Effects of aging

In this section we investigate if BactoBox® enumerates bacteria in aged/senescent cultures.

Does BacTotal-10 count aged bacteria as cells?

It depends. Most aged cultures of Gram-negative bacteria are still found within the BacTotal-10 cell-definition. In contrast, some Gram-positive species may change electrical properties that differ from the BacTotal-10 cell-definition.

You can use custom gating to include the aged cell populations. This is an advanced feature. We only support advanced features through a custom solution. Contact support for details.

Aged cultures and BacTotal-10

The default BactoBox® configuration uses BacTotal-10. Objects classified as cells are found within the region with lavender shading. The distributions correspond to late-exponential phase (green) and aged (red) cultures.

Some aged cultures stay within the BacTotal-10 cell-definition. This is the case for Salmonella enteritidis (2 days), Escherichia coli (5 days), Lactobacillus rhamnosus (8 days), Lactobacillus casei (6 days), and Lactobacillus salivarius (4 days). The distributions shift left upon aging, but they are still within the BacTotal-10 cell-definition.

Examples of aged cultures that stay within the BacTotal-10 cell-definition.

Cultures of some aged bacterial species move outside the BacTotal-10 cell-definition. This is exemplified with Bifidobacterium bifidum (6 days), Bifidobacterium longum (3 days), Pediococcus acidilactici (6 days), Lactobacillus plantarum (5 days), and Staphylococcus epidermidis (2 days).

The aged cells of Gram-positive bacteria shift to the same conductive region as where we found the alcohol-treated cells. BacTotal-10 will not count these objects as cells.

Examples of aged cultures that move outside the BacTotal-10 cell-definition.

Why aged electrical signatures may differ between Gram-positive and Gram-negative bacteria

Autolysins are present in both Gram-negative and Gram-positive bacteria. These enzymes remodel the peptidoglycan cell wall during cell division and aging.

In Gram-positive cells the peptidoglycan cell wall is the only load-bearing structure. Nick it and the lipid membrane herniates through the breach and the membrane tears immediately. There is essentially no "damaged wall, intact membrane" intermediate state. The Gram-positive cells often become more conductive as they age.

Gram-negatives are different: the outer membrane plus a thin peptidoglycan layer means they can die non-lytically and persist for a long time. Some of the cytoplasm is lost during aging and the cell shrinks. Overall, the Gram-negative cells usually become less conductive during aging.

Illustration of the fates of Gram-positive and Gram-negative bacteria during aging. Gram-positive bacteria often become highly conductive, while the Gram-negative bacteria often become less conductive.

Summary

Cells in aged cultures may move outside the BacTotal-10 cell-definition. Custom gating may be required for BactoBox® to enumerate these objects.

Advanced feature 🧩

Custom gating is an advanced feature. We only support advanced features through a custom solution. Contact support for details.