Papaya (Carica papaya L.) cultivars and breeding lines were evaluated for resistance to Enterobacter cloacae (Jordan) Hormaeche & Edwards, the bacterial causal agent of internal yellowing disease (IY), using a range of concentrations of the bacterium. Linear regression analysis was performed and IY incidence was positively correlated with increasing inoculum concentrations for susceptible cultivars Kapoho Solo and Laie Gold but not for resistant cultivars or lines. It was determined that the inoculum concentration of 9 to 10 Log10 colony-forming units per milliliter (cfu/mL) was able to reliably differentiate resistant and susceptible papaya germplasm. Red-fleshed cultivars SunUp and Sunrise were the most resistant papaya groups evaluated at this dose concentration. Yellow-fleshed cultivars, Kapoho Solo and Laie Gold, were susceptible to E. cloacae. ‘Rainbow’, an F1 hybrid between ‘SunUp’ and ‘Kapoho Solo’ that is yellow-fleshed, was moderately resistant to E. cloacae, exhibiting limited symptoms of the disease. Yellow-fleshed I-Rb F5/F6, an advanced inbred line derived from ‘Rainbow’, is resistant and offers the potential of improving resistance of yellow-fleshed commercial cultivars. A colorimeter was used to objectively measure internal flesh color and distinguish between infected and non-infected tissue in red- and yellow-fleshed papayas using L*C*H* color space analysis. Symptomatic tissue (72.4 and 79.0°) had higher hue angle means than non-symptomatic tissue (62.8 and 75.0°) for all cultivars or lines in red- and yellow-fleshed papayas, respectively. Yellow (“Y”) hue color also distinguished infected tissue from non-infected tissue. Symptomatic tissue that had Y hue color resulted in 79 to 81° hue angle means among red- or yellow-fleshed papayas. Our results demonstrated the usefulness of colorimetry to help detect infected papaya tissue. In surveys of naturally infected papaya, high populations (8.57 × 107 cfu/g) of E. cloacae were recovered in infected fruit of ‘Kapoho Solo’ and represent a food safety concern for fresh and processed papaya. In isolations from inoculated fruits, we observed decreases of ≈1 to 2 Log10 cfu/g in final bacterial populations when high-dose range inoculum concentrations (9 to 12 Log10 cfu/mL) were used. This dose range may represent a saturation range for E. cloacae inoculation.