Application of pseudomonas in treatment and / or prevention of Alzheimer's disease

Abstract

The invention discloses application of pseudomonas in treatment and / or prevention of Alzheimer's disease, and belongs to the technical field of microorganisms. The invention provides pseudomonas salmonella E14-1 for treating and / or preventing Alzheimer's disease, and the pseudomonas salmonella E14-1 can improve the autophagy level of nematodes and the unfolded protein reaction of endoplasmic reticulum, can reduce the latency period of AD mouse water maze, and can increase the number of platform crossing times. The invention further provides a composition for treating and / or preventing the Alzheimer's disease, a microbial agent and application. The composition and the microbial agent can effectively relieve the symptoms of the Alzheimer's disease.

Description

Technical Field

[0001] This invention relates to the field of microbial technology, and in particular to the use of a strain of Pseudomonas in the treatment and / or prevention of Alzheimer's disease. Background Technology

[0002] Alzheimer's disease (AD) is a chronic, progressive neurodegenerative disease and the most common cause of dementia. Its typical pathological features include senile plaques formed by the deposition of β-amyloid (Aβ) protein in the brain and neurofibrillary tangles formed by hyperphosphorylated tau protein. Early symptoms include memory loss, followed by language impairment, behavioral changes, and a decline in daily living abilities.

[0003] Pseudomonas spp. ( Pseudomonas *Pseudomonas salmaria* is a Gram-negative, aerobic, rod-shaped bacterium with single-ended flagella, belonging to the phylum Proteobacteria, class Gamma-Proteobacteria. It measures approximately 0.5–1.0 μm × 1.5–5.0 μm, is non-spore-forming, and some species have capsules. It is oxidase-positive, does not ferment sugars, and often produces water-soluble pigments such as pyocyanin and luciferin. It grows well on ordinary culture media at 30–35°C, with some species tolerating 4°C or 42°C. Over 200 species have been named in this genus. *Pseudomonas salmaria* is widely distributed in water, soil, and air, and is a key functional group in soil ecosystems. It can inhibit pathogens, decompose organic pollutants, and is used in bioremediation, agricultural biocontrol, and food preservation; this genus has significant ecological application value. This invention proposes a species identified by sequencing as *Pseudomonas salmaria*, which has been shown to alleviate Alzheimer's disease. Summary of the Invention

[0004] Based on the technical problems to be solved by the present invention, the present invention proposes the application of a strain of Pseudomonas in the treatment and / or prevention of Alzheimer's disease.

[0005] One of the objectives of this invention is to provide a strain of Pseudomonas E14-1 for the treatment and / or prevention of Alzheimer's disease (… Pseudomonas salmasensis The strain is deposited at the China General Microbiological Culture Collection Center (CGMCC) with accession number CGMCC No. 37145.

[0006] Furthermore, the strain was able to enhance autophagy levels and endoplasmic reticulum unfolded protein responses in nematodes.

[0007] Furthermore, the strain was able to reduce the latency period in the water maze of AD mice and increase the number of platform crossings.

[0008] A second objective of this invention is to provide a composition for treating and / or preventing Alzheimer's disease, the composition comprising Pseudomonas E14-1, a culture of the strain, or a dried product of the culture of the strain as an active ingredient.

[0009] Furthermore, the composition has the effect of improving cognitive and memory functions.

[0010] A third objective of this invention is to provide a bacterial agent for treating and / or preventing Alzheimer's disease, wherein the bacterial agent is a powder obtained by drying a bacterial solution containing Pseudomonas E14-1.

[0011] The fourth objective of this invention is to provide the use of Pseudomonas E14-1, a composition, or a bacterial agent in the preparation of a medicament for the treatment and / or prevention of Alzheimer's disease.

[0012] Furthermore, the drug contains Pseudomonas E14-1 and a pharmaceutically acceptable carrier.

[0013] Furthermore, the carrier comprises one or more of pharmaceutically acceptable protectants, fillers, disintegrants, and lubricants.

[0014] Furthermore, the drug is a drug for treating and / or preventing important pathological features caused by Alzheimer's disease, including Aβ plaque deposition, a large number of Aβ plaques, and a large Aβ plaque area.

[0015] Compared with the prior art, the present invention proposes the application of a strain of Pseudomonas aeruginosa in the treatment and / or prevention of Alzheimer's disease, which has the following beneficial effects: The Pseudomonas E14-1 strain proposed in this invention can alleviate Alzheimer's disease by increasing autophagy levels and endoplasmic reticulum unfolded protein responses. Attached Figure Description

[0016] Figure 1 The diagram illustrates how a strain of Pseudomonas aeruginosa, according to an embodiment of the present invention, can alleviate the paralysis phenotype in an AD model.

[0017] Figure 2 An evolutionary tree diagram of a Pseudomonas strain according to an embodiment of the present invention is shown.

[0018] Figure 3 The diagram illustrates the effect of a Pseudomonas strain on autophagy levels according to an embodiment of the present invention; wherein, the fluorescence display diagram of PHA-4::GFP is shown, n=25-46; data are expressed as mean ± standard error (t-test). <0.01, scale bar, 20μm.

[0019] Figure 4This diagram illustrates the elevated endoplasmic reticulum unfolded protein reaction of a Pseudomonas strain according to an embodiment of the present invention; wherein, P hsp-4 ::GFP fluorescence display diagram, n=25-45, data are expressed as mean ± standard error (t-test). <0.0001; Scale bar, 100μm.

[0020] Figure 5 This paper illustrates how a strain of *Pseudomonas* improved the latency time in the water maze of Alzheimer's disease (AD) mice, as shown in the statistical graph of the number of platform crossings. The data are presented as mean ± standard error (one-way ANOVA). <0.05, ns indicates no statistical difference. Detailed Implementation

[0021] Exemplary embodiments of the present application will now be described in more detail with reference to the accompanying drawings. While exemplary embodiments of the present application are shown in the drawings, it should be understood that the present application may be implemented in various forms and should not be limited to the embodiments set forth herein. Rather, these embodiments are provided so that this application will be thorough and complete, and will fully convey the scope of the present application to those skilled in the art.

[0022] Unless otherwise specified, the technical means used in the following embodiments are conventional means well known to those skilled in the art, and the reagents and materials in this invention are obtained from the market or other public channels.

[0023] This invention provides the application of a *Pseudomonas* strain in the treatment and / or prevention of Alzheimer's disease. The *Pseudomonas* strain is E14-1, a strain collected from the environment, screened, and cultured in TSB medium, possessing properties for treating and / or preventing Alzheimer's disease. This strain is deposited at the China General Microbiological Culture Collection Center (CGMCC) with accession number CGMCC No. 37145; its classification name is *Pseudomonas salmasidium* (…). Pseudomonas salmasensis ); Deposit date: December 24, 2025; Deposit address: Institute of Microbiology, Chinese Academy of Sciences, No. 1 Beichen West Road, Chaoyang District, Beijing; The test result of this strain is that it is viable.

[0024] See Figure 1 The Pseudomonas strain E14-1 of this invention can alleviate the paralysis phenotype in an AD model. The paralysis rate was statistically analyzed by culturing AD model nematodes Aβ and then heat shocking them at 25°C.

[0025] Example 1 This invention proposes an experiment for obtaining and identifying Pseudomonas.

[0026] Mainly includes: This embodiment uses Caenorhabditis elegans (Caenorhabditis elegans) Caenorhabditis elegans Large-scale unbiased screening was performed using a model. Different bacterial strains were collected from the environment, and single colonies were isolated by streaking. Single colonies were then placed in TSB medium (tryptic-soybean peptone liquid medium) and incubated at 28°C for 24 hours to obtain independent bacteria. TSB medium, based on two peptones (tryptic peptone + soybean peptone), is nutritionally balanced and can support the growth of various microorganisms. Tryptic peptone, derived from trypsin digests of casein, provides abundant nitrogen, carbon, and growth factors, serving as the main nutritional basis of the medium. Soybean peptone, derived from soybean hydrolysate, provides additional nitrogen and vitamins, especially B vitamins, further enriching the nutrition and aiding in the cultivation of fastidious bacteria. The medium also contains sodium chloride to provide an isotonic environment and dipotassium hydrogen phosphate as a buffer. The bacterial culture was centrifuged to remove the supernatant. The resulting bacterial blocks were lysed and subjected to PCR 16S rRNA sequencing. 16S result comparison confirmed that the bacterium (E14-1) belongs to the genus *Pseudomonas*. E14-1 strain and Pseudomonas salmas SWRI126T (JABWQJ010000063) P. paracarnis V5 / DAB / 2 / 5T(KX186983) P.lactis DSM 29167T(JYLO01000038), P. pergaminensis 1008T(CP078013) gathered in one ( Figure 2 The similarity between this strain and the four strains was 99.45%.

[0027] The original genome data of strain E14-1 was 1310 Mb, and the data size after quality control was 1294 Mb. The G+C content of the entire genome was 60.08%. Based on the calculation results, the ANI and dDDH of strain E14-1 and strains with high homology are shown in Table 1. (The text then abruptly shifts to a seemingly unrelated topic: strain E14-1 and...) P. salmasensis The ANI was 98.63%, compared to P. paracarnis The ANI was 95.7%, compared to P.lactis The ANI was 95.2%, compared to P. pergaminensis The ANI was 87.08%, indicating that they may be the same species. Meanwhile, strain E14-1 and... P. salmasensis The dDDH was 88.7%, compared to P. paracarnis , P.lactis , P. pergaminensis All dDDH values ​​were below the species-defined threshold (<70%), therefore strain E14-1 can be identified as... P. salmasensis .

[0028] Table 1. List of ANI and dDDH values ​​(%) of E14-1 and closely related species. Example 2 This invention proposes a method by which Pseudomonas aeruginosa alleviates the phenotype of the Alzheimer's disease (AD) nematode model by enhancing autophagy and increasing the endoplasmic reticulum unfolded protein response.

[0029] Mainly includes: This embodiment uses autophagy markers to detect the effect of *Pseudomonas salmasidium* on autophagy. PHA-4 is a transcription factor that regulates autophagy in *C. elegans*. Since PHA-4 expression is divided into two aggregation regions in the nematode head, the number of nuclei entering the nucleus in each region was counted. See the fluorescence display diagram of the PHA-4::GFP fusion protein for details. Figure 3 The study revealed the temporal and spatial expression patterns of PHA-4 as a key transcription factor, and these patterns were visualized using GFP fluorescence. Based on observations and nucleus counts of 25-46 live samples of *C. elegans* expressing the PHA-4::GFP fusion protein, it was concluded that *Pseudomonas salmas* enhanced autophagy.

[0030] Furthermore, studies have shown that increased endoplasmic reticulum unfolded protein responses can combat AD and alleviate neurodegenerative diseases. This was demonstrated after feeding *Pseudomonas salmas*, based on studies of 25-45 cells expressing P… hsp-4 For observation of live samples of *Caenorhabditis elegans* containing GFP, see [link to relevant documentation]. Figure 4 Compared with the OP50 control group, the fluorescence intensity of nematodes was significantly increased. Therefore, it was concluded that *Pseudomonas salmasidium* can increase the unfolded protein response of the endoplasmic reticulum.

[0031] Example 3 This invention presents an experimental method for reducing the latency period in AD mice treated with Pseudomonas salmasidium.

[0032] Mainly includes: This embodiment conducts a water maze experiment, a classic behavioral experiment used in neuroscience research to assess the spatial learning and memory abilities of rodents, such as mice. This experiment examines how mice learn and remember the spatial location of an escape platform relative to surrounding visual cues through exploration and memorization, testing the mice's referential memory of their spatial environment, which depends on hippocampal function. In AD model mice, this typically manifests as prolonged latency and reduced time spent in the target quadrant. This experiment utilizes the mice's natural aversion to water, guiding them to find a platform hidden underwater. See [link to relevant documentation]. Figure 5 The statistical graphs of latency time and platform crossing number of AD mice in the water maze show that AD mice fed with Pseudomonas salmas experienced a reduction in latency during the training period, and AD mice fed with Pseudomonas salmas experienced a reduction in latency and an increase in platform crossing number during the testing period.

[0033] In summary, this invention proposes the application of a Pseudomonas strain in the treatment and / or prevention of Alzheimer's disease. The Pseudomonas E14-1 strain can alleviate Alzheimer's disease by increasing autophagy levels and endoplasmic reticulum unfolded protein responses.

[0034] It should be noted that the term "comprising," or any other variation thereof, is intended to cover non-exclusive inclusion, such that a process, method, article, or apparatus that comprises a list of elements includes not only those elements but also other elements not expressly listed, or elements inherent to such a process, method, article, or apparatus. Without further limitation, an element defined by the phrase "comprising..." does not exclude the presence of other identical elements in the process, method, article, or apparatus that includes said element.

[0035] The above are merely embodiments of this application and are not intended to limit the scope of this application. Various modifications and variations can be made to this application by those skilled in the art. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of this application should be included within the scope of the claims of this application.

Claims

1. A strain of Pseudomonas E14-1 used for the treatment and / or prevention of Alzheimer's disease ( Pseudomonas salmasensis ), characterized in that, The strain is deposited at the China General Microbiological Culture Collection Center, accession number CGMCC No. 37145.

2. The *Pseudomonas* E14-1 for treating and / or preventing Alzheimer's disease according to claim 1, characterized in that, The Pseudomonas E14-1 is Pseudomonas Salmas E14-1; The strain of *Pseudomonas salmasidium* E14-1 was able to enhance autophagy levels and endoplasmic reticulum unfolded protein response in nematodes.

3. The *Pseudomonas* E14-1 for treating and / or preventing Alzheimer's disease according to claim 1, characterized in that, The strain was able to reduce the latency of the water maze in AD mice and increase the number of platform crossings.

4. A composition for treating and / or preventing Alzheimer's disease, characterized in that, The composition comprises, as an active ingredient, the Pseudomonas E14-1 strain culture or the dried product of the strain culture as described in claim 1.

5. The composition for treating and / or preventing Alzheimer's disease according to claim 4, characterized in that, The composition can improve the body's cognitive and memory functions.

6. A bacterial agent for treating and / or preventing Alzheimer's disease, characterized in that, The bacterial agent is a powder obtained by drying a bacterial solution containing the Pseudomonas E14-1 of claim 1.

7. The use of Pseudomonas E14-1 according to any one of claims 1 to 3, or the composition according to any one of claims 4 to 5, or the bacterial agent according to claim 6 in the preparation of a medicament for treating and / or preventing Alzheimer's disease.

8. The application according to claim 7, characterized in that, The drug contains the Pseudomonas E14-1 as described in claim 1 and a pharmaceutically acceptable carrier.

9. The application according to claim 7, characterized in that, The carrier contains one or more of pharmaceutically acceptable protectants, fillers, disintegrants, and lubricants.

10. The application according to claim 7, characterized in that, The drug is a drug for treating and / or preventing important pathological features caused by Alzheimer's disease, including Aβ plaque deposition, a large number of Aβ plaques, and a large Aβ plaque area.

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