For any bioprocess where the product is expressed intracellularly, cell lysis is an essential unit operation to release the desired bioproduct into the extracellular space. Specifically in Gram-negative bacteria, there exists three barriers to the release of an intracellular bioproduct (Silhavy et al., 2010): the lipid cell membrane, the peptidoglycan cell wall, and the outer membrane; each of which must be disrupted to release bioproducts into the surrounding medium. It is worth noting that the addition of a secretion signal in Gram-negative bacteria can target proteins to the periplasmic space only (Karyolaimos and de Gier, 2021), and not the extracellular space.

Traditional cell lysis techniques can be grouped into 4 main groups: mechanical lysis, chemical lysis, heat lysis, and enzymatic lysis (Shehadul Islam et al., 2017) (Fig. 1). Mechanical lysis (e.g., homogenisation) can result in the creation of shear forces that may be damaging to the intracellular bioproduct (Carlson et al., 1995). This is typically not an issue with proteins, however there is increasing evidence that shear and interfacial stresses may contribute to loss of product quality due to protein aggregation (Moino et al., 2024). Diverse bioproducts are produced in engineered bacterial host cells, such as a wide range of different nucleic acids (Curry et al., 2024, Daròs, 2021, Godiska et al., 2010), biopolymers (Moradali and Rehm, 2020), and sometimes more sensitive proteins; mechanical lysis may be unsuitable for these applications. Shear forces from mechanical lysis may lead to the degradation of nucleic acids and biopolymers, making such disruption methods unsuitable for these products (Bhat et al., 2024, Carlson et al., 1995; Levy et al., 1999). Chemical lysis typically involves the addition of a chemical that can destabilize the different layers of the bacterial cell envelope, such as detergents or alkaline buffers (Shehadul Islam et al., 2017). However, similarly to mechanical lysis, chemical lysis can result in damage to the desired bioproduct (Clemson and Kelly, 2003). Thermal lysis is another common approach but is limited in scale due to heat transfer challenges when scaling up. Enzymatic lysis, although highly efficient, requires the costly procurement of purified lytic enzymes, therefore leading to cost limitations when scaling. Altogether, traditional cell lysis techniques suffer from two main drawbacks: harsh conditions leading to product degradation or lack of scalability.

Autolysis, defined as the intracellular expression of lytic proteins to trigger self-lysis of the host cell, has been an attractive alternative method for large scale intracellular bioproduct release. It addresses the challenges facing traditional cell lysis techniques and is a gentler approach for large scale bioproduct recovery. Employing autolysis in a bioprocess will allow for the integration of two steps into a single unit operation: fermentation and cell disruption. This reduction in unit operations may potentially allow for a bioprocess that is more cost-efficient and environmentally sustainable. As a technology, autolysis has existed for decades (Morita et al., 2001), and autolysis is utilised for a wide range of applications, including intracellular product release (Borrero-de Acuña et al., 2017), control of synthetic microbial consortia (Diao et al., 2021), production of bacterial ghost adjuvants (Ma et al., 2022), and targeted drug delivery (Din et al., 2016). However, a number of challenges have prevented the wide adoption of autolysis in bioprocessing. The engineering of autolytic bacterial strains and its broader significance have been thoroughly reviewed elsewhere (Dong et al., 2024). Therefore, this review will focus on the challenges preventing the wide adoption of inducible autolysis in large scale bioprocessing and explore solutions proposed in the literature.