Inhibition of the early stage of Salmonella enterica serovar Enteritidis biofilm development on stainless steel by cell-free supernatant of a Hafnia alvei culture

Abstract

Compounds present in Hafnia alvei cell-free culture supernatant cumulatively negatively influence the early stage of biofilm development by Salmonella enterica serovar Enteritidis on stainless steel while they also reduce the overall metabolic activity of S. Enteritidis planktonic cells. Although acylhomoserine lactones (AHLs) were detected among these compounds, the use of several synthetic AHLs was not able to affect the initial stage of biofilm formation by this pathogen.

FIG. 1.

(A) Strongly attached/biofilm S. Enteritidis cells (log CFU cm⁻²) on SS coupons. Coupons were incubated at 18°C for up to 72 h in growth media containing 0% (□), 20% (▪), or 50% (▤) H. alvei CFS. (B) Conductance detection times (DTs; h) corresponding to data presented in panel A. The bars represent the mean values ± standard deviations. Mean values sharing at least one common number shown above the bars are not significantly different at a P value of <0.05. Within a group with the same incubation period (12 to 72 h; x axis), mean values sharing at least one common letter are not significantly different at a P value of <0.05.

FIG. 2.

Typical changes for broth conductance (μS) versus time (h). Each Malthus tube contained an SS coupon which had previously been incubated for 12 h in growth medium containing 0% (⧫), 20% (⋄), or 50% (×) H. alvei CFS. Sigmoidal curves represent the fitting of the three data series with the model of Baranyi and Roberts (R² > 0.99). The detection times (DTs; h) calculated by the model are also depicted. Similar DTs were recorded for 0 and 20% CFS (DT1); they were shorter than the DT recorded in the case of 50% CFS (DT2).

FIG. 3.

Linear regression plots between biofilm counts (log CFU cm⁻²) and corresponding conductance detection times (DTs; h). For biofilm formation, SS coupons were incubated at 18°C for up to 72 h in growth media containing 0% (⧫), 20% (⋄), or 50% (×) H. alvei CFS. Data points represent mean values. For greater clarity, error bars are not included. The mathematical equations of the three regression plots, together with their relevant regression coefficients (R²), are also shown.

FIG. 4.

Characteristic TLC profiles of AHLs in extracts from the three growth media at 12 h of incubation. Chromatographs were developed using the reporter strain A. tumefaciens A136. Lanes: 1, C6-HSL standard; 2, 0% CFS; 3, 50% CFS; 4, 20% CFS.