Tag Archives: Salmonella Typhimurium

Study highlights two strategies used by Salmonella to escape the human body’s defenses

Like thieves that constantly look for ways to evade capture, Salmonella enterica, a disease-causing bacterium, uses various tactics to escape the human body’s defense mechanisms. In a new study, researchers from the Department of Microbiology and Cell Biology (MCB), IISc, highlight two such strategies that the bacterium uses to protect itself, both driven by the same protein.

When Salmonella enters the human body, each bacterial cell resides within a bubble-like structure known as Salmonella-containing vacuole (SCV). In response to the bacterial infection, the immune cells in our body produce reactive oxygen species (ROS) and reactive nitrogen species (RNS), along with pathways triggered to break down these SCVs and fuse them with cellular bodies called lysosomes or autophagosomes, which destroy the bacteria. However, these bacteria have developed robust mechanisms to maintain vacuolar integrity, which is crucial for their survival. For example, when a bacterial cell divides, the vacuole surrounding it also divides, enabling every new bacterial cell to be ensconced in a vacuole. This also ensures that more vacuoles are present than the number of lysosomes which can digest them.

In the study published in Microbes and Infection, the IISc team deduced that a critical protein produced by Salmonella, known as SopB, prevents both the fusion of SCV with lysosomes as well as the production of lysosomes, in a two-pronged approach to protect the bacterium. “[This] gives the upper hand to bacteria to survive inside macrophages or other host cells,” explains Ritika Chatterjee, former PhD student in MCB and first author of the study. The experiments were carried out on immune cell lines and immune cells extracted from mice models.

SopB acts as a phosphatase – it aids in removing phosphate groups from phosphoinositide, a type of membrane lipid. SopB helps Salmonella change the dynamics of the vacuole – specifically it alters the type of inositol phosphates in the vacuole membrane – which prevents the vacuole’s fusion with lysosomes.

A previous study from the same team had reported that the number of lysosomes produced by the host cells decreases upon infection with Salmonella. The researchers also found that mutant bacteria that were unable to produce SopB were also unable to reduce host lysosome numbers. Therefore, they decided to look more closely at the role that SopB was playing in the production of lysosomes, using advanced imaging techniques.

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What they found was that SopB prevents the translocation of a critical molecule called Transcription Factor EB (TFEB) from the cytoplasm of the host cell into the nucleus. This translocation is vital because TFEB acts as a master regulator of lysosome production.

This is the first time we deciphered that SopB can work in a dual manner – it changes the phosphoinositide dynamics of SCV and affects TFEB’s translocation into the nucleus. While other groups have already reported the function of SopB in mediating invasion in epithelial cells, the novelty of our study lies in identification of the function of SopB in inhibiting the vacuolar fusion with existing autophagosomes/lysosomes, and the second mechanism, which provides Salmonella with a survival advantage by increasing the ratio of SCV to lysosomes.”

Dipshikha Chakravortty, Professor at MCB and corresponding author of the study

The researchers suggest that using small molecule inhibitors against SopB or activators of TFEB can help counter Salmonella infection.

In subsequent studies, the team plans to explore the role of another host protein called Syntaxin-17 whose levels also reduce during Salmonella infection. “How do the SCVs reduce the levels of Syntaxin-17? Do they exchange it with some other molecules, or do the bacteria degrade it? We [plan to] look into it next,” says Chakravortty.

Source:
Journal reference:

Chatterjee, R., et al. (2023) Deceiving The Big Eaters: Salmonella Typhimurium SopB subverts host cell Xenophagy in macrophages via dual mechanisms. Microbes and Infection. doi.org/10.1016/j.micinf.2023.105128.

Novel assay based on hybrid DNA-RNA probe for detecting food contaminated with salmonella

A team of researchers have developed an easy-to-use colorimetric assay for the detection of food contaminated with salmonella. The assay is based on a novel nucleic acid probe that is cleaved by an RNase enzyme specific to the salmonella species. As the team report in the journal Angewandte Chemie, this specific enzymatic cleavage principle made it possible to build a sensitive but simple and portable test system using colloidal gold.

Novel assay based on hybrid DNA-RNA probe for detecting food contaminated with salmonella​​​​​​​

Image Credit: Angewandte Chemie

Consumption of food contaminated with Salmonella typhimurium, whether eggs, ground meat, or chicken, can lead to severe food poisoning. However, suspected cases of salmonella are usually only confirmed several days later, when the bacteria are detected in microbiology laboratories by growing them in culture. A team of researchers led by Yingfu Li, Tohid Didar, and Carlos Filipe of McMaster University in Hamilton, Canada, have now developed a novel test system based on a hybrid DNA-RNA probe that specifically and rapidly detects salmonella, without the need for microbiological diagnostics or expensive analytical equipment.

Using a multi-round selection process, the McMaster team uncovered an artificial DNA-RNA hybrid probe that is a substrate for a salmonella-specific form of an RNase H enzyme. Based on this highly specific enzymatic recognition, the team first developed a fluorescence-based assay on salmonella RNase H, and then extended the principle to a simple, portable salmonella assay based on a colloidal gold colorimetry.

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Colloidal gold is a common color reagent familiar to many of us from its use in SARS-CoV-2 antigen test strips. In a slight departure from this methodology, however, the team did not use a paper strip as the basis for their assay, but instead turned to plastic pipette tips, which are commonly used in the laboratory to measure specific amounts of liquids.

For the preparation of the colorimetric assay, the inner wall of a pipette tip was first coated with DNA-functionalized nanogold. A mixture of reagents composed of nanogold-DNA and the DNA-RNA probe were then sucked up into the pipette tip, causing a double layer of nanogold to form on the walls, because the DNA-RNA hybrid probe links both layers.

However, when the sample mixture contains salmonella, the upper layer is released thanks to the salmonella RNase H specifically cleaving the DNA-RNA linker probe. When the gold-containing solution is then drained onto an absorbent pad with a nylon membrane, a clear red spot indicates the presence of salmonella in the sample being tested. The team also tested the specificity of their system, finding it did not falsely detect the presence of other bacteria containing RNAse H.

The authors highlight that the test is not only much less complex than other methods for detecting salmonella, but also much faster. In contrast to other methods, only one hour of incubation in a pipette tip is required for highly sensitive detection of salmonella, for example, in ground beef. In the future, the team envision developing more nucleic acid probes which can specifically detect other infectious pathogens, for example coliform bacteria such as E. coli.

Source:
Journal reference:

Li, J., et al. (2023). A Simple Colorimetric Au‐on‐Au Tip Sensor with a New Functional Nucleic Acid Probe for Food‐borne Pathogen Salmonella typhimurium. Angewandte Chemie International Edition. doi.org/10.1002/anie.202300828.