Trypanosoma brucei gambiense Translationnally Controlled Tumour Protein interacts with Glossina midgut bacteria

Trypanosoma brucei gambiense

Translationally Controlled Tumour Protein

interacts with Glossina midgut bacteria

S.Ravel

African trypanosomiasis

actors

S.Ravel

Domestic / Wild animals and Human

 Molecular dialogue between host and parasite

 Parasite destabilize environment to survive inside tse tse fly by secreted protein (secretome)

 Circulating inside tse tse fly (midgut, gut)

 During a host blood meal infected by trypanosoma

Parasite’s strategy

Approach

 Identified secretome’s protein contained in vesicule

 Selected proteins and study them as therapeutic and/or diagnostic target

Translationally Controled Tumour Protein (TCTP)

Histidine

GST TCTP

Check list from secretome’s protein

Produced in baculovirus system Purified by affinity colunm

1.

2.

Isolated from 39 bacteria species

from

Glossina

midguts

Physical interaction test

50 KDa

Western blot : HRP antihistidine monoclonal

Tbg TCTP control

No interaction Interaction

Results

Interaction : 24 bacteria species

No interaction : 15 bacteria species

3.

Concentration of

Tbg

TCTP (1.6 µg to 200 µg) Reading 0h, 24h, 48h, 72h, 96h Temperature 25°C and 37°C Atmosphere aerobiosis microaerobiosis anaerobiosis

Impact of Tbg TCTP on the bacteria growth

Conclusion

Global results

• 25°C and 37°C bacteria have a similar growth with any

Tbg

TCTP concentration

Results at 25°C

• In majority no effect regardless atmostphere excepted to few bacteria species at three

concentration of Tbg TCTP 0 500 1000 1500 sans TCTP TCTP 200 TCTP 100 TCTP 50 TCTP 25 TCTP 12,5 TCTP 6,25 TCTP 3,15 TCTP 1,6 0 100 200 300 400 500 600 700 800 no TCTP TCTP 200 TCTP 100 TCTP 50 TCTP 25 TCTP 12,5 TCTP 6,25 TCTP 3,15 TCTP 1,6

Parameters

Temperature effect 25°C cam 33C 24h 0 50 100 150 200 sans TCTP TCTP 200 TCTP 100 TCTP 50 TCTP 25 TCTP 12,5 TCTP 6,25 TCTP 3,15 TCTP 1,6

Inhibition effect independant time,

atmosphere, and Tbg TCTP dose Activation modulating

Identification of Tbg TCTP in vesicules Production and purification

Physical interaction with number of Glossina bacteria species

Impact of bacteria growth is highly disturbed regardless the concentration of Tbg TCTP in vitro In vivo, in Glossina, this bacteriome modulated could be associated to vector competence

cam 32C 72h

Géraldine Bossard, CIRAD, UMR INTERTRYP, F-34398 Montpellier, France

Manon Bartoli, IRD-MIO, F-13009 Marseille, France

Fardeau Marie Laure, IRD-MIO, F-13009 Marseille, France Olivier Bernard, IRD-MIO, F-13009 Marseille, France

Holzmuller Philippe, CIRAD, UMR ASTRE, F-34398 Montpellier, France Anne Geiger, IRD, UMR INTERTRYP, F-34398 Montpellier, France

Contact : geraldine.bossard@cirad.fr

Website : https://umr-intertryp.cirad.fr

Trypanosome Tse tse fly

Coomassie _{Western blot}

Relativie activity Relative activity Relative activity 50 60 70 80 90 100 110 120 130 140 150 24h 48h 72h 96h Gmm1 Ang24C 0 500 1000 1500 2000 2500 24h 48h 72h 96h Cam15B

Relative activity _{Relative activity}

Characterization of recombinant Trypanosoma brucei gambiense

Translationally Controlled Tumor Protein (rTbgTCTP) and its interaction with Glossina midgut bacteria.

Bossard G, Bartoli M, Fardeau ML, Holzmuller P, Ollivier B, Geiger A.

Gut Microbes. 2017 Sep 3;8(5):413-427. doi:

10.1080/19490976.2017.1331833. Epub 2017 Jun 6.

Results at 37°C

Methods

Bacteria + Tbg TCTP 2 hours at 4°C, 3 washes in PBS, and loading in acrylamide gel follow

transfer on a nitrocellulose membrane,

revealing a with HRP antihistidine monoclonal

IAD -T our s Mar s 2 01 8 TCTP concentration TCTP concentration Time Time TCTP concentration No No

Selection filtrer : no signal peptide Physical transporter : microvesicule