Affiliation:
1. Department of Microbiology and Immunology, The University of Texas Medical Branch, Galveston, Texas 77555-1070
Abstract
ABSTRACT
Many gram-negative bacteria use a type III secretion system (TTSS) to deliver effector proteins into host cells. Here we report the characterization of a TTSS chromosomal operon from the diarrheal isolate SSU of
Aeromonas hydrophila
. We deleted the gene encoding
Aeromonas
outer membrane protein B (AopB), which is predicted to be involved in the formation of the TTSS translocon, from wild-type (WT)
A. hydrophila
as well as from a previously characterized cytotoxic enterotoxin gene (
act
)-minus strain of
A. hydrophila
, thus generating
aopB
and
act/aopB
isogenic mutants. The
act
gene encodes a type II-secreted cytotoxic enterotoxin (Act) that has hemolytic, cytotoxic, and enterotoxic activities and induces lethality in a mouse model. These isogenic mutants (
aopB
,
act
, and
act/aopB
) were highly attenuated in their ability to induce cytotoxicity in RAW 264.7 murine macrophages and HT-29 human colonic epithelial cells. The
act/aopB
mutant demonstrated the greatest reduction in cytotoxicity to cultured cells after 4 h of infection, as measured by the release of lactate dehydrogenase enzyme, and was avirulent in mice, with a 90% survival rate compared to that of animals infected with Act and AopB mutants, which caused 50 to 60% of the animals to die at a dose of three 50% lethal doses. In contrast, WT
A. hydrophila
killed 100% of the mice within 48 h. The effects of these mutations on cytotoxicity could be complemented with the native genes. Our studies further revealed that the production of lactones, which are involved in quorum sensing (QS), was decreased in the
act
(32%) and
aopB
(64%) mutants and was minimal (only 8%) in the
act/aopB
mutant, compared to that of WT
A. hydrophila
SSU. The effects of
act
and
aopB
gene deletions on lactone production could also be complemented with the native genes, indicating specific effects of Act and the TTSS on lactone production. Although recent studies with other bacteria have indicated TTSS regulation by QS, this is the first report describing a correlation between the TTSS and Act of
A. hydrophila
and the production of lactones.
Publisher
American Society for Microbiology
Subject
Infectious Diseases,Immunology,Microbiology,Parasitology
Cited by
84 articles.
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