An anti-HIV microbicide engineered in commensal bacteria: secretion of HIV-1 fusion inhibitors by lactobacilli.
OBJECTIVE
To engineer Lactobacillus spp. to secrete HIV-1 fusion inhibitors with potent neutralizing activity against primary HIV-1 isolates.


METHODS
HIV-1 fusion inhibitors (FI-1, FI-2, and FI-3) were introduced into the previously developed shuttle vector pTSV2 and transformed in L. plantarum and L. gasseri. The signal peptide Usp45 from L. lactis was used to achieve high secretion efficiency of peptides into the bacterial supernatant. The antiviral activity of lactobacillus-derived HIV-1 fusion inhibitors was tested against a panel of primary HIV-1 isolates and a chimeric simian/HIV (SHIV-162P3) using the TZM infection assay. TZM-bl cells are engineered HeLa cells that express CD4, CCR5, and CXCR4 and contain integrated reporter genes for firefly luciferase and beta-galactosidase under the control of an HIV-1 long terminal repeat. The amount of secreted fusion inhibitor FI-3 was determined by Western blot analysis and the antiviral specificity verified by antibody-mediated depletion of peptide FI-3 and HIV-1 infection with VSV-G envelope pseudotyped virions.


RESULTS
Viral infectivity of primary HIV-1 isolates and SHIV-162P3 was neutralized by up to 98% and 72%, respectively, by 10% (v/v) lactobacillus supernatant containing fusion inhibitor FI-3. The antiviral activity of the lactobacillus-derived fusion inhibitor FI-3 was clearly shown to be attributable to the secreted fusion inhibitor peptide.


CONCLUSIONS
The development of recombinant lactobacilli expressing HIV-1 fusion inhibitors with potent neutralizing activity represents an important step toward the development of a live microbial (topical) microbicide against HIV-1 transmission.
Pusch
Oliver
O
Center of Anatomy and Cell Biology, Medical University of Vienna, Vienna, Austria.
Kalyanaraman
Roopa
R
Tucker
Lynne D
LD
Wells
Jerry M
JM
Ramratnam
Bharat
B
Boden
Daniel
D
D000818:Animals; D019380:Anti-HIV Agents; D019149:Bioreactors; D015153:Blotting, Western; D023581:HIV Fusion Inhibitors; D015658:HIV Infections / Q000188:drug therapy; D015497:HIV-1 / Q000187:drug effects; D006367:HeLa Cells; D006801:Humans; D007778:Lactobacillus / Q000502:physiology*; D048191:Lactiplantibacillus plantarum / Q000378:metabolism; D009500:Neutralization Tests; D030781:Organisms, Genetically Modified; D011994:Recombinant Proteins / Q000378:metabolism; D015302:Simian Immunodeficiency Virus / Q000187:drug effects; D014161:Transduction, Genetic / Q000379:methods
D019380:Anti-HIV Agents; D023581:HIV Fusion Inhibitors; D011994:Recombinant Proteins