Vibrio parahaemolyticus, enterotoxigenic Escherichia coli, enterohemorrhagic Escherichia coli and Vibrio cholerae

This review highlighted the following: (i) pathogenic mechanism of the thermostable direct hemolysin produced by Vibrio parahaemolyticus, especially on its cardiotoxicity, (ii) heat-labile and heat-stable enterotoxins produced by enterotoxigenic Escherichia coli, especially structure–activity relationship of heat-stable enterotoxin, (iii) RNA N-glycosidase activity of Vero toxins (VT1 and VT2) produced by enterohemorrhagic Escherichia coli O157:H7, (iv) discovery of Vibrio cholerae O139, (v) isolation of new variant of Vibrio cholerae O1 El Tor that carries classical ctxB, and production of high concentration of cholera toxin by these strains, and (vi) conversion of viable but nonculturable (VBNC) Vibrio cholerae to culturable state by co-culture with eukaryotic cells

Protein kinase C and clostridial neurotoxins affect discrete and related steps in the secretory pathway

1. The effects on catecholamine secretion of activation of protein kinase C and clostridial neurotoxins were examined in digitonin-permeabilized bovine adrenal chromaffin cells.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/44281/1/10571_2004_Article_BF00711564.pd

Expression of tetanus toxin subfragments in vitro and characterization of epitopes.

To define epitopes of tetanus toxin, we compared four different in vitro systems in terms of their ability to produce tetanus toxin-specific subfragments from cloned DNA. A transcription-translation system developed from a nontoxigenic strain of Clostridium tetani was found to yield predominantly full-sized peptides. Such peptides were used to map six different epitopes for eight monoclonal antibodies. The toxin-neutralizing properties of the antibodies were determined in an in vitro assay, based on the toxin-mediated inhibition of norepinephrine release from rat brain particles. Two monoclonal antibodies recognizing epitopes within the regions Ser-744 to Ser-864 and Ile-1224 to Asp-1315 could neutralize the toxin. A third nonneutralizing antibody was shown to recognize the synthetic peptide Phe-947 to Glu-967 derived from the tetanus toxin sequence. This peptide contains a human T-cell epitope

Evaluation of DNA probes for specific detection of Vibrio cholerae O139 Bengal.

Two DNA probes, 2R1 and 2R3, prepared from a region in the chromosome specific for the lipopolysaccharide O side chains of Vibrio cholerae O139 (M.K. Waldor and J.J. Mekalanos, Lancet 343:1366, 1994) were examined for their specificity and sensitivity. Both probes did not hybridize with any strain of V. cholerae belonging to serogroups other than O139 and to any of the other species examined belonging to the family Vibrionaceae. Among the 126 strains of V. cholerae O139 examined, probe 2R1 hybridized with 125 strains while probe 2R3 hybridized with all 126 strains. Both probes were found to be highly specific and sensitive and can be used for the specific identification of V. cholerae O139

Exogenous mRNA encoding tetanus or botulinum neurotoxins expressed in Aplysia neurons.

Injection of exogenous mRNA purified from various tissue preparations into cellular translation systems such as Xenopus oocytes has allowed expression of complex proteins (e.g., receptors for neurotransmitters). No evidence for expression of injected exogenous mRNA, however, has been reported in terminally differentiated neurons. If achieved, it would allow the study of long-lasting changes of properties of nerve cells in their functional context. To obtain evidence of such expression, we chose two proteins that produce a detectable effect even at very low intracellular concentrations. Tetanus toxin and botulinum neurotoxin fulfill this criterion, being the most potent neurotoxins known. Both toxins block neurotransmitter release at nanomolar intracellular concentrations. These di-chain proteins, consisting of a light chain and a heavy chain, have recently been sequenced. Their active sites are located (or partly located) on the light chain. mRNAs encoding the light chain of either toxin were transcribed in vitro from the cloned and specifically truncated genes of Clostridium tetani and Clostridium botulinum, respectively, and injected into presynaptic cholinergic neurons of the buccal ganglia of Aplysia californica. Depression of neurotransmitter release appeared in less than 1 hr, demonstrating successful expression of foreign mRNA injected into a neuron in situ

MOLECULAR-BIOLOGY OF CLOSTRIDIAL TOXINS - EXPRESSION OF MESSENGER-RNAS ENCODING TETANUS AND BOTULINUM NEUROTOXINS IN APLYSIA NEURONS

mRNAs encoding the light chain of tetanus and botulinum neurotoxins were transcribed, in vitro, from the cloned and specifically truncated genes of Clostridium tetani and Clostridium botulinum, respectively, and injected into presynaptic identified cholinergic neurons of the buccal ganglia of Aplysia californica. The size of the current response measured in the voltage clamped postsynaptic neuron was taken as indicator of the quantity of acetylcholine released. Depression of neurotransmitter release similar to that observed when native light chains of the two toxins were injected but needing an additional delay of 30 to 40 minutes, demonstrated a successful expression of a foreign mRNA injected into a neuron in situ

Characterization of phenotypic, serological, and toxigenic traits of Vibrio cholerae O139 bengal.

Biochemical and physiological traits of a collection of strains of Vibrio cholerae O139 Bengal isolated from India, Bangladesh, and Thailand showed that these strains formed a phenotypically homogeneous group with identical characteristics that were essentially similar to those of the O1 serogroup. Resistance to 150 micrograms of the vibriostatic agent O/129 (2,4-diamino-6,7-diisopropylpteridine) and Mukherjee's El Tor phage 5 and classical phage IV and the nonagglutinability of the strains with O1 antiserum were the only discernible differences between the O139 and O1 serogroups. Extensive serological characterization further revealed the O139 serogroup to be distinct from the existing 138 serogroups of V. cholerae. Antiserum raised against the O139 serogroup required absorption with the R reference strain CA385 and with the reference strain representing serogroup O22 to remove cross-reacting agglutinins. All of the 223 representative strains of V. cholerae O139 examined hybridized with DNA probes specific for the cholera toxin (CT) gene, zonula occludens toxin gene, and El Tor hemolysin gene but not with the probe specific for the heat-stable enterotoxin gene. The amount of CT present in stool samples of patients infected with the O139 serogroup was higher than that found in stools of patients infected with O1 El Tor, and this echoed findings that the amount of CT produced by O139 strains in vitro was higher than that produced by the O1 El Tor strains. The nucleotide sequences of the genes encoding the A and B subunits of CT of the O139 serogroup were identical to the sequences reported for the CT gene of O1 El Tor. The CT gene of O139 strains could be amplified by using primers developed for detection of the CT gene of the O1 serogroup by a PCR assay, which could also be used to detect the CT gene in stool samples of patients infected with strains of the O139 serogroup