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The viable but nonculturable (VBNC) state has been recognized as a strategy for bacteria to cope with stressful environments; in this state, bacteria fail to grow on routine culture medium but are actually alive and can resuscitate into a culturable state under favorable conditions. The VBNC state may pose a great threat to food safety and public health. To date, more than 100 VBNC microorganism species have been proven to exist in fields of food safety, environmental application, and agricultural diseases. Most harsh conditions can induce these microorganisms into the VBNC state, including food processing and preservation methods, adverse environmental conditions, and plant‐disease controlling means. The characteristics of VBNC state cells differ from those of normally growing cells and dead cells, based on which of the various detection methods are developed, and they are of great significance for potential risk assessment. To provide molecular level insights into this state, many studies on induction and resuscitation mechanisms have emerged over the past three decades, including research on omics, specific genes, or proteins involved in VBNC state formation and the roles of promoters in resuscitation from the VBNC state. In this review, microorganism species, induction and resuscitation factors, detection methods, and formation and resuscitation mechanisms of the VBNC state are comprehensively and systematically summarized.

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ABSTRACTrRNAs of dormant spores of Bacillus subtilis were >95% degraded during extended incubation at 50°C, as reported previously (E. Segev, Y. Smith, and S. Ben-Yehuda, Cell 148:139 -114, 2012, doi:http://dx.doi.org/10.1016/j.cell.2011.11.059), and this was also true of spores of Bacillus megaterium. Incubation of spores of these two species for ϳ20 h at 75 to 80°C also resulted in the degradation of all or the great majority of the 23S and 16S rRNAs, although this rRNA degradation was slower than nonenzymatic hydrolysis of purified rRNAs at these temperatures. This rRNA degradation at high temperature generated almost exclusively oligonucleotides with minimal levels of mononucleotides. RNase Y, suggested to be involved in rRNA hydrolysis during B. subtilis spore incubation at 50°C, did not play a role in B. subtilis spore rRNA breakdown at 80°C. Twenty hours of incubation of Bacillus spores at 70°C also decreased the already minimal levels of ATP in dormant spores 10-to 30-fold, to <0.01% of the total free adenine nucleotide levels. Spores depleted of rRNA were viable and germinated relatively normally, often even faster than starting spores. Their return to vegetative growth was also similar to that of untreated spores for B. megaterium spores and slower for heat-treated B. subtilis spores; accumulation of rRNA took place only after completion of spore germination. These findings thus strongly suggest that protein synthesis is not essential for Bacillus spore germination. Spores of Bacillus species formed in sporulation are metabolically dormant and extremely resistant to many severe treatments, including heat, radiation, and toxic chemicals (1-3). Although these spores can survive in this dormant resistant state for many years, if given the appropriate stimulus, generally small molecules that are found in environments that are appropriate for the growth of these bacteria, the spores can rapidly break dormancy in the process of spore germination and then outgrowth (1, 4, 5). Notably, spores' extreme resistance properties are lost in the process of germination and early outgrowth, and thus, germinated spores are much easier to kill than dormant spores. This property has significant applied implications, as dormant spores are the vectors for much food spoilage, as well as some foodborne and other human diseases. Consequently, there is significant interest in mechanisms that determine rates of spore germination and how sporulation conditions, as well as storage conditions, modify spores' resistance properties and their rates of germination.In addition to the applied interest in spore resistance and germination, there has also long been much basic interest in the mechanism of spore germination, in particular, the signal transduction pathways operating during the process (4, 5). The major proteins involved in Bacillus spore germination are as follows. (i) Germinant receptors (GRs) in spores' inner membrane (IM) recognize and respond to the physiological germinants specific for spores of each species/strain. (ii)...

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The Gram positive bacterium Bacillus subtilis and its relatives are capable of forming a durable dormant long-lasting spore. Although spores can remain dormant for years, they possess the remarkable capacity to rapidly resume life and convert into actively growing cells. This cellular transition initiates with a most enigmatic irreversible event, termed germination, lasting only for a few minutes. Germination is typified by a morphological conversion that culminates in loss of spore resilient properties. Yet, the molecular events occurring during this brief critical phase are largely unknown. The current widely accepted view considers germination to occur without the need for any macromolecule synthesis; however, accumulating data from our laboratory and others, highlighted here, provide evidence that both transcription and translation occur during germination and are required for its execution. We further underline numerous overlooked studies, conducted mainly during the 1960s-70s, reinforcing this notion. We propose to revisit the fascinating process of spore germination and redefine it as a pathway involving macromolecule synthesis. We expect our perspective to shed new light on the awakening process of a variety of spore-forming environmental, commensal and pathogenic bacteria and possibly be applicable to additional organisms displaying a quiescent life form.