The Conjugation Window in an Escherichia coli K-12 Strain with an IncFII Plasmid

Abstract

Many studies have examined the role that conjugation plays in disseminating antibiotic resistance genes in bacteria. However, relatively little research has quantitively examined and modeled the dynamics of conjugation under growing and non-growing conditions beyond a couple of hours. We therefore examined growing and non-growing cultures of E. coli over a 24 hour time period to try and understand the dynamics of bacterial conjugation in the presence and absence of antibiotics with pUUH239.2, an IncFII plasmid containing multi-antibiotic and metal resistant genes. Our data indicates that conjugation occurs after E. coli divide and before they have transitioned to a non-growing phase. The result is that there is only a small window of opportunity for E. coli to conjugate with pUUH239.2 under both growing and non-growing conditions. Only a very small percentage of the donor cells likely are capable of even undergoing conjugation and not all transconjugants can become donor cells due to molecular regulatory controls and not being in the correct growth phase. Once a growing culture enters stationary phase the number of capable donor cells decreases rapidly and conjugation slows to produce a plateau. Published models did not provide accurate descriptions of conjugation under non-growing conditions. A modified modeling approach is presented that accurately describes observed conjugation behavior under growing and non-growing conditions. Importance There has been growing interest in horizontal gene transfer of antibiotic resistant plasmids as the antibiotic resistance crisis has worsened over the years. Most studies examining conjugation of bacterial plasmids focus on growing cultures of bacteria for short time periods, but in the environment most bacteria grow episodically and at much lower rates than the laboratory. We examined conjugation of an IncFII antibiotic resistant plasmid in E. coli under growing and non-growing conditions to understand the dynamics of conjugation under which the plasmid is transferred. We found that conjugation occurs in a narrow time frame when E. coli is transitioning from a growing to non-growing phase and that the conjugation plateau develops because of a lack of capable donor cells in growing cultures. From an environmental aspect, our results suggest that episodic growth in nutrient depleted environments could result in more conjugation than sustained growth in a nutrient rich environment.

Conjugative plasmid transfer is limited by prophages but can be overcome by high conjugation rates

“…given that for phage infection and subsequent lysogenization to occur, prophages first have to be induced (which happens at a low spontaneous rate) and need to reach a high enough phage-to-bacteria ratio for lysogenization to take place at a significant frequency [7,8]. In agreement with our data however, it has been shown before that conjugation frequency for some plasmids increases only slowly over the exponential growth phase and peaks at the transition to stationary phase [16]. Another, mutually non-exclusive, explanation could be that recipients pay a plasmid acquisition cost (in addition to the plasmid metabolic burden), which slows down initial growth and spread of transconjugants [36].…”

“…Another unexpected observation in this one-donor experiment was the rapid emergence of transconjugants at the first measurement point (Figure 3B, 3C), which did not allow us to estimate the initial rate of increase in those experiments. This might be caused by induction of lysogens continuously killing a small part of the plasmid donor population, meaning that overnight cultures might undergo more turnover [35] and that a larger fraction of the population might still be capable of conjugation [16].…”

“…The plasmid can be transferred from the plasmid donor (DP) to recipient cells ( R ), phage donors ( DL ) or recipient lysogens ( RL ) with a certain conjugation rate ( p ), leading to recipient plasmid ( RP ), recipient lysogen plasmid ( RLP ) and phage donor plasmid ( DLP ) cells after τ p time units needed for transfer. Plasmids can be transferred from any plasmid-carrying population, but conjugation slows down when cells approach stationary phase [15, 16] - which is scaled via k p (Equ. 16), as our empirical data suggested that conjugation is not completely inhibited during stationary phase (see increase of transconjugants and double-carriers from T7 to T24 in Figure 1B).…”

Plasmid Transfer by Conjugation in Gram-Negative Bacteria: From the Cellular to the Community Level

“…One such regulation involves the silencing of P Y , P M , and P J promoters by the chromosome-encoded histone-like nucleoid structuring protein (H-NS) [ 18 , 19 ]. The H-NS copy number per cell varies during growth [ 20 ], thus rendering the F plasmid transfer rate growth phase-dependent, i.e., maximum in the exponential phase, reduced in the mid-exponential phase, and mostly abolished in the stationary phase [ 21 , 22 ]. However, during the exponential phase, H-NS repression activity is itself counteracted by the cooperative binding of TraJ and the host protein ArcA (aerobic respiration control of anoxic redox control) to the P Y promoter [ 23 ].…”

Bacteria DNA Conjugation: from The Cellular to The Community Level

“…One such regulation involves the silencing of PY, PM, and PJ promoters by the chromosome encoded histone-like nucleoid structuring protein H-NS [18,19]. H-NS copy number per cell varies during growth [20], thus rendering F plasmid transfer rate growth-phase-dependent, i.e., maximum in exponential, reduced in mid-exponential and mostly abolished in stationary phase [21,22]. Yet, during the exponential phase, H-NS repression activity is itself counteracted by the cooperative binding of TraJ and the host protein ArcA (Aerobic respiration control of anoxic redox control) to the PY promoter [23].…”