Research ArticleLIFE SCIENCES

Bacteria: A novel source for potent mosquito feeding-deterrents

See allHide authors and affiliations

Science Advances  16 Jan 2019:
Vol. 5, no. 1, eaau6141
DOI: 10.1126/sciadv.aau6141
  • Fig. 1 Description of the membrane feeding and feeding-deterrent screening system.

    (A) Components of the feeding system, including (top/bottom panel, left to right) Hemotek temperature controller, feeder-housing assembly, metal feeder assembled with cocktail diet (red color) secured with collagen casing and O-ring, two layers of cotton cloth, metal feeder with cotton cloth secured via rubber bands (not visible), and metal feeder assembly secured to feeder housing. ID, inside diameter. Photo credit: Mayur Kumar Kajla, University of Wisconsin–Madison. (B and C) Results of the feeding assays. (B) Plot showing the number of A. aegypti mosquitoes that fed when water was applied to the cotton cloth as control, or DEET or picaridin (0.95 mg/cm2) [equivalent to 1.0% (v/v)] was applied in three replicate experiments. Each replicate consisted of 20 mosquitoes for each of the control as well as tests. (C) Representative image depicting appearance of fed (red abdomens) versus unfed Aedes mosquitoes resulting from the bioassay. Images show engorged abdomens and red dye in fed mosquitoes. Absence of both color and engorgement of abdomens indicates that no feeding occurred with 1% DEET (or picaridin) as a positive control.

  • Fig. 2 Mosquito feeding-deterrent activity of Xbu Peak#3 with C. pipiens, A. gambiae, and A. aegypti.

    (A) Plots show feeding-deterrent activity of Xbu compounds tested at 0.057 mg/cm2. Data from three replicate experiments are shown. Each replicate consisted of 20 mosquitoes for each of the control (water only) as well as tests (Xbu Peak#3), respectively. A. aegypti mosquitoes were included for comparison. (B) Appearance of fed (red abdomens) and unfed Anopheles and Culex mosquitoes. In this experiment, one-layer muslin cloth was used. Fisher’s exact test was used to assess differences between groups using Stata statistical software.

  • Fig. 3 MALDI-TOF spectrum of flash and HPLC C18 reversed-phase chromatography separated mosquito feeding-deterrent active peak fraction.

    (A) MALDI-TOF analysis of the C18 flash fractionated feeding-deterrent active fraction yielded several major masses (and related molecular species) at m/z 1302.94 (1346.97), 1312.96 (1356.98), and 1430.07 (1474.10), as well as m/z 1238.953. The mass difference of m/z 44 in these pairs may be due to addition of C2H4O moieties (11). (B) MALDI-TOF analysis of HPLC-separated feeding-deterrent active Xbu Peak#3 shows enrichment of the same two abundant (and related) molecular species at m/z 1302.92 and 1346.95.

  • Table 1 Determination of feeding-deterrence dose (50 and 90%) of DEET, picaridin, and Xbu compounds against A. aegypti.

    RE90 is the relative efficacy of Xbu Peak#3 derived as a ratio of the dose causing 90% reduction in mosquito feeding of DEET or picaridin to that of Xbu Peak#3.

    CompoundSlope ± SE50% Feeding-
    deterrence dose
    (mg/cm2)
    95% Fiducial limits90% Feeding-
    deterrence dose
    (mg/cm2)
    95% Fiducial limitsRE90
    DEET1.09 ± 0.360.0120.000–0.0320.1780.066–6.7851
    Picaridin1.73 ± 0.350.0860.044–0.1560.4710.237–2.0030.38
    Xbu Peak#32.08 ± 0.360.0140.010–0.0200.0570.035–0.1498.26
  • Table 2 Comparison of mosquito feeding-deterrent activity of DEET, picaridin, and Xbu Peak#3 against A. aegypti.

    Table shows the Probit-analyzed comparison among the least-squares estimates for each compound’s effect (32). Feeding-deterrence comparison based on adjusted P values among DEET and picaridin was significant (adjusted P < 0.05); DEET and Xbu Peak#3 were not significant (adjusted P > 0.05); and Xbu Peak#3 was significantly different from picaridin (adjusted P < 0.05).

    Differences of compound least-squares means adjustment for multiple comparisons: Tukey-Kramer
    CompoundCompoundEstimateSEz valuePr > |z|Adj P
    DEETPicaridin1.0910.2943.710.0000.000
    DEETXbu Peak#3−0.1730.276−0.620.5320.806
    PicaridinXbu Peak#3−1.2640.309−4.08<0.0000.000

Supplementary Materials

  • Supplementary material for this article is available at http://advances.sciencemag.org/cgi/content/full/5/1/eaau6141/DC1

    Fig. S1. Enrichment of the mosquito feeding-deterrent active compounds via flash chromatography on reversed-phase C18 column.

    Fig. S2. Enrichment of the mosquito feeding-deterrent active compounds via HPLC on an analytical reversed-phase C18 column.

    Fig. S3. Elution profile of feeding-deterrent active fraction.

    Fig. S4. MALDI-TOF MS analysis of HPLC-enriched feeding-deterrent Peak#3.

    Fig. S5. Electrospray ionization–Orbitrap MS analysis of the HPLC-enriched deterrent active Xbu Peak#3 showing observed charged states.

    Fig. S6. MS/MS spectra of doubly-charged molecular species at m/z 651.9 and 673.9.

    Fig. S7. Amino acid analyses of purified feeding-deterrent fraction.

    Fig. S8. Appearance of cotton cloths used in the Hemotek-based membrane-feeding assays.

    Fig. S9. Typical example of Xbu streaked on LBTA (Luria broth agar supplemented with indicator stains bromothymol blue and triphenyltetrazolium chloride) plate.

    Table S1. Mosquito feeding data used for Probit analysis to generate Tables 1 and 2.

    Table S2. Results of the N-terminal Edman sequencing trial.

    Movie S1. Video clip showing feeding response of mosquitoes with water control and at a 90% feeding-deterrence dose of Xbu Peak#3.

  • Supplementary Materials

    The PDF file includes:

    • Fig. S1. Enrichment of the mosquito feeding-deterrent active compounds via flash chromatography on reversed-phase C18 column.
    • Fig. S2. Enrichment of the mosquito feeding-deterrent active compounds via HPLC on an analytical reversed-phase C18 column.
    • Fig. S3. Elution profile of feeding-deterrent active fraction.
    • Fig. S4. MALDI-TOF MS analysis of HPLC-enriched feeding-deterrent Peak#3.
    • Fig. S5. Electrospray ionization–Orbitrap MS analysis of the HPLC-enriched deterrent active Xbu Peak#3 showing observed charged states.
    • Fig. S6. MS/MS spectra of doubly-charged molecular species at m/z 651.9 and 673.9.
    • Fig. S7. Amino acid analyses of purified feeding-deterrent fraction.
    • Fig. S8. Appearance of cotton cloths used in the Hemotek-based membrane-feeding assays.
    • Fig. S9. Typical example of Xbu streaked on LBTA (Luria broth agar supplemented with indicator stains bromothymol blue and triphenyltetrazolium chloride) plate.
    • Table S1. Mosquito feeding data used for Probit analysis to generate Tables 1 and 2.
    • Table S2. Results of the N-terminal Edman sequencing trial.
    • Legend for movie S1

    Download PDF

    Other Supplementary Material for this manuscript includes the following:

    • Movie S1 (.mp4 format). Video clip showing feeding response of mosquitoes with water control and at a 90% feeding-deterrence dose of Xbu Peak#3.

    Files in this Data Supplement:

Navigate This Article