Potential Regrowth and the Role of Viable but Nonculturable Bacteria

As practitioners in the compost industry, we assume that when the temperature of our compost has reached 55 C, we have killed all potential pathogens. There are many studies that demonstrate the effect of temperature on the viability of potential pathogens. When we find elevated fecal coliform or E. coli after the high temperature phase, we assume that its either regrowth, or false positives due to “other” bacteria. However, the elevated fecal coliform following high temperature composting may be due to Viable but Nonculturable Bacteria (VBNC). Sunar et al. (2009) observed that high temperatures during composting killed E. coli and Salmonella as measured using traditional plating methods, but found that using polymerase chain reaction (PCR) methodology indicated that the E. coli had survived.

There are some excellent review articles on VBNC (Fakruddin et al. 2013, Li et al. 2014, Ramamurthy et al. 2014, Pinto et. al. 2015, Zhao et al. 2017). The concept was first reported in 1982 (Xu et al. 1982). Li et al. (2014) described as bacteria reducing their function as a survival mechanism allowing them to wait for suitable conditions to revive. and also described that bacteria are not able to survive the VBNC state for extended periods of time. Ramamurthy et al. (2014) described how the resuscitation process of VBNC bacteria required favorable growth conditions with a source of energy, which we observe during composting when the temperature decreases following the thermophilic phase.

While we would like to think that its only the non-pathogenic bacteria that can survive high temperatures, its not true.  Li et al. (2014) reported that 51 potential human pathogens are able to enter the VBNC state including those commonly found in our composting processes.

Some excellent work was done on the topic of potential pathogen kill and viable but nonculturable bacteria during composting by the Engineering group at the University of Alberta (Isobaev 2014). They concluded that:

“Gradual exposure to TTC [time temperature criteria] induces a VBNC state in E. coli and Salmonella. The VBNC state helps both E. coli and Salmonella survive at appreciable concentration throughout the 56 days long composting cycle. With certain constraints the VBNC at the early state in E. coli and Salmonella can be reverted when optimum growth conditions are supplied”.

“it is not recommended to view the temperature as an effective stand-alone sanitation factor. According to the collected evidences, pathogens like E. coli and Salmonella can survive thermophilic conditions, similar to those in the composting pile. The cells, when exposed to 55°C for more than 3 consecutive days can induce stress-response mechanism and subsequently transit into VBNC state. During direct process validation the organisms in VBNC successfully skip culture-based detection methods and pose the risk to regrow during storage and transportation. The stakeholders should always keep that in mind when distributing the product. At least the existing direct process validation methods should be amended to incorporate the pathogens in VBNC.”

The humbling news is that potentially pathogenic organisms may survive the high temperature phase of composting.

The good news is that potentially pathogenic bacteria that enter the VBNC state are only able to remain in that state for a limited length of time. Other studies have confirmed that when the compost matures, there is no readily available carbon left for the potentially pathogenic organisms, and they are no longer able to resuscitate. This confirms that when we follow adequate composting and curing procedures, we are destroying potential pathogens.

References

Fakruddin, M., K.S.B. Mannan and S. Andrews. 2013. Viable but nonculturable bacteria: food safety and public health perspective. ISRN Microbiology. http://dx.doi.org/10.1155/2013/703813.

Isobaev, P. 2014. Developing and Testing a Framework to Measure the Sanitation Efficacy on a Random Particle Level in the Composting Industry. PhD Thesis. Department of Civil and Environmental Engineering, University of Alberta.

Li, L., N. Mendis, H. Trigui, J.D. Oliver and S.P. Faucher. 2014. The importance of the viable but non-culturable state in human bacterial pathogens. Frontiers in Microbiology. Doi 10.3389/fmicb.2014.00258

Pinto, D. V., M.A. Santos and I. Chambel.  2015. Thirty years of viable but nonculturable state research: unsolved molecular mechanisms. Critical Reviews of Microbiology 41: 61-76. review

Ramamurthy, T., A. Ghosh, G.P. Pazhani and S. Shinoda. 2014. Current perspectives on viable but non-culturable (VBNC) pathogenic bacteria. Frontiers in Public Health. Doi: 10.3389/pubh.2014.00103. review

Sunar, N.M., D.I. Stewart, E.I Stentiford, and L.A. Fletcher. 2009. A rapid molecular approach to determine the occurrence of pathogen indicators in compost. Proceedings of the Twelfth International Waste Management and Landfill. Sardinia 2009 Symposium.

Xu, H.S., N. Roberts, F.I Singleton, R.W. Attwell, D.J. Grimes and R.R. Colwell. 1982. Survival and viability of nonculturable Escherichia coli and Vibrio cholerae in the estuarine and marine environment. Microbial Ecology 8: 313-323.

Zhao, X. J. Zhong, C Wei, C-W Lin and T. Ding. 2017. Current perspectives on viable but non-culturable state in food-borne pathogens. Frontiers in Microbiology doi: 10.3389/fmicb.2017.00580

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