Search PubMed⌕ Search

Biomedical subjects

M M Levine

Publications and source records attributed to M M Levine.

At least 127 records · Page 7Linked to original sources
PubMed1994

Longus: a long pilus ultrastructure produced by human enterotoxigenic Escherichia coli.

Enterotoxigenic Escherichia coli (ETEC) causes an acute cholera-like diarrhoea in both humans and animals. We describe a new pilus termed longus produced by ETEC, which can extend for over 20 microns from the cell surface. Longus is composed of a repeating subunit of 22 kDa and its NH2-terminal amino acid sequence revealed homology with the toxin-coregulated pilus of Vibrio cholerae, the bundle-forming pilus of enteropathogenic E. coli and type IV pilins of some Gram-negative bacterial pathogens. The longus structural gene (lngA) is encoded in a large plasmid and was cloned in a 5 kb fragment, which proved to be sufficient for pilus production and assembly in E. coli K-12. The presence of lngA was restricted to human ETEC strains. In contrast to other ETEC pili, lngA was widely distributed among ETEC strains independent of their geographical origin, serotype, toxin production, or other pili antigens expressed. Longus is a new member of the type IV pili family, which may represent a highly conserved intestinal colonization factor of ETEC. Common antigenic determinants exist among longus and their pilin subunits, produced by heterologous ETEC. Longus could be significant in the immunoprophylaxis of diarrhoeal disease caused by ETEC, especially against those strains in which no colonization factors have been identified and that produce heat-stable toxin only.

Amino Acid Sequence↗
PubMed1994

Construction and characterization of attenuated delta aroA delta virG Shigella flexneri 2a strain CVD 1203, a prototype live oral vaccine.

We engineered an oral Shigella vaccine prototype that can invade intestinal epithelial cells but cannot undergo extensive intracellular replication or extend to adjacent epithelial cells. Strain CVD 1203, derived from wild-type Shigella flexneri 2a by introducing deletions in chromosomal aroA and invasion plasmid virG, was highly attenuated in the Sereny test. Two 10(9)-CFU orogastric doses (2 weeks apart) stimulated production of secretory immunoglobulin A antibodies to S. flexneri 2a and protected against conjunctival sac challenge with virulent S. flexneri 2a.

Administration, Oral↗
PubMed1994

Potential for reacquisition of cholera enterotoxin genes by attenuated Vibrio cholerae vaccine strain CVD 103-HgR.

The potential for reacquisition of ctxA genes by attenuated Vibrio cholerae O1 vaccine strain CVD 103-HgR was examined by performing a series of mating experiments under a variety of in vivo and in vitro conditions. We found no evidence that CVD 103-HgR could reacquire ctxA genes from wild-type V. cholerae O1 strains. However, if the donor V. cholerae O1 strains were genetically manipulated to add genes that allow chromosomal gene transfer, then ctxA sequences could be acquired by CVD 103-HgR. The minimal excretion of CVD 103-HgR by vaccinees and the refractoriness to reacquisition of ctxA sequences suggest that this well-tolerated, highly immunogenic live oral cholera vaccine will have a minimal environmental impact.

Cholera Vaccines↗
PubMed1994

Vaccination against typhoid fever: present status.

Typhoid fever remains an underestimated important health problem in many developing countries, causing more than 600,000 deaths annually in the world. Because of the reactogenicity of the parenteral, killed whole-cell vaccine, research has been oriented towards vaccination orally using live organisms and purified antigen. Live vaccine Ty21a, given by the oral route, has been extensively tested in several studies in developing countries. Its liquid formulation was the most effective, providing more than 60% protection after 7 years of follow-up. A Vi polysaccharide vaccine has been elaborated and provided more than 65% protection; after 3 years of follow-up the Vi antibody level was still at a high level. These two vaccines are therefore candidates for use in public health control programmes. Before such use, however, they need further evaluation for safety and protective efficacy when administered to the EPI-targeted age groups. The question of whether typhoid fever vaccines interfere with the response to simultaneously administered measles vaccine must also be studied. New live vaccines, given by the oral route in one dose, have been constructed through genetic engineering. The first results are promising, but they must be improved before use in a large-scale study. These strains could be used as live vector to deliver foreign antigens to the intestinal mucosa.

Administration, Oral↗
PubMed1994

Recombinant Salmonella vectors in vaccine development.

A diversity of means are available for the attenuation of Salmonella which can be used to immunize animals and humans orally to elicit mucosal, humoral and cellular immune responses. Avirulent Salmonellae can be genetically engineered to express foreign antigens and the recombinant avirulent Salmonellae are capable of stable, high-level expression of the foreign antigen in the orally immunized animal or human host. The resulting vaccines are safe, efficacious, and are easy and economical to use.

Administration, Oral↗
PubMed1993

Immunologic response to oral cholera vaccination in a crossover study: a novel placebo effect.

The authors conducted a two-period crossover study of the reactogenicity and immunogenicity of live oral cholera vaccine CVD 103-HgR among US university students. Subjects ingested 5 x 10(8) colony forming units of either killed Escherichia coli K12 placebo or vaccine, followed by the opposite treatment one week later. Surprisingly, the dynamics of the immunologic response were influenced by prior ingestion of placebo. Subjects who received placebo first showed stronger vibriocidal antibody responses 2 weeks after vaccination compared with subjects who received vaccine first; this same pattern was seen for antitoxin titers. The authors suggest that ingestion of E. coli K12 one week prior to immunization boosts the immunologic response to vaccine by an unknown mechanism. Future crossover studies that examine immunologic outcomes might be designed to explore the ubiquity of such an effect.

Administration, Oral↗
PubMed1993

Epidemiologic studies of Escherichia coli diarrheal infections in a low socioeconomic level peri-urban community in Santiago, Chile.

The incidence of diarrhea due to six categories of diarrheogenic Escherichia coli was determined in two pediatric cohorts in a low socioeconomic level community in Santiago, Chile, with access to chlorinated water. An age cross-sectional cohort of 340 children aged birth to 47 months was assembled. A newborn cohort was assembled by enrolling 10-12 newborns monthly for 12 months. Episodes of diarrhea were detected by twice weekly household visits. E. coli from stool cultures of cases and matched controls were hybridized with DNA probes specific for enterotoxigenic, enteroinvasive, enteropathogenic, enterohemorrhagic, enteroaggregative, and diffuse adherence E. coli. Overall, the incidence of diarrhea was low (2.1 episodes/infant/year). Nevertheless, a putative E. coli enteropathogen was found in a large proportion of diarrheal episodes, particularly during the summer. In both cohorts, enterotoxigenic E. coli were important pathogens. Enteropathogenic E. coli were incriminated during the first year of life in the newborn cohort, where they were found significantly more often in cases (p = 0.021) than in controls; beyond this age, isolation rates were similar. In contrast, the relative risk of isolation of diffuse adherence E. coli increased with age in the age cross-sectional cohort, where, overall, the difference in rate of isolation between cases and controls was significant (p = 0.0024). Enteroinvasive and enterohemorrhagic E. coli were isolated infrequently. Enteroaggregative E. coli were encountered equally in cases and controls. Facile transmission of E. coli enteropathogens is occurring in this community despite the availability of potable water.

Case-Control Studies↗
PubMed1993

Construction of genetically marked Vibrio cholerae O1 vaccine strains.

Attenuated Vibrio cholerae O1 vaccine strains lacking the gene encoding the A subunit of cholera toxin have proven efficacious in preventing experimental cholera. As these strains move from closed, contained testing environments to large-scale field trials, a readily assayable phenotypic trait to distinguish a vaccine strain from wild-type V. cholerae O1 is desirable. We have constructed three derivatives of the attenuated V. cholerae strain CVD 103 which carry a mercury resistance or urease marker in the hlyA gene. CVD 103-HgR was constructed using a protracted marker-exchange procedure; this strain was found to have somewhat lowered colonisation efficiency in infant mice in comparison to its parent strain, CVD 103. The insertion of the resistance marker was repeated using a suicide vector system; CVD 103-HgR2 was found to colonise infant mice as efficiently as CVD 103. Strain CVD 103-UR, in which sequences encoding urease were inserted using a suicide vector, also colonised infant mice as well as CVD 103. The genetically marked strains CVD 103-HgR, CVD 103-HgR2 and CVD 103-UR form the basis for a generation of defined oral vaccines that may give single-dose, long-lasting protection to populations at risk from cholera.

Animals↗
PubMed1993

Structural studies of the O-antigenic polysaccharide of an enteroaggregative Escherichia coli strain.

The polysaccharide part of the lipopolysaccharide obtained from an enteroaggregative Escherichia coli strain isolated from a young child with diarrhoea in Santiago, Chile (strain 73-1) was investigated. Sugar and methylation analyses of native and partially degraded polysaccharide together with 1H-NMR and 13C-NMR spectroscopy revealed that the polysaccharide is built of pentasaccharide repeating units. The structure of the repeating unit of E. coli strain 73-1 O-polysaccharide is (formula: see text)

Carbohydrate Conformation↗
PubMed1993

Enteroaggregative Escherichia coli heat-stable enterotoxin 1 represents another subfamily of E. coli heat-stable toxin.

Enteroaggregative Escherichia coli (EAggEC) are associated with persistent diarrhea in young children. Some of these organisms produce a low-molecular-weight, heat-stable, plasmid-encoded enterotoxin that has been named EAggEC heat-stable enterotoxin 1 (EAST1). We have cloned a 4.4-kb DNA fragment from the virulence plasmid of prototype EAggEC strain 17-2, which expresses enterotoxic activity as measured by electrogenic response in Ussing chambers mounted with rabbit ileal tissue. DNA-sequence analysis of this fragment identified an open reading frame (ORF) encoding a cysteine-rich polypeptide of 38 amino acids (M(r), 4100). Insertional and deletional mutations in this ORF resulted in loss of enterotoxic activity. The ORF was cloned into a T7 expression vector, and postinduction culture filtrates exhibited enterotoxic activity and increased ileal tissue cGMP levels. A synthetic peptide consisting of predicted amino acid residues 8-29 also showed enterotoxic activity. These data indicate that this ORF, named astA (EAggEC heat-stable enterotoxin), represents the EAST1 structural gene. EAST1 shows significant homology with the enterotoxic domain of heat-stable enterotoxin a (STa) of enterotoxigenic E. coli and with guanylin, a mammalian analog of STa. Unlike STa, which requires six cysteines and three disulfide linkages for full biological activity, both EAST1 and guanylin contain four cysteine residues. Based on the cGMP data and the sequence homology to STa and guanylin, it is predicted that EAST1 stimulates the particulate form of guanylate cyclase through the same receptor-binding region as STa and guanylin.

Amino Acid Sequence↗
PubMed1993

Immunization of rabbits with enterotoxigenic E. coli colonization factor antigen (CFA/I) encapsulated in biodegradable microspheres of poly (lactide-co-glycolide).

We have searched for an effective oral delivery system for a purified enterotoxigenic Escherichia coli (ETEC) fimbrial adhesin, CFA/I, which elicits anti-colonization immunity. Purified CFA/I antigen encapsulated in biodegradable polymer microspheres of poly (DL-lactide-co-glycolide) (PLG) induced a vigorous, systemic CFA/I IgG antibody response in rabbits immunized once via intragastric tube; oral, unencapsulated CFA/I induced little or no circulating antibody. CFA/I-specific, S-IgA coproantibody was detected in one of three rabbits fed with the CFA/I-PLG microsphere vaccine. We conclude that PLG microspheres protected CFA/I from degradation in the stomach and effectively delivered the antigen for processing by the host's immune system.

Adhesins, Escherichia coli↗
PubMed1993

Live oral vaccines against cholera: an update.

One hundred years elapsed between the first (live, parenteral) cholera vaccine that entered clinical trials in 1885 and the field trials of two oral inactivated cholera vaccines undertaken in Bangladesh in the mid-1980s. The oral inactivated vaccines advanced the art by establishing, convincingly, that oral vaccines could protect (although multiple doses were required) and that (at least in adults) protection could last 3 years. Attenuated Vibrio cholerae O1 strain CVD 103-HgR (deleted of the cholera toxin A subunit gene and harbouring a gene encoding resistance to Hg++) constitutes another significant advance. This live oral vaccine is well tolerated and highly immunogenic in adults and children and highly protective (in adult volunteer challenge studies) following ingestion of of a single dose.

Administration, Oral↗
PubMed1993

Vaccines to prevent enteric infections.

Considerable progress has been made in the last decade in developing vaccines against the enteric infections of greatest public health importance. A quadrivalent rotavirus vaccine consisting of rhesus rotavirus vaccine (which contains serotype 3 neutralization antigen) and three reassortant viruses (rhesus virus expressing neutralization antigens of serotypes 1, 2 or 4) is undergoing placebo-controlled field trials of efficacy in the USA and in two developing countries. Two new vaccines against typhoid fever (oral Ty21a and parenteral Vi polysaccharide) have been licensed in many countries. Even newer generations of typhoid vaccines are undergoing clinical testing, including new attenuated S. typhi strains and Vi polysaccharide-carrier protein conjugate vaccines. Two inactivated oral cholera vaccines, consisting of inactivated V. cholerae O1 bacteria alone or in combination with purified B subunit of cholera toxin, each conferred 50-53% protection over 3 years in a field trial in Bangladesh where subjects were immunized with a three-dose regimen. In extensive clinical trials in adults and children in less-developed countries, an engineered live oral cholera vaccine, strain CVD 103-HgR, has been shown to be well tolerated and highly immunogenic following administration of just a single oral dose; a large-scale field trial in 70,000 subjects is underway to investigate the efficacy of this vaccine. Several candidate vaccines against Shigella and enterotoxigenic Escherichia coli are in clinical trials. Accumulating knowledge on pathogenesis of enteric infections and advances in mucosal and cellular immunology, coupled with the application of modern biotechnology, have resulted in a plethora of vaccine candidates. It is expected that in future years efforts will be directed to construct vaccines against other enteric pathogens.

Gastroenteritis↗
PubMed1993

Relationship between humoral immune responses and oral temperature during human shigellosis.

The relationship between oral temperature and serum IgA and IgG antibody responses to Shigella sonnei infection was examined in 65 experimentally infected adult volunteers. Although the IgA antibody response appeared to correlate directly with the rise in oral temperature following infection, stepwise regression analysis revealed other associated signs and symptoms as the critical elements underlying this apparent correlation. There was no evidence of a positive correlation between the IgG antibody response and the peak increment in oral temperature after infection. Although published data obtained in vitro suggest that fever might have a potentiating effect on antibody production, no such effect was seen in human volunteers with acute shigellosis.

Adult↗
PubMed1993

Safety and immunogenicity of live oral cholera vaccine candidate CVD 110, a delta ctxA delta zot delta ace derivative of El Tor Ogawa Vibrio cholerae.

The current pandemic of cholera is caused primarily by Vibrio cholerae O1 of the El Tor biotype. Live attenuated classical biotype V. cholerae vaccine strains prevent severe and moderate cholera due to either biotype in challenged volunteers but may provide less protection against mild cholera due to El Tor organisms. CVD 110, a new ctxA-deleted vaccine strain derived from an El Tor Ogawa parent, lacks zona occludens toxin (Zot), accessory cholera enterotoxin (Ace), and hemolysin/enterotoxin. Ten healthy adult volunteers were given 10(8) cfu of CVD 110 with buffer; 7 developed diarrhea (mean stool volume, 861 mL). Vaccine organisms were shed in stool by all vaccines and were recovered from duodenal fluid in three-quarters of vaccinees. After vaccination, the geometric mean peak reciprocal vibriocidal titer among vaccinees was 17,829. CVD 110 is a powerful immunogen but insufficiently attenuated despite the absence of known potential enterotoxins of V. cholerae. Another unrecognized toxin or colonization alone may be responsible for diarrhea after ingestion of this strain.

Administration, Oral↗
PubMed1993

Secondary Vibrio cholerae-specific cellular antibody responses following wild-type homologous challenge in people vaccinated with CVD 103-HgR live oral cholera vaccine: changes with time and lack of correlation with protection.

Peripheral blood immunoglobulin A antibody-secreting-cell (ASC) responses are thought to reflect the mucosal immune response to locally presented antigens. We evaluated the ASC response to cholera toxin (CT) and Inaba lipopolysaccharide (LPS) in 26 North American volunteers following immunization with a single oral dose of live attenuated Vibrio cholerae O1 vaccine strain CVD 103-HgR and again upon homologous wild-type challenge with V. cholerae classical Inaba 569B. Challenge occurred at either 7, 30, or 180 days after vaccination. The CT and LPS ASC responses of volunteers following vaccination (83 and 55%, respectively) were similar in magnitude and frequency to those of unvaccinated controls following wild-type challenge (80 and 60%, respectively [0.1 < or = P < or = 0.9]). The responses were primarily immunoglobulin A. Vaccinated volunteers challenged within 30 days of vaccination had reduced or nondetectable CT and LPS ASC responses. Challenge at 6 months resulted in a heightened ASC response to LPS, confirming the existence of mucosal memory. ASC responses to CT upon challenge at 6 months were detectable but not different from that seen following primary immunization, suggesting that secondary ASC responses to different antigens from a single vaccine operate independently. In spite of these variable ASC responses, the vaccine efficacy was 100% following challenge for all vaccinees. V. cholerae-specific ASC responses following antigenic reexposure gave information on the presence of mucosal B memory cells but did not correlate with protective immunity. As such, these ASC assays will have limited usefulness for evaluating vaccine responders in vaccine field trials in cholera-endemic areas where prior V. cholerae O1 exposure is unknown.

Administration, Oral↗
PubMed1993

Safety and immunogenicity of Salmonella typhi Ty21a vaccine in young Thai children.

Salmonella typhi Ty21a vaccine in a liquid formulation was evaluated in 634 Thai children 2 to 6 years of age. The seroconversion rate was 69% for those who received vaccine versus 14% for those who received placebo (P < 0.005). The immune responses among vaccine recipients ranged from 60% in 3-year-olds to 81% for 6-year-olds.

Age Factors↗