase chain reaction, which amplifies a portion of the hepatitis A virus genome, did not demonstrate viral nucleic acid in cerebrospinal fluid. These studies suggest that the patient may have suffered from a post-viral hepatitis A encephalitis from which she fully recovered.

Multistate outbreak of hepatitis A associated with frozen strawberries.

A multistate outbreak of hepatitis A was traced to frozen strawberries processed at a single plant. Among 827 students and 60 teachers at an elementary school in Georgia during a 2-week period, 15 developed hepatitis A. Three months later, among 174 residents and 467 staff in an institution for the developmentally disabled in Montana during a 3-week period, 13 developed hepatitis A. Primary attack rates were 10% in the school and 8% in the institution. Cohort analysis in the school implicated consumption of strawberry shortcake in hepatitis A virus (HAV) infection (relative risk, 7.6; 95% confidence interval, 1.04-55.6). In the institution, such analysis implicated desserts and uncooked strawberries as the most biologically plausible vehicle of HAV transmission. Molecular analysis of HAV from patients in the two outbreaks revealed that the viral genomes were genetically identical and distinct from other known US strains. Contamination of food products before retail distribution is rare but should be considered in investigating common-source outbreaks of hepatitis A.

Genetic relatedness of hepatitis A virus strains recovered from different geographical regions.

A pairwise comparison of the nucleic acid sequence of 168 bases from 152 wild-type or unique cell culture-adapted strains of hepatitis A virus (HAV) revealed that HAV strains can be differentiated genetically into seven unique genotypes (I to VII). In general, the nucleotide sequence of viruses in different genotypes differs at 15 to 25% of positions within this segment of the genome. Viruses from four of the genotypes (I, II, III and VII) were recovered from cases of hepatitis A in humans, whereas viruses from the other three genotypes (IV, V and VI) were isolated only from simian species developing a hepatitis A-like illness during captivity. Among non-epidemiologically related human HAV strains, 81 were characterized as genotype I, and 19 as genotype III. Within each of these major genotypes, there were two distinct groups (subgenotypes), which differed in sequence at approximately 7.5% of base positions. Each genotype and subgenotype has a characteristic amino acid sequence in this region of the polyprotein, with the most divergent genotypes differing at 10 of 56 residues. Strains recovered from some geographical regions belonged to a common (endemic) genotype, whereas strains from other regions belonged to several, probably imported, genotypes. Thus, HAV strains recovered in North America were for the most part closely related at the nucleotide sequence level, whereas in other regions, such as Japan and Western Europe, HAV strains were derived from multiple genotypes or sub-genotypes. These data indicate that patterns of endemic transmission can be differentiated from situations in which infections are imported due to travel.

Characterization of a genetic variant of human hepatitis A virus.

Human isolates of hepatitis A (HAV) are a single serotype; however, recent genetic surveys using limited nucleotide sequencing have provided evidence that more than one genotype is responsible for HAV infection in different parts of the world (Jansen et al. [1990]: Proc Natl Acad Sci USA 87:2867-2871; Robertson et al. [1991] J Infect Dis 163:286-292). One of these genotypes was originally isolated from Panamanian owl monkeys (strain PA21), but has subsequently been found associated with human cases of HAV from Sweden in 1979 (H-122) and the United States of America in 1976 (GA76). The nucleic acid sequence of the exposed capsid polypeptide region of GA76 differs from other human HAV sequences by approximately 20%, yet differs by only 2.4% when compared with P1 sequence of the PA21 strain. The 20% nucleic acid variability between GA76 and other human HAV results in limited amino acid changes (3%), while a comparison with PA21 revealed only four homologous amino acid substitutions within VP2, VP3, and VP1 polypeptides. HAV infected stool specimens from Nepal and northern India during 1989 and 1990 were found to contain virus whose genetic makeup was related to the PA21 and GA76 isolates. This genotype of HAV appears to be circulating in some parts of the world where HAV is hyperendemic, and is a potential cause of hepatitis A infection within a susceptible population.

Epidemiologic patterns of wild-type hepatitis A virus determined by genetic variation.

Hepatitis A virus (HAV) isolates from different parts of the world are a single serotype. However, genetic analysis of the VP1 genome region of published HAV sequences suggested that distinct genotypes of HAV could be defined based upon the geographic source of the original isolates. To circumvent the process of cell culture adaptation or animal passage, a 247-bp segment within the VP1 genome region of wild-type HAV was amplified by reverse transcription followed by polymerase chain reaction amplification in the presence of negative- and positive-sense primers. From the sequences obtained from 22 epidemiologically distinct HAV isolates, three genetic groups of HAV could be delineated. Two of the groups differed by 10%, while the third group differed from other isolates by approximately 20%. These investigations indicate that HAV isolates from different parts of the world can be differentiated genetically, which will facilitate studies of epidemiologic transmission.

Altered hepatitis A VP1 protein resulting from cell culture propagation of virus.

The published sequence of hepatitis A virus (HAV), strain HAS-15, after 20-30 cell culture passages contains an 18 nucleotide deletion (Ovchinnikov et al., 1985) within the VP1 genome region. This results in a significant amino acid difference of the VP1 protein when this strain of HAV is compared with other published HAV sequences. Comparison of the polyacrylamide gel electrophoretic migration of HAS-15 HAV and two other strains of HAV revealed that the HAS-15 VP1 molecule migrated faster than the VP1 molecule of the other two strains. Enzymatic amplification of viral RNA derived from the original stool suspension and cell culture adapted HAS-15 using the polymerase chain reaction followed by hybridization analyses with selected synthetic oligonucleotide probes revealed that the original wild type virus did not contain the deletion. These results confirm that cell culture adapted HAS-15 contains an eighteen nucleotide deletion which apparently was selected during cell culture adaptation.

Structure of the hepatitis A virion: identification of potential surface-exposed regions.

Iodination of highly purified hepatitis A (HAV) virus results in the selective labeling of two viral polypeptides, which are identified as the the VP 1 and VP 2 capsid polypeptides. Based upon the kinetics of labeling, the exposed region of VP 1 appears to be more accessible to iodination, although the ultimate proportion of label present within VP 1 and VP 2 is approximately equal. By utilizing iodinated whole virions, isolated VP 1, VP 2, and the tryptic digest derived from VP 1 and VP 2, binding by heterologous anti-160 S antibody indicated that a significant portion of the antibodies was directed against an epitope on VP 2 that was not affected by denaturation. Identification of the regions exposed for iodination on these two polypeptides was accomplished by tryptic digestion of the isolated polypeptides followed by characterization of the iodinated tryptic peptide by gel filtration and reverse-phase chromatography. The results indicate that tyrosine 100 on VP 2 and a large tryptic peptide composed of amino acids 222 through 260 on VP 1 which contains four tyrosine residues are two regions that are surface-exposed on these molecules.

An International Outbreak of Salmonella Nima from Imported Chocolate.

Twenty-nine confirmed cases of Salmonella nima enterocolitis in Canada and four cases in the United States were traced to gold-foil wrapped chocolate coins from Belgium. Illness in Canadian cases occurred between September 1985 and October 1986 where the primary case in each of 24 affected families was a child ≤ 14 years of age. A product recall was issued on October 3, 1986 in Canada. Quantitative analysis of four composite samples of suspect chocolate by the most probable number (MPN) technique showed levels of 4.3 to 24.0 S. nima per 100 g product. These levels of contamination and consumption of approximately 25 g of chocolate by primary cases suggest that small numbers of S. nima precipitated clinical symptoms.

Large scale production of hepatitis A virus in cell culture: effect of type of infection on virus yield and cell integrity.

Approaches to cell culture propagation of hepatitis A virus (HAV) have used either acute infection by passage of infected cell lysates or supernatants into uninfected cells or the passage of persistently infected cells. The findings presented here demonstrate that the growth and recovery of purified virus from foetal rhesus monkey kidney (FRhK4) cells persistently infected with HAV isolate HAS-15 decreased over a 2 to 3 month period. In contrast, high multiplicity acute infection of FRhK4 cells with purified HAS-15 HAV resulted in degeneration of the cell monolayer 2 to 3 weeks later. Large scale propagation of acutely infected cells followed by traditional picornavirus purification procedures reproducibly yielded milligram amounts of purified virus.

Ethambutol-induced hyperuricaemia.

A prospective study involving 134 cases of pulmonary tuberculosis was conducted to determine the incidence and severity of ethambutol-induced hyperuricaemia. The cases were randomly allocated to two groups: one group (71 cases) received ethambutol (20 mg/kg bodyweight/day) streptomycin and isoniazid (SHE), the other group (60 cases) received streptomycin, isoniazid and thioacetazone (SHT). All the cases were hospitalized. A significant rise in serum uric acid levels was found in 66% of SHE patients during the first 60-90 days of treatment, but there was no such change in the SHT patients. In one patient who received ethambutol generalized arthralgia developed and in another acute gouty arthritis was observed. In both cases, symptoms abated when ethambutol was withdrawn and reappeared when it was resumed.