Table 1.
M. marinum infection stages in D. discoideum and methods used for their analysis.
| Infection stage | Method used for analysis | D. discoideum strains/markers | M. marinum strains/markers | ||
|---|---|---|---|---|---|
| 1. Phagocytosis | FACS (Sattler et al., 2013) | wt | – | wt | pMSP12::GFP |
| IFA (Hagedorn et al., 2009) | wt | F-actin (phalloidin) | wt | pMSP12::GFP | |
| InfectChip (Delince et al., 2016) | wt | – | wt | pCherry10 | |
| Live microscopy (Hagedorn et al., 2009) | wt | GFP-ABD | wt | pR2Hyg | |
| TEM (Hagedorn et al., 2009) | wt | – | wt | – | |
| 2. Niche establishment | IFA (Hagedorn and Soldati, 2007; Kolonko et al., 2014) | wt, wshA- | ArpC4-GFP, calmodulin, cathepsin D, Coronin-GFP, F-actin, p80, vacuolins, VMC, VatA, GFP-WASH | wt, L1D | pCherry3, pMSP12::GFP |
| Live microscopy (Kolonko et al., 2014; Barisch et al., 2015a,b; Gerstenmaier et al., 2015; Barisch and Soldati, 2017b; Cardenal-Muñoz et al., 2017) | wt | ABD-GFP, AmtA-mCherry/GFP, DQ Green BSA, LysoSensor Green, NR, GFP-Rab5a, GFP-Rab7a, GFP-Rab11c, TRITC-dextran, VacA-GFP, VatB-RFP, VatM-GFP | wt, ΔRD1 | pCherry3/10, pMSP12::DsRed/GFP, Vibrant DyeCycle Ruby | |
| TEM (Cardenal-Muñoz et al., 2017) | wt | – | wt | – | |
| 3. MCV membrane damage | IFA (Cardenal-Muñoz et al., 2017) | wt, atg1- | Atg8a, GFP-p62, Ub (FK2) | wt, ΔCE | pCherry10 |
| Immunoblot (Cardenal-Muñoz et al., 2017) | wt | p-4E-BP1 (T70), Abp1, p-Raptor (S863) | wt, ΔRD1 | pCherry10, pMSP12::DsRed | |
| Live microscopy (Cardenal-Muñoz et al., 2017) | wt, atg1- | GFP-Atg8a, GFP-Atg18, DQ Green BSA, GFP-Ub | wt, ΔRD1, ΔRD1::RD1-2F9 | pCherry10, pMSP12::DsRed, Vibrant DyeCycle Ruby | |
| qPCR (Cardenal-Muñoz et al., 2017) | wt | atg1, atg8a, atg8b, p62 | wt, ΔRD1 | – | |
| 4. MCV membrane repair | IFA (Cardenal-Muñoz et al., 2017) | wt, atg1-, atg1- Atg1-GFP, atg8- | Ub (FK2) | wt | pCherry10 |
| Immunoblot (Cardenal-Muñoz et al., 2017) | wt | p-4E-BP1 (T70), Abp1, p-Raptor (S863) | wt, ΔRD1 | pCherry10, pMSP12::DsRed | |
| Live microscopy (Cardenal-Muñoz et al., 2017) | wt | GFP-Lamtor1, GFP-Lst8, GFP-Raptor, GFP-Rheb | wt, ΔRD1 | pCherry10, pMSP12::DsRed | |
| TEM (Cardenal-Muñoz et al., 2017) | wt, atg1- | – | wt | – | |
| 5. Bacteria replication within the MCV | FACS (Hagedorn and Soldati, 2007; Hagedorn et al., 2009; Lelong et al., 2011; Kolonko et al., 2014) | wt, kil2-, racH-, vacB-, wshA- | MB38::ESAT-6 | wt, L1D, ΔRD1 | pMSP12::GFP |
| IFA (Solomon et al., 2003; Hagedorn and Soldati, 2007; Lelong et al., 2011) | wt, kil2-, racH-, vacB- | vacuolins, coronin-GFP, p80 | wt, L1D | map24::GFP, pMSP12::GFP | |
| Luminescence recording in microplate reader (Ouertatani-Sakouhi et al., 2017) | wt | – | wt | pMV306::lux | |
| TEM (Hagedorn et al., 2009) | wt | – | wt | – | |
| 6. MCV rupture | IFA (Hagedorn and Soldati, 2007; Hagedorn et al., 2009; Lelong et al., 2011; Cardenal-Muñoz et al., 2017) | wt, kil2-, racH-, vacA-, vacB- | MB38::ESAT-6, p80, Ub (KF2), vacuolins | wt, ΔRD1 | pCherry10, pMSP12::GFP |
| Live microscopy (Barisch et al., 2015b) | wt | VacA-GFP | wt | pCherry10 | |
| TEM (Hagedorn et al., 2009) | wt, racH- | – | wt, ΔRD1 | – | |
| 7. Escape from the MCV | IFA (Hagedorn and Soldati, 2007; Hagedorn et al., 2009; Gerstenmaier et al., 2015; Cardenal-Muñoz et al., 2017) | wt, racH-, vacA- vacB- | Atg8a, F-actin (phalloidin), MB38::ESAT-6, GFP-p62/Sqstm1, p80, Ub (FK2), vacuolins | wt, ΔRD1 | pCherry10, pMSP12::GFP |
| Live microscopy (Barisch et al., 2015a,b; Barisch and Soldati, 2017b) | wt | AmtA-mCherry/GFP, RFP/GFP-Plin, VacA-GFP | wt, ΔRD1 | pCherry10 | |
| TEM (Hagedorn et al., 2009) | wt, racH- | – | wt, ΔRD1 | – | |
| 8. Cytosolic replication | Luminescence recording in microplate reader (Cardenal-Muñoz et al., 2017) | wt, atg1-, atg8a-, p62- | – | wt, ΔRD1 | pMV306::lux |
| 9.1. Egress by ejection | CLEM (Gerstenmaier et al., 2015) | wt | F-actin (Lifeact-GFP) | wt | pCherry3 |
| FACS (Gerstenmaier et al., 2015) | wt, atg1- | PI | wt | pMSP12::GFP | |
| IFA (Hagedorn et al., 2009; Kolonko et al., 2014; Gerstenmaier et al., 2015) | wt, atg1-, atg5-, atg6-, atg7-, p62/sqstm1-, racH-, vacA-, vacB-, wshA-, wshA- GFP-WASH | GFP-2xFYVE, Arp3, Atg8a, GFP-Atg18, Coronin, F-actin (phalloidin), GFP, MB38::ESAT-6, myoII, myoB, GFP-p62/Sqstm1, p80, PM4C4, Ub (FK2) | wt, ΔRD1 | anti-mar, pCherry3, pMSP12::GFP, pR2Hyg | |
| Live microscopy (Hagedorn et al., 2009; Gerstenmaier et al., 2015) | wt | ABD-GFP, GFP-ABD, DAPI, TRITC-dextran | wt | pMSP12::GFP, pR2Hyg | |
| TEM (Gerstenmaier et al., 2015) | wt, atg1- | – | wt | – | |
| 9.2. Egress by host cell death | FACS (Gerstenmaier et al., 2015) | wt, atg1- | PI | wt | pMSP12::GFP |
| InfectChip (Delince et al., 2016) | wt | – | wt | pCherry10 | |
| 9.3. Exocytosis | IFA (Hagedorn et al., 2009) | wt | GFP-ABD, F-actin (phalloidin), p80, PM4C4, vacuolin | wt | pMSP12::GFP |
| InfectChip (Delince et al., 2016) | wt | – | wt | pCherry10 | |
| Live microscopy (Gerstenmaier et al., 2015) | wt | ABD-GFP, TRITC-dextran | wt | pMSP12::GFP | |
| SEM (Hagedorn et al., 2009) | wt | – | wt | – | |
| TEM (Hagedorn et al., 2009) | wt | – | wt | – | |
| 10. Intercellular dissemination | Dissemination/Transmission assay [fixed fluorescence microscopy (Hagedorn et al., 2009) or FACS (Gerstenmaier et al., 2015)] | wt, atg1-, racH- | GFP-ABD, Lifeact-RFP | wt | pMSP12::GFP, pR2Hyg |
| IFA (Hagedorn et al., 2009) | wt | F-actin (phalloidin), p80 | wt | pMSP12::GFP | |