NL2035175B1

NL2035175B1 - Lactobacillus paracasei ty-f15 and application thereof

Info

Publication number: NL2035175B1
Application number: NL2035175A
Authority: NL
Inventors: Zhao Lijuan; Zhu Xu; Deng Tao; Gu Mingyu; Zhang Feng; Zhang Zhen; Wang Jing; Hong Yang; Chen Zuguo; Li Wanzhu
Original assignee: Chongqing Tianyou Dairy Co Ltd
Priority date: 2023-03-01
Filing date: 2023-06-26
Publication date: 2024-03-25
Also published as: NL2035175A; CN118580982A

Abstract

A lactobacillus paracasei TY—F15 to which the accession number of CGMCC No. 25742 is assigned. The lactobacillus paracaseì TY—F15 has the capability of inhibiting fungal growth. Using the lactobacillus paracasei as an auxiliary starter to ferment cow milk may delay mold development in fermented milk, such that the fermented milk may be stored by more than 22 days at 10°C, and a storage period of the fermented milk is prolonged by more than 10d-28d compared with fermented milk that is not added with the TY-F15. The fermented milk may also be stored more than 6 days at room temperature of 25°C, such that the storage period is prolonged by more than 2d-8d compared with the fermented milk that is not added with the TY—F15, and a tissue state is better than a tissue state that is not added with the TY—F15.

Description

LACTOBACILLUS PARACASEI TY-F15 AND APPLICATION THEREOF

TECHNICAL FIELD

The present application belongs to the technical field of microorganism applications, and specifically, relates to a lactobacillus paracasei TY-F15 and an application thereof.

BACKGROUND

Fungi are the main microorganisms that cause spoilage of fermented products. Every year, food contamination caused by the fungi leads to significant waste and causes great economic losses to our country. As the number one fermented dairy product in China, yogurt is rich in nutrients and has health functions, which is very popular among people. However, as the yogurt contains active lactobacilli, the yogurt has a short shelf life and is highly susceptible to contamination by acid-resistant fungi during processing, packaging, transportation and storage, especially the fungi (such as trichoderma longibrachiatum, aspergillus aculeatus, aspergillus fumigatus, and saccharomyces) whose growth pH is similar to that of the yogurt and that may multiply in large numbers in yogurt, such that the shelf life of the yogurt is seriously affected. In addition, the proliferation of the fungi in the yogurt also causes peculiar smell, package expansion, bulging lids and whey precipitation during the shelf life of the yogurt, resulting in food safety problems and economic losses.

Currently, methods for preventing or reducing fungal contamination in fermented milk are mainly to add chemical preservatives or improve processing devices and packaging measures. However, the addition of preservatives is potentially harmful, and improved processing devices and packaging measures have the drawback of significantly increasing production costs.

Therefore, reducing fungal contamination by avoiding use of chemical preservatives without increasing production costs is of great significance for preparation of the fermented milk.

SUMMARY in view of the above problems, the present application is intended to provide a lactobacillus paracasei TY-F15, which is derived from home-made traditionally-fermented yak yogurt from herdsmen in Hongyuan County, Aba Tibetan and Qiang Autonomous Prefecture,

Sichuan Province. The lactobacillus paracasei TY-F15 has the capability of obviously inhibiting fungal growth, and may delay mold development in fermented milk by being used as a starter or an auxiliary starter to ferment the fermented milk, so as to improve post-acidification of the fermented milk, thereby effectively prolonging the shelf life of the fermented milk, and a tissue state is better than a tissue state that is not added with the TY-F15. In addition, there is also no potential harm from chemical preservatives, and processing devices and packaging measures do not need to be improved to increase production costs.

In order to achieve the above purpose, the present application may use the following technical solutions.

One aspect of the present application provides a lactobacillus paracasei TY-F15, which is assigned with the accession number of CGMCC No. 25742.

Another aspect of the present application provides a composition, The composition includes one or a combination of a plurality of the following substances: (a) the viable lactobacillus paracasei TY-F15; (b) lysate of the lactobacillus paracasei TY-F15; (c) a culture of the lactobacillus paracasei TY-F15; and (d) a fermentation broth of the lactobacillus paracasei

TY-F15.

Still another aspect of the present application provides a preparation. The preparation may include the lactobacillus paracasei TY-F15 or the composition, and a carrier. The carrier is a medicinal carrier or an edible carrier.

Still another aspect of the present application provides an application of the lactobacillus paracasei TY-F15 in resisting spoilage bacteria.

The collection information of the lactobacillus paracasei TY-F15 in the present application includes the following. The lactobacillus paracasei TY-F15 is deposited with the China General

Microbiological Culture Collection Center {CGMCC) at No. 3, Yard 1, BeiChen West Road,

Chaoyang District, Beijing on September 16, 2022 and is assigned with the accession number of CGMCC No. 25742, with the classification name being lactobacillus paracasei.

The beneficial effects of the present application include the following. (1) The lactobacillus paracasei TY-F15 provided in the present application has the capability of obviously inhibiting fungal growth {for example, different species and genera of fungi {trichoderma longibrachiatum (ID), aspergillus aculeatus (LM), aspergillus fumigatus (DHZ), penicillium sp {CICC2515} and aspergillus niger (CICC2487)), especially the fungal species (such as the trichoderma longibrachiatum, the aspergillus aculeatus and the aspergillus fumigatus) that spread in the air of a yogurt production plant which can cause contamination to, and grow in, the yogurt, {2} Using the lactobacillus paracasei TY-F15 provided in the present application as the auxiliary starter to ferment the cow milk may delay mold development in the fermented milk,

such that the fermented milk may be stored more than 22 days at 10°C, and a storage period is prolonged by more than 10d-28d compared with fermented milk that is not added with the

TY-F15; and the fermented milk may also be stored more than 6 days at room temperature of 25°C, such that the storage period is prolonged by more than 2d-8d compared with the fermented milk that is not added with the TY-F15. In addition, there is also no potential harm from chemical preservatives. (3) By means of using the lactobacillus paracasei TY-F15 provided in the present application as the starter to ferment the cow milk, production costs increased by improving the processing devices and the packaging measures to avoid fungal contamination may be avoided. {4) By means of using the lactobacillus paracasei TY-F15 provided in the present application as the starter to ferment the cow milk, compared with the fermentation of the commercial strains that are not added with the lactobacillus paracasei TY-F15, the stability and overall sensory evaluation of the fermented milk may be improved.

BRIEF DESCRIPTION OF THE DRAWINGS

Fig. 1 shows colonial morphology of common spoilage fungi and fungal standard strains in dairy products.

Fig. 2 shows colonial morphology of TY-F15.

Fig. 3 shows a gram staining result of TY-F15.

Fig. 4 shows a plate inhibiting effect of TY-F15 on indicator bacteria.

Fig. 5a shows effects of penicillium indicator bacteria and addition of TY-F15 on mold development in a yogurt at 10°C.

Fig. 5b shows effects of aspergillus niger indicator bacteria and addition of TY-F15 on mold development in a yogurt at 10°C.

Fig. 5c shows effects of aspergillus fumigatus indicator bacteria and addition of TY-F15 on mold development in a yogurt at 10°C,

Fig. 5d shows effects of trichoderma longibrachiatum indicator bacteria and addition of

TY-F15 on mold development in a yogurt at 10°C.

Fig. 5e shows effects of aspergillus aculeatus indicator bacteria and addition of TY-F15 on mold development in a yogurt at 10°C.

Fig. 6a shows effects of penicillium indicator bacteria and addition of TY-F15 on mold development in a yogurt at 25°C.

Fig. 6b shows effects of aspergillus niger indicator bacteria and addition of TY-F15 on mold development in a yogurt at 25°C.

Fig. 6c shows effects of trichoderma longibrachiatum indicator bacteria and addition of

TY-F15 on mold development in a yogurt at 25°C.

Fig. 6d shows effects of aspergillus aculeatus indicator bacteria and addition of TY-F15 on mold development in a yogurt at 25°C.

Fig. 6e shows effects of aspergillus fumigatus indicator bacteria and addition of TY-F15 on mold development in a yogurt at 25°C.

Fig. 7 shows the impact of TY-F15 on a fermentation state of a yogurt.

Fig. 8 shows the impact of TY-F15 on fermentation time of a yogurt.

Fig. 9 shows the impact of TY-F15 on a pH value and titratable acidity of a yogurt during storage.

Fig. 10 shows the impact of TY-F15 on stability (water holding capacity and clarification index) of a yogurt during storage.

Fig. 11 shows the impact of TY-F15 on the texture of a yogurt during storage.

In Fig. 7 to Fig. 11, Group TY-F15 refers to the group that TY-F15 assists in fermentation of a commercial starter.

DETAILED DESCRIPTION OF THE EMBODIMENTS

The embodiments are given to better describe the present application, but the content of the present application is not limited only to the embodiments given. Therefore, non-essential improvements and adjustments to the embodiments made by a person skilled in the art in accordance with the content of the above present application still fall within the scope of protection of the present application.

The terms used herein are only intended to describe specific embodiments and are not intended to limit the present disclosure. Expressions in the singular form include those in the plural form unless the expressions have a distinctly different meaning in the context. As used herein, it is to be understood that terms such as "include", "have", "contain", and the like are intended to indicate the presence of features, figures, operations, or combinations. The terms of the present application are disclosed in the specification and are not intended to exclude the possibility that one or more other features, figures, operations, or combinations thereof may exist or may be added. As used here, "/" may be interpreted as "and" or "or", as appropriate.

An embodiment of the present application provides a lactobacillus paracasei TY-F15, which is assigned with the accession number of CGMCC No. 25742.

It is to be noted that, the lactobacillus paracasei TY-F15 is separated from home-made traditionally-fermented yak yogurt from herdsmen in Hongyuan County, Aba Tibetan and

Qiang Autonomous Prefecture, Sichuan Province. By means of sequencing, a 16S rDNA sequence of the lactobacillus paracasei TY-F15 is shown as SEQ ID NO.1. 5 In addition, the lactobacillus paracasei TY-F15 is a probiotic. The probiotics are active microorganisms that can colonize in a human body and may change the composition of the flora in a certain part of the human body, and may achieve the effect of promoting nutrient absorption and maintaining intestinal health by means of regulating immune functions of the human mucosa and system, or by regulating the balance of gut microbiota in the human body.

The lactobacillus is the most common type of traditional probiotics, which may ferment carbohydrates to produce lactic acid and is functional flora that regulates human intestinal health, including lactobacillus spp and bifidobacterium.

Another embodiment of the present application further provides a composition. The composition may include one or a combination of a plurality of the following substances: (a) the viable lactobacillus paracasei TY-F15; (b) lysate of the lactobacillus paracasei TY-F15; (c) a culture of the lactobacillus paracasei TY-F15; and (d) a fermentation broth of the lactobacillus paracasei TY-F15. it is to be noted that, the lactobacillus paracasei TY-F15 may be used in different forms, or in combination, as a component of a composition. For example, according to specific requirements, one or a combination of a plurality of forms of the viable lactobacillus paracasei

TY-F15, the lysate, the culture and the fermentation broth may be combined in the composition. The viable lactobacillus paracasei TY-F15 may be bacterial fluid, or may be in the form of bacterial powder. On the basis of a probiotic anti-fungal function of the lactobacillus paracasei TY-F15, the composition containing the lactobacillus paracasei TY-F15 also has the probiotic anti-fungal function.

In some specific embodiments, the composition may further include one or a combination of probiotics, prebiotics, dietary fiber and traditional Chinese drugs.

It is to be noted that, the lactobacillus paracasei TY-F15 and different forms thereof may also be used in combination with one or the combination of probiotics, dietary fiber and a pharmacologically active compound. For example, the lactobacillus paracasei TY-F15 may be used in combination with bacillus subtilis, bifidobacterium or lactobacillus, so as to cause the composition to simultaneously have the effects of the lactobacillus paracasei TY-F15 and other probiotics. For another example, the lactobacillus paracasei TY-F15 may be used in combination with prebiotics, and the prebiotics may provide an energy source for the lactobacillus paracasei TY-F15, such that the effect of the lactobacillus paracasei TY-F15 may be improved. For another example, the lactobacillus paracasei TY-F15 may be used in combination with the dietary fiber, and the dietary fiber may assist in colonization of the lactobacillus paracasei TY-F15, such that the effect of the lactobacillus paracasei TY-F15 may be improved. For another example, the lactobacillus paracasei TY-F15 may be used in combination with the traditional Chinese drugs to form the composition, such that the effects of the lactobacillus paracasei TY-F15 and the traditional Chinese drugs may be simultaneously achieved.

Still another embodiment of the present application provides a preparation. The preparation may include the lactobacillus paracasei TY-F15 or the composition, and a carrier.

The carrier is a medicinal carrier or an edible carrier.

It is to be noted that, drugs or edible food or health care products may be prepared by adding the medicinal carrier or the edible carrier to the composition including the lactobacillus paracasei TY-F15 and different forms thereof. The medicinal carrier or the edible carrier is known in the art and may be selected according to the dosage form as needed. For example, the preparation of tablets mainly uses a diluent (such as starch, dextrin, sucrose or sugar), an absorbent (such as calcium sulfate, calcium hydrogen phosphate or light magnesium oxide), an adhesive (such as povidone, syrup or hydroxypropyl! methylcellulose), a wetting agent (such as water), or a disintegrating agent {such as dry starch, sodium hydroxymethyl starch or cross-linked povidone). For example, the preparation of the liquid preparation mainly uses a bulking agent, a suspending agent, an emulsifying agent, a colorant, or the like.

In some specific embodiments, the preparation may be tablets, pills, capsules, powder, gel, granules or a liquid preparation, etc.

It is to be noted that, solid dosage forms such as the tablets, the pills, the granules or the capsules may be product forms such as probiotic tablets, probiotic sugar pills, probiotic powder or probiotic capsules. The liquid preparation may be a product form such as a probiotic beverage. The gel may be product forms such as probiotic jelly, probiotic milk foam or solidified yogurt.

Still another embodiment of the present application provides an application of the lactobacillus paracasei TY-F15 in resisting spoilage bacteria. it is to be noted that, as described above, the lactobacillus paracasei TY-F15 may act as a safe edible bacterium, and metabolites of the lactobacillus paracasei TY-F15 have a significant inhibiting effect on the spoilage bacteria, such that the lactobacillus paracasei TY-F15 has the potential to develop into a natural preservative.

In some specific embodiments, in the above application, the spoilage bacteria may include one or more of penicillium, aspergillus niger, trichoderma longibrachiatum, aspergillus aculeatus, and aspergillus fumigatus.

It is to be noted that, the lactobacillus paracasei TY-F15 especially has a strong inhibiting effect on the 5 spoilage bacteria mentioned above. In particular, the trichoderma longibrachiatum, the aspergillus aculeatus and the aspergillus fumigatus are three fungal species that can grow in the fermented milk and need to be focused on prevention in actual fermented milk processing production.

In some specific embodiments, the application may include any one of the following: (a) applying the lactobacillus paracasei TY-F15 to preparation of fermented milk as a fermented strain; (b) applying a combination of the lactobacillus paracasei TY-F15 and another fermented strain to preparation of the fermented milk as the fermented strain; and (c) applying the viable lactobacilius paracasei TY-F15 or a metabolite thereof to drugs or food as an additive.

It is to be noted that, there are two main factors affecting the shelf life of the fermented milk. On the one hand, fungal contamination during production and processing generally causes the fermented milk to bulge and produce peculiar smell; and on the other hand, the flavor of the fermented milk changes to be too sour, affecting the taste of consumers, which is mainly caused by post-acidification of yogurt during storage.

The lactobacillus paracasei TY-F15 in the present application has an anti-fungal function, such that the lactobacillus paracasei may be applied to preparation of the fermented milk by being used as a fermentation strain or in combination with another fermentation strain, to reduce fungal contamination as a biologic preservative, so as to delay mold development of the fermented milk. As described above, the trichoderma longibrachiatum, the aspergillus aculeatus and the aspergillus fumigatus are the fungal species that can grow in the fermented milk, and are the fungal species that need to be prevented in actual fermented milk processing production. The TY-F15 has a certain inhibiting effect on the above fungi below 10°C or at about 25°C, such that by means of applying the lactobacillus paracasei to preparation of the fermented milk, the storage period of the fermented milk may be prolonged.

In some specific embodiments, compared with normal fermentation without using the

TY-F15, if the fermented milk simultaneously contains the penicillium sp, the aspergillus niger, the aspergillus aculeatus, and the aspergillus fumigatus, the storage period using the TY-F15 at 10°C may also be prolonged by about 10 days; if the fermented milk does not contain the aspergillus niger in the above 5 molds, the storage period using the TY-F15 at 10°C may be prolonged by about 16 days; if the fermented milk does not contain the aspergillus niger and the aspergillus fumigatus in the above 5 molds, the storage period using the TY-F15 at 10°C may be prolonged by about 18 days; if the fermented milk does not contain the aspergillus niger, aspergillus fumigatus and the trichoderma longibrachiatum in the above 5 molds, the storage period using the TY-F15 at 10°C may be prolonged by about 22 days; and if the fermented milk does not contain the aspergillus niger, the aspergillus aculeatus, the aspergillus fumigatus, and the trichoderma longibrachiatum in the above 5 molds, the storage period using the TY-F15 at 10°C may be prolonged by more than 28 days. That is, according to the fungal strains carried in the fermented milk, the storage period, using the TY-F15, of the fermented milk added with the TY-F15 at 10°C is prolonged by more than 10d-28d.

In addition, compared with the normal fermentation without using the TY-F15, if the fermented milk contains all the 5 molds, the storage period using the TY-F15 at 25°C may also be prolonged by about 2 days; if the fermented milk does not contain the aspergillus niger in the above 5 molds, the storage period using the TY-F15 at 25°C may be prolonged by about 5 days; if the fermented milk does not contain the aspergillus niger and the aspergillus fumigatus in the above 5 molds, the storage period using the TY-F15 at 25°C may be prolonged by about 6 days; and if the fermented milk does not contain the aspergillus niger, the aspergillus aculeatus, the aspergillus fumigatus, and the trichoderma longibrachiatum in the above 5 molds, the storage period using the TY-F15 at 25°C may be prolonged by about 8 days. That is, according to the fungal strains carried in the fermented milk, the storage period of the fermented milk added with the TY-F15 at 25°C may be prolonged by more than 2d-8d. it is further to be noted that, the TY-F15 has a poor inhibiting effect on the trichoderma longibrachiatum in a plate, but has good inhibiting effect in the fermented milk, such that the

TY-F15 may be well applied to preparation of the fermented milk.

In addition, when the lactobacillus paracasei TY-F15 works together with another fermentation strain, the yogurt fermented by using the TY-F15 as the auxiliary starter is smooth in surface, uniform in color, good in tissue state, and has no peculiar smell. In some specific embodiments, addition of the TY-F15 in the fermented milk may improve the slightly rough and non-smooth surface of the fermented milk fermented with the commercial starter,

thereby improving the overall acceptance of the fermented milk.

In addition, as auxiliary inoculation of the TY-F15, there is no significant effect on fermentation time of the fermented milk, because the TY-F15 proliferates slowly and does not affect a fermentation process of the fermented milk during fermentation. In some specific embodiments, the TY-F15 proliferates as a dominant bacterium in the yogurt during the late storage period, inhibiting the growth of the lactobacillus bulgaricus {which is the main strain causing the post-acidification of the yogurt), such that the post-acidification of the yogurt is alleviated to a certain extent, which has a positive effect on prolonging the shelf life of the yogurt.

For the above, the lactobacillus paracasei TY-F15 in the present application has a positive role in prolonging the shelf life of the fermented milk by being applied to preparation of the fermented milk; and the fermented milk obtained by means of fermentation is good in tissue state, uniform in color, and desirable in stability and sensory quality, and has no peculiar smell.

The tissue state is better than that of the fermented milk prepared without adding the lactobacillus paracasei TY-F15. In addition, there is also no potential harm from chemical preservatives. Therefore, the TY-F15 has the potential of developing fermented food with long shelf life such as the fermented milk, and has wide application prospects.

In addition to this, the viable lactobacillus paracasei TY-F15 or a metabolite thereof may also be used as an additive to be directly added to drugs or food as a probiotic element, such that the TY-F15 not only has the function of probiotics, but also may delay mold development in drugs or food, thereby prolonging the shelf life. It is to be noted that, the metabolite may include a fermentation broth, a culture, and lysate.

In some specific embodiments, in the above application, the fermented milk may be a yogurt.

In some specific embodiments, in the above application, another fermented strain comprises one or a combination of a plurality of the following: a lactobacillus bulgaricus and a streptococcus thermophilus. it is to be noted that, the lactobacillus bulgaricus was the main strain causing the post-acidification of the yogurt, and the TY-F15 proliferates as the dominant bacterium in the yogurt in the later stage of storage, inhibiting the growth of the lactobacillus bulgaricus, such that the post-acidification of the yogurt is alleviated to a certain extent.

In order to better understand the present application, the content of the present application is further described below with reference to specific embodiments, but is not only limited to the following examples.

Embodiment 1: Separation, purification and identification of common molds in yogurt (1) Spoiling of experimental samples

A Tianyou classic original yogurt, a Tianyou blueberry grape fruit yogurt, and a Tianyou red date yogurt, which were newly prepared by Chongqing Tianyou Dairy Co., Ltd, were respectively placed, in an open manner, in a yogurt production workshop for air exposure spoiling treatment; the yogurts were placed for 7d at room temperature; and after airborne microorganisms have naturally settled on the yogurts and colonies visible to the naked eye have been formed, molds were separated, and purification and biological identification were performed on the molds. (2) Preparation of spoiled sample diluent

Colonies with different forms on the yogurts were selected under an aseptic condition to 5 mL of sterilized distilled water containing glass beads; full shaking and well mixing were performed for 2h on a 37°C constant temperature shaking table, so as to form sample diluent of 1:10; a density gradient dilution method was used to dilute the samples to 107 in sequence, the diluent with three gradients of 10%, 10% and 10° was selected, 100 uL was taken for each gradient and spread on the surface of a rose-bengal solid culture medium, three plates were made for each gradient (10%, 10° and 10%}, and inverted culture was performed for 3-5 days at 25°C; single colonies with different forms were selected and placed on a Potato Dextrose

Agar (PDA) solid culture medium to perform purification for 2-3 times, so as to obtain single indicator bacteria; the purified single bacteria were inoculated on a slope and temporarily stored at 4°C for later experiment use. (3) Counting and storage of spoilage fungi

Under an aseptic condition, spores on the surface of the PDA solid culture medium were placed in a 0.05% (V/V) Tween-80 solution for full shaking; bacteria were filtered by using sterilized gauze; the concentration of the spore solution was regulated by using sterile saline to an appropriate concentration, and a hemocytometer was used to count the spores under a microscope; and 10% was finally regulated for later use.

The purified single indicator bacteria were inoculated into the PDA solid culture medium, and after the spores grown on the surface of the culture medium in a culture dish, washing was performed with a small amount of sterile water, so as to prepare into spore suspension of the indicator bacteria; sandy soil was sieved, dried, split, and sterilized; under the aseptic condition, the spore suspension was dropped in a sandy soil tube, and placed in a dryer for drying, and the sandy soil tube was sealed for long-term storage after moisture was completely evaporated; and the indicator bacteria spores were in a dormant state in the sandy soil, and might be revived by placing the spores in a moist environment when the spores were used again. (4) 26SrDNA sequence analysis and molecular identification of spoilage fungi

After air exposure treatment was performed on the yogurts for 7d according to the above method, 9 colonies visible to the naked eye were found; after the colonies were transferred to the rose-bengal culture medium for separation, purification, and expansion cultivation, 3 spoilage bacteria were totally separated from three different yogurts. The spoilage bacterium grown in the Tianyou classic original yogurt was named as JD; the spoilage bacterium grown in the Tianyou blueberry grape fruit yogurt was named as LM; and the spoilage bacterium grown in the Tianyou red date yogurt was named as DHZ. 1 mL of the purified JD, LM and DHZ indicator bacterial spore suspension was respectively taken; after centrifugation, precipitates were placed and frozen for 30 min at -80°C, and then a Cetyltrimethylammonium Ammonium Bromide {CTAB) method was used to extract the DNA of precipitated bacteria; and after a fungal Internal Transcribed Spacer (ITS) amplification universal primer ITS1 (5'-TCCGTAGGTGAACCTGCGG-3’SEQ ID NO.5) and an ITS4 (5'-TCCTCCGCTTATTGATATGC-3’SEQ ID NO.6) were used to amplify the extracted DNA, Sangon

Biotech (Shanghai) Co., Ltd. was entrusted to sequence qualified PCR amplification products, and after the sequence was obtained, searching and similarity comparison were performed in

GenBank by using BLAST(http://www.ncbi.nlm.nih.gov/BLAST).

According to 265rDNA sequence analysis, the JD was identified to be in trichoderma, and a sequence of the JD was shown as SEQ ID No.2; and by means of similarity comparison, a matching degree with trichoderma longibrachiatum reached 100%, such that the JD was identified as the trichoderma longibrachiatum. The LM was identified to be in aspergillus, and a sequence of the LM was shown as SEQ ID No.3; and by means of similarity comparison, a matching degree with aspergillus aculeatus reached 100%, such that the LM was identified as the aspergillus aculeatus. The DHZ was identified to be in the aspergillus, and a sequence of the DHZ was shown as SEQ ID No.4; and by means of similarity comparison, a matching degree with aspergillus fumigatus reached 100%, such that the DHZ was identified as the aspergillus fumigatus.

Embodiment 2 Screening of lactobacilli inhibiting spoilage bacteria

The trichoderma longibrachiatum {JD}, the aspergillus aculeatus (LM), and the aspergillus fumigatus (DHZ) were considered to be the main contaminating bacteria for yogurt spoilage.

In the embodiments of the present application, a double layer plate method was used; common spoilage fungi (the trichoderma longibrachiatum {JD}, the aspergillus aculeatus (LM), and the aspergillus fumigatus (DHZ) that were separated and purified in Embodiment 1) in dairy products and purchased fungal standard strains (fungal standard strains which were purchased from the China Center of Industrial Culture Collection: penicillium sp {CICC2515) and aspergillus niger (CICC2487)) were used as indicator bacteria; and then lactobacilli with potential anti-fungal properties were screened. Colonial morphology of the trichoderma longibrachiatum (JD), the aspergillus aculeatus (LM), the aspergillus fumigatus (DHZ), the penicillium sp (CICC2515), and the aspergillus niger (CICC2487} in a PDA plate was shown in

Fig. 1. {1} Separation and purification of lactobacilli

Home-made traditionally-fermented yak yogurt from herdsmen in Hongyuan County, Aba

Tibetan and Qiang Autonomous Prefecture, Sichuan Province: the home-made Qula from the herdsmen was taken by using a sterile spoon, and was put into a 15 mL sterile capped centrifuge tube containing a proper amount of sterile calcium carbonate and soluble starch {a ratio of the calcium carbonate to the soluble starch being 1:1). The tube was screwed up after the mixture was uniformly stirred, then put into a freezer, and transported back to a laboratory for immediate purification and separation of lactobacilli. Under an aseptic condition, 1 mL of a sample was pipetted to 9 mL of sterile saline, and a sample diluent of 10? was obtained after the mixture was well mixed by means of vortex. Then 10-fold gradient dilution was performed to 107. 100 uL of diluent at 10%, 10%, or 107 was selected, and uniformly spread on an MRS plate for inverted culture for 48h at 37°C. After culture was completed, colonial morphology on the MRS plate was observed. Typical colonies of lactobacilli were selected to purify strains by using a streak plate method, and this streaking operation was repeated until the purified strains were obtained.

Single colonies were formed in the MRS solid culture medium after the strains were purified; and the colonies were small, circular, translucent, raised and slightly white, and had rough surfaces and irregular edges, as shown in Fig. 2. {2} Observation of morphological structures of lactobacilli inhibiting molds

The purified strains were inoculated in a 5 mL sterile MRS broth for culture for 18h at 37°C. 1 mL of bacterial fluid was taken and centrifuged for 1 min at 12000 r/min. After the bacterial fluid was washed for two times with sterile saline, isochoric sterile saline was then added to resuspend the bacteria. A small amount of the bacteria was taken and spread uniformly on a glass slide by using an inoculating loop. After fixation, Gram staining, microscopic examination, and photographing were performed. Gram-positive bacteria (G+) cells stained were blue-purple, and Gram-negative bacteria (G-) cells were red. Cell morphology and Gram staining results were observed and recorded.

After gram staining, the cells were all purple under a microscope, such that the strains were determined as gram-positive bacteria (G+), with rod-like shapes, as shown in Fig. 3 {gray processing was performed in Fig. 3, and an original drawing was purple).

The morphology of the strains conformed to features of a lactobacillus, and morphological structures were uniform, such that it indicated that the strains were pure. (3) PCR amplification of 16S rDNA sequences of lactobacilli inhibiting fungi

PCR amplification was performed by using a 25 pul of a reaction system, including 1 pL of a template, 1 uL of primers (bacterial universal primer, 27F (TACGGYTACCTTGTTACGACTTSEQ

ID NO.7), 1492R{ AGAGTTTGATCMTGGCTCAGSEQ ID NO.8)) {10 uM) each, and 12.5 pL of 2xTaq PCR Master Mix, and making up to 25 ul with sterile ultrapure water. PCR amplification conditions included: performing pre-denaturation at 94°C for 5 min, performing denaturation at 94°C for 30 s, performing annealing at 55°C for 30 s, and performing extension at 72°C for 1 min, there being a total of 35 cycles; and performing end extension at 72°C for 10 min. After sequence amplification, Sangon Biotech (Shanghai) Co., Ltd. is entrusted to sequence qualified

PCR amplification products. After the sequence is obtained, searching and similarity comparison are performed in GeneBank by using BLAST(http://www.ncbi.nim.nih.gov/BLAST).

Results showed that, the TY-F15 was a lactobacillus paracasei strain, the lactobacillus, that is, a sequence of a 16S rDNA gene amplification product of the lactobacillus paracasei TY-F15 {hereinafter also referred to as TY-F15) was shown as SEQ ID No.1. (4) Screening of lactobacilli inhibiting molds

The double layer plate method was used; 15 mL of the MRS solid culture medium at about 45°C was uniformly poured in a plate as a bottom layer; after complete solidification, an inoculating needle wet with the purified lactobacillus liquid was used to draw two horizontal lines with a length being 2.5 cm and a width being 1.5 cm in the center of a culture dish; and inverted culture was performed for 24h in a 37°C constant temperature incubator. 10 mL of a

PDA semi-solid culture medium containing an indicator bacterial spore solution with a concentration being 10% was uniformly spread at an upper part of the MRS solid culture medium as an upper layer; and after the PDA semi-solid culture medium was completely solidified, culture was performed for 48h at 25°C, with three parallel controls per group, and the consistency of bacteriostatic circles among the parallel controls was observed. The growth of the indicator bacteria around the lactobacilli was observed on a regular basis, and the sizes of the bacteriostatic circles were recorded and photographed for the culture dishes where the bacteriostatic circles appeared.

According to the above method, the penicillium sp, the aspergillus niger, the JD, the LM and the DHZ were used as the indicator bacteria, and strains, which was the lactobacillus paracasei TY-F15, having the optimal antibacterial effect were screened from a strain bank (including the purified lactobacillus paracasei TY-F15) owned by Tianyou Dairy. Results were shown in Fig. 4 and Table 1.

Table 1 Size of bacteriostatic circle of TY-F15 against molds

I CC) Co CL

From Fig. 4 and Table 1, it may be learned that, the lactobacillus paracasei TY-F15 had the most obvious antibacterial effect on the penicillium sp. It may be seen that, in the presence of the lactobacillus paracasei TY-F15, the growth of the indicator bacteria was significantly inhibited, the indicator bacteria did not grow or the growth of the indicator bacteria was significantly reduced.

Embodiment 3 Verification of TY-F15 for inhibiting fungi in yogurt

In the embodiments of the present application, a commercial starter used was a lactobacillus bulgaricus and a streptococcus thermophilus, and was purchased from Danisco (China). In the embodiments of the present application, a lactobacillus paracasei TY-F15 with antifungal potential that was obtained by means of screening in Embodiment 2 was added in a yogurt as an auxiliary starter for fermentation, and a mold spore solution was dropped on the surface of the yogurt to observe the time when the yogurt was spoiled. (1) Yogurt making

A yogurt starter was divided into 2 groups, which were respectively a group in which only the commercial starter was added, and a group in which the commercial starter and the

TY-F15 were simultaneously added; after activation of two generations of the TY-F15, washing was performed with sterile saline for two times, resuspending was performed in skim milk, and the final addition (a final concentration in the yogurt) of the strain was 1x10°CFU/mL; skimmed reconstituted milk {the addition of defatted milk powder being 120g/L, and the addition of sugar being 60g/L) was heated to 95°C in a water bath, after sterilization was performed for 10-15 min, a temperature was cooled to about 42°C, and then the two starter groups were inoculated; and fermentation was performed at a constant temperature of 42°C until complete coagulation, then the starters were placed at 4°C for cold storage and after-ripening, and the texture and fermentation of the yogurt were observed. (2} Verification measurement of lactobacillus yogurt inhibiting fungi

The fungal spore solution (JD, LM and DHZ) with the concentration being 10% prepared in

Embodiment 1 and the fungal standard strains were respectively used as indicator bacteria, uniformly dropped on the surface of the prepared yogurt, and placed at room temperature conditions of 10°C and 25°C respectively for spoiling time observation; there were 3 portions for all samples; mold development and the number of molds on the surface of the yogurt were observed every 48h; and the indicator bacterial spore solution was dropped on the surface of the yogurt group only containing the commercial starter as a control group, and the group in which the commercial starter and the TY-F15 were added served as a test group.

Since microbial spoilage was a microscopic phenomenon that could not be observed with the naked eye at the beginning, and since the shelf life of the yogurt was measured in days, an observation time point set here was 48h.

Mold development effects of different indicator bacteria in the yogurt at 10°C were shown in Fig, 5a, Fig. 5b, Fig. 5¢, Fig. 5d and Fig. 5e; and mold development time of the yogurt at 10°C was shown in Table 2.

Table 2 Mold development of yogurt at 10°C

Fp op A A A Js

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6 [3 jo [B 19 Je jo [B 9 [9 0

B [3 jo [B jo 2 jo [B jor 10 wm 3 0 13 jo [2 jo [B jo 2 [9 2 | jo B It B 0 Bo 2 0 2 3 jo [B Dt [B jo Bo Bo x 3 0 13 |B 13 jo B 2 3° 2 3 jo Bp 3 3 jo B23 0 2 jo pF Bo [B 2 B 0 2 3 0 fr |B 13 jo B 2 13 [9

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EEE

Extension | >28 = 10 x 22 = 16 = 18 ow

In Table 2, = is because the samples may start to develop molds when there was 1 sample, or 2 samples, or 3 samples, such that there was certain difference in specific mold development time.

From Fig. 5 and Table 2, it may be learned that, the penicillium sp began to appear in the yogurt at Day 14; a mold development rate reached 100% at Day 16; and after the TY-F15 was added, no sample was molded, indicating that the TY-F15 had an obvious inhibiting effect on the penicillium. The aspergillus niger began to appear in the yogurt at Day 12; the 3 samples were completely molded at Day 14; and after the TY-F15 was added, mold development began to appear in the yogurt at Day 22, and the 3 samples were completely molded until Day 26, indicating that the TY-F15 still had a certain inhibiting effect on the aspergillus niger. The

LM began to appear in the yogurt at Day 18; the 3 samples were completely molded at Day 22; and after the TY-F15 was added, the yogurt was molded at Day 40. The DHZ began to appear in the yogurt at Day 10, and the 3 samples were completely molded at Day 12; and after the

TY-F15 was added, mold development began to appear in the yogurt at Day 26. The JD began to appear in the yogurt at Day 18, and the 3 samples were completely molded at Day 24; and after the TY-F15 was added, mold development began to appear in the yogurt at Day 36.

The above results showed that, the TY-F15 had obvious inhibiting effects on the penicillium, the aspergillus niger, the aspergillus aculeatus (LM), the aspergillus fumigatus (DHZ), and the trichoderma longibrachiatum {JD} in the yogurt at 10°C, and had the optimal inhibiting effect on the penicillium sp and the aspergillus aculeatus (LM). It indicated that if the yogurt contained the penicillium sp, the aspergillus niger, the aspergillus aculeatus, the trichoderma longibrachiatum and the aspergitlus fumigatus, the storage period at 10°C may also be prolonged by about 10 days; if the yogurt did not contain the aspergillus niger in the above 5 molds, the storage period at 10°C may be prolonged by about 16 days; if the yogurt did not contain the aspergillus niger and the aspergillus fumigatus in the above 5 molds, the storage period at 10°C may be prolonged by about 18 days; if the yogurt did not contain the aspergillus niger, the aspergillus fumigatus and the trichoderma longibrachiatum in the above 5 molds, the storage period at 10°C may be prolonged by more than 22 days; and if the yogurt did not contain the aspergillus niger, the aspergillus aculeatus, the aspergillus fumigatus, and the trichoderma longibrachiatum, the storage period at 10°C may be prolonged by more than 28 days. That is, according to the fungal strains carried in the yogurt, the storage period of the yogurt added with the TY-F15 at 10°C is prolonged by more than 10d-28d. it is further to be noted that, the proliferation rate of the 5 molds was inhibited by low temperatures, and if the temperature is lower, the proliferation rate is slower, such that the proliferation rate of the 5 molds below 10°C was slower than that at 10°C, and the storage time of the fermented milk was longer.

In addition, mold development effects of different indicator bacteria in the yogurt at 25°C were shown in Fig. 6a, Fig. 6b, Fig. 6¢, Fig. 6d and Fig. 6e; and mold development results of the yogurt at 25°C were shown in Table 3.

Table 3 Mold development of yogurt at 25°C

EO A CN A A CN CC CI

EO EL EA A A EC re

FF EEE Ene Ee

FEE TE fe FEE Je

EE

Extension | = 8 = 2 = 6 = 4

Ee

In Table 3, = is because the samples may start to develop molds when there was 1 sample, or 2 samples, or 3 samples, such that there was certain difference in specific mold development time.

From Fig. 6 and Table 3, it may be learned that, the penicillium sp, the aspergillus niger, the LM, the JD and the DHZ all began to appear in the yogurt samples at Day 4, and some samples were completely molded; after TY-F15 was added, the initial mold development time of the penicillium sp was prolonged to Day 12, and the samples were completely molded at

Day 16; the initial mold development time of the aspergillus niger was prolonged to Day 6, , and the samples were completely molded; the initial mold development time of the LM was prolonged to Day 10, and the samples were completely molded at Day 14; the initial mold development time of the DHZ was prolonged to Day 8, and the samples were completely molded at Day 10; and the initial mold development time of the JD was prolonged to Day 10, and the samples were not completely molded after 42 days.

The above results showed that, the TY-F15 still had the obvious inhibiting effects on the penicillium, the aspergillus niger, the trichoderma longibrachiatum (ID), the aspergillus aculeatus (LM), and the aspergillus fumigatus (DHZ) in the yogurt at room temperature of 25°C, and had the optimal inhibiting effect on the penicillium, the aspergillus aculeatus and the trichoderma longibrachiatum. it indicated that if the yogurt contained all the 5 molds, the storage period at 25°C may also be prolonged by about 2 days; if the yogurt did not contain the aspergillus niger in the above 5 molds, the storage period at 25°C may be prolonged by about 4 days; if the yogurt did not contain the aspergillus niger and the aspergillus fumigatus in the above 5 molds, the storage period at 25°C may be prolonged by about 6 days; and if the yogurt did not contain the aspergillus niger, the aspergillus aculeatus, the aspergillus fumigatus and the trichoderma longibrachiatum in the above 5 molds, the storage period at 25°C may be prolonged by about 8 days. That is, according to the fungal strains carried in the yogurt, the storage period of the yogurt added with the TY-F15 at 25°C may be prolonged by more than 2d-8d. it is further to be noted that, the proliferation rate of the 5 molds was inhibited by low temperatures, and if the temperature is lower, the proliferation rate is slower, such that the proliferation rate of the 5 molds below 25°C was slower than that at 25°C, and the storage time of the fermented milk was longer.

To sum up, the TY-F15 had the inhibiting effects on the penicillium, the aspergillus niger, the aspergillus aculeatus (LM), the trichoderma longibrachiatum (JD), and the aspergillus fumigatus (DHZ) in the yogurt stored below 10°C and below 25°C, had a certain effect on prolonging the shelf life of the yogurt, and had the potential of becoming the auxiliary starter for long shelf life yogurt.

Embodiment 4 Impact of TY-F15 on quality of yogurt

In the embodiments of the present application, a commercial starter (a lactobacillus bulgaricus and a streptococcus thermophilus) was purchased from Danisco (China). In the embodiments of the present application, the impact of physical and chemical properties and sensory quality of a fermented yogurt by auxiliary bacteria was studied by using the TY-F15 as an auxiliary starter and the commercial starter as a main starter.

The TY-F15 was inoculated in the optimized liquid culture medium according to a 5% inoculation amount and cultured at 37°C for 18h; after activation for three generations, 2 mL was taken and centrifuged for 15 min at 10000 r/min, so as to collect a bacterial precipitate; and the bacterial precipitate was washed for 2 times with a sterile PBS solution, and resuspended in isochoric sterile saline, so as to obtain bacterial suspension. In one group, the well prepared bacterial suspension and 6 g/L of the commercial starter were simultaneously added into HUT sterilized milk according to 105CFU/L; in the other group, 6 g/L of the commercial starter was added into the HUT sterilized milk as a control group; and fermentation was performed on two groups for 7h at 42°C simultaneously, 7 days were taken as a time mode after cold storage and after-ripening, and the state and quality of the yogurt in the two groups were recorded and compared. (1) Impact of TY-F15 on appearance of yogurt

Appearance of the yogurt fermented by using the TY-F15 as the auxiliary starter and after 1 day of cold storage and after-ripening in the control group was shown in Fig. 7. The yogurt fermented by using the TY-F15 as the auxiliary starter is smooth in surface, uniform in color, good in tissue state, and has no peculiar smell, while the yogurt in the control group is slightly rough and non-smooth in surface. Therefore, as the auxiliary starter, the TY-F15 has a positive effect on improving the product quality of the yogurt. (2) impact of TY-F15 on fermentation time of yogurt.

In the embodiments of the present application, a CINAC dairy fermentation monitoring instrument was used to track and record changes in the pH of the above yogurt fermentation process. A dynamic change curve of the pH with fermentation time was shown in Figure 8.

The auxiliary inoculation of the TY-F15 does not significantly affect the fermentation time of the yogurt, because the TY-F15 proliferates slowly and does not affect a fermentation process of the fermented mitk during fermentation. (3) impact of TY-F15 on pH value and titratable acidity of yogurt during storage.

In the embodiments of the present application, measurement of the pH of the yogurt during yogurt storage was performed by using a precise pH meter; and titratable acidity during yogurt storage is measured with reference to GB 5009.239-2016 "National Food Safety

Standard for Measurement of Food Acidity” (at the end of yogurt fermentation, lactobacilli continued to convert lactose to lactic acid, changing the pH and acidity of the yogurt). Results were shown in Fig. 9. The TY-F15 had the impact on the acidity of the yogurt, and can delay the post-acidification of the yogurt to a certain extent at the later storage period. The lactobacillus bulgaricus was the main strain causing the post-acidification of the yogurt, and the TY-F15 proliferated as a dominant bacterium in the yogurt during the late storage period, inhibiting the growth of the lactobacillus bulgaricus, such that the post-acidification of the yogurt was alleviated to a certain extent, which had a positive effect on prolonging the shelf life of the yogurt.

(4) Impact of TY-F15 on stability of yogurt during storage.

In the embodiments of the present application, measurement of Water Holding Capacity (WHC) of the yogurt was performed according to the following method. 20 g of a sample was accurately weighed, centrifugation was performed for 20 min at 4000 r/min, the mass of a precipitate was weighed, and WHC was calculated according to an equation {1).

WHC 2% =(1- Mass of precipitate 1001)

Mass of samples

In the embodiments of the present application, measurement of the stability of the yogurt was performed according to the following method. An optical analysis centrifugal machine (LUMi Sizer L.U.M.GmbH, Germany) was used for measurement, and about 0.4mL of

AMD was filled in a standard reaction cup. Measured instrument parameters were set as follows: a temperature was 25°C; a rotary speed was 4000 rpm; a time interval was 10 seconds; experimental period was 4490s; and each experiment was made in triplicate, and each result was expressed as clarification index average result t standard deviation.

In the embodiments of the present application, the texture of the yogurt was measured according to the following method. The method included: measuring the texture of a yogurt by using a texture analyzer (TA XTplus, Micro Stable System Co.,UK); performing a penetration test by using a cylindrical probe with a diameter being 10 mm; immediately performing measurement after the samples were taken from a refrigerator (4°C); and penetrating 15 mm in the yogurt samples at a speed of 1mm/s and a trigger force of 1 g.

Changes {as a living microorganism, the addition of the lactobacilli to the yogurt may disrupt the steady state of a yogurt system, causing serious problems of whey precipitation or poor texture, which were caused by deterioration of the stability of the yogurt) in the WHC and clarification index of the yogurt at 4°C during storage were detected , and results were shown in Fig. 10. The activity of the TY-F15 in the early stage of storage, such that the stability of the yogurt is not significantly affected; and the TY-F15 proliferated as a dominant bacterium in the yogurt in the later stage of storage, the WHC of the group added with the TY-F15 gradually increased, and the clarification index (transmittance) gradually reduced, such that the stability of products added with the TY-F15 is significantly improved.

Changes in the texture of the yogurt added with the TY-F15 at 4°C during storage were shown in Fig. 11. The hardness, viscosity, elasticity, cohesion, adhesiveness, and chewiness of the yogurt were improved to a certain extent by adding the TY-F15 in the later stage of storage.

To sum up, the stability of a yogurt system may be improved by adding the TY-F15. (5) Measurement of impact of TY-F15 on sensory quality of yogurt

Sensory quality was the most important criterion for consumers to accept or reject food.

Food sensory evaluation, also known as food sensory analysis, was an inspection and analysis process by using scientific analysis methods to sense food characteristics or properties by means of vision, smell, taste, and hearing, and performing qualitative and quantitative analysis of food. The metabolism of the probiotic lactobacilli produced different products that give the yogurt certain special flavors, directly affecting consumer acceptance and purchase intention, such that the metabolism of the probiotic lactobacilli was an important factor affecting quality of the products.

In the embodiments of the present application, a 9-point scaling method was used, 200 sensory evaluators (100 men and 100 women) scored the appearance, color, aroma, texture, taste and overall acceptance of the yogurt samples prepared in Embodiment 1 respectively on the basis of their habits and interests for consuming the yogurt, with a total score of 9 {1=very dislike; 3=dislike; 5=neither like nor dislike; 7=like; 9=very like); and each result was expressed as average result t standard deviation. Results were shown in Table 4.

Table 4 Sensory evaluation table

As shown in Table 4, results showed that, the appearance, texture, and taste of a yogurt product added with the TY-F15 were obviously improved, and the scores for overall acceptance were slightly higher than those in the control group, but none of them were significantly different (P>0.05). Most people believed that the yogurt products with TY-F15 had a better appearance and texture, with a special fermented flavor, indicating that the

TY-F15 had a certain improvement effect on the product quality of the yogurt, and had higher degree of consumer acceptance.

It is finally to be noted that, the above embodiments are merely for describing and not intended to limit the technical solutions of the present application. Although the present application is described in detail with reference to the preferred embodiments, those of ordinary skill in the art should understand that the technical solutions of the present application may be modified or equivalently replaced without departing from the purpose and scope of the technical solutions of the present application, and shall all fall within the scope defined by the claims of the present application.

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ER <INSDQualifier valus>genomic DNA</INSDQuali fier valus> 26 </INSDOualifier> 37 <INSDOQualifier id="q8"> 38 <IN3DQualifier namevorganism“/INSDQualifier name> 38 <INSDgvelifier wvalue>Aspergillus fumigatus </INgDQualifier value» 100 </INSDQualifier>

LOL </INSDFeaturs quals>

LOE </IN3DFeature> 103 “/INSDSeqg fesature-table> ijd <iNSDSeq sequence» tgggtccaacctcccaccegtgtctatcgtaccttgttgettcggcgggccecgeegtttcgacggccgcecgggg aggcettgecgccccegggccegegcccgcecgaagaccccaacatgaacgctgttctgaaagtatgcagtctgag ttgattatcgtaatcagttaaaactttcaacaacggatctcttggttccggcatcgatgaagaacgcagcgaaa tgcgataagtaatgtgaattgcagaattcagtgaatcatcgagtectttgaacgcacattgegccccctggtatt ccggggggcatgcctgtccgagegtcattgetgccctcaagcacggcttgtgtgttgggccccegtcecccetct cccgggggacgggcccgaaaggcagcggecggcaccgegtceggtcctecgagegtatggggetttgtcacctget ctgtaggcccggcecggecgccagccgacacccaactttatttttctaaggttgacctcggat </INSDSeg sequence> 108 </INSDSeg> 10a </SeguenceDatas

LO <SequenceData sequenceIDNumber="5n>

LO5 <INSDSeqd»>

LOG <INSDSeq length>19</INSD5eq length» iiD <INSDSeq moltype>DNA</INSDSeg moltype> ill <INSDSeq divislion»PAT</INSDSeqg division» iië <INSDSeg Iearturertabie» iia <INSDFsature> 114 <IN3DFeature key>source</IN3DFeature key>

Lijs <IN3DFeature location»l..19</INSDFeaturs location ie <INSDFsature qualsg>

LAT <INSDuuelifier>

LLS <INSDoualifier name>mol type“ /INSDQualifier name>

LLS <INSDQualifisr value>other DNA</INSDQualifier valuer 1E0 </INSDOualifier> iz <INSDOualifier id="gqgi0r> 122 <IN3DQualifier name>organism</INSDQualifisr name> 123 <INSDQualifiler valuersynthetic construct </INSDDualifier valued 124 </INSDOualifier>

Las </THSDFeaturs guals> 126 </INSDFeatures 127 </IN3DSeqg feature-table> ief <INSDSeq sequence>teegtaggtgaacetgegg-/INSD3eg sequences 12% </TNSDSeg> 130 </SeguenceDaia>

Lj <Sequencebata zedquencelDNumser="&N>

LS <INSDSeq> 133 <INSDSeq length>20</INSDSeq lengths 134 <INSDSeq moltype>DNA</INSDSea moliype> ijs <INSDSeq division»PAT</INSD3ea division” 138 <INSDSeq feature-table>

Lat <INSDFeature> 138 <IN3DFeature key>source</IN3DFeature key»

Lis <INSDFeature location»l1..20</INSDFeature location»

TAL <INSDFeature quals> u 141 <INSDOualifiers

LAY <INSDQualifier name>mol type</INSDQualifier name> 142 <INSDOualifier wvalue>other DNA</INSDCualifier value» 144 </INSDQualifier> u 145 <INSDOualifier id="gien> 14a <IN3DQualifier namerorganism“/INSDQuali fier name> 4 <INSDOualifier valuersynthetic construct </INSDQualifier value» 14% </INSDOualifier> 140 </INSDFeaturs guals> 1540 </TNSDFeaturer 151 </INBDSeq feature-table> i582 ZINSDSeq sequenas>tecteegettattgatatge</IN3D3ey sequenae> 153 </INSDSeq> 154 </Zequencelata> 1a <Sequencebata seguenasibNumbar=snits 156 <INSDieg> 137 <INSDSeq length>22</IN3DSeq length» 158 <INSDSeq moltype>DNA</INSDSeq moltype> 153 <IN3DSeq division»PAT</INSD3eq division»

Lan <INSDSeq fearure-tabier iel <INSDFeabture>

Le: <INSDFeature key>source“/INSDFeature key>

Les <INSDFeature location>l..22</iN3DFeature lozation>

184 <INSDFeaturs qualsy 165 <INSDQualifier»> aa <INSDQualifier name>mol type</iNSDQualifier name> ia <INSDuuelifier value>other DNA</INSDOualifier valued 148 </INSDQualifier> u ies <INSDQualifier id="gidnx>

LEO <INSDQualifier namerorganism</INSDQualiifier name>

EL <INSDQualifier valuersynthetic construct </INSDguali fier values»

LTE </INSDQualifiers> 172 </IN3DFeature gualsd ijd </INSDFeature> u 17h </INSDSeg features table»

Lie <INSDSeg seguence>rtacggytaccttgttacgactt</INSDS=4 zeguence>

LT </INSDSeg>

LG </SequenceData>

Lijn <SequernceData sequenceliNuec="S"> 1840 <INSDSeqg> 18d <INSDSeq length>20</INSDSeq length> 182 <INSDSeq moltype>DNA-/INSDSeg moltype> i183 ZINSDSeq division>PAT</INSDSeq division» 184 <INSDSeq feabure-table> ign <INSDfeasture>

LEE <INSDFeature key>source</INSDFeature key>

HS <INSDFeature location>l..20</INSDFeature locations iës <INSDFealurse guals> 153 <INSDOQualifier» 150 <IN3DQualifier namedmol type</INSDQualifisr name> 151 <INSDUvaelifier valuerother DNA</INSDGualifier value> ien </INSDQualifier» Ĳ 1893 <INSDguelifier id="gig*>

RSE <INSDOQualifier namerorganism</INSDQualifier name>

Les <INSDOualifier value>synthetic construct </INSDQuali fier value> 184 </INSDOualifier> 137 </IN3DFeature guala> ian </INSDFeaturs> u 18% </INSDSegy featurs-table> 200 <INSDSeq sequence>agagtttgatemtggetecag</INZlSeq sequence

ZOL </INSDSeg>

LQ </SequenceData> 203 </8TZ2eSaguenceld sting

Claims

CONCLUSIONS

1. Lactobacillus paracasei (Lactobacillus paracasei) TY-F15 to which the collection number CGMCC No. 25742 was awarded by the General Microbiological Center of the Chinese Microbiological Culture Collection Management Committee.

Lactobacillus paracasei TY-F15 according to claim 1, characterized in that a 16S rDNA sequence as sequence SEQ ID No. 1 is shown.

3. Composition, characterized in that the composition contains one or a combination of a plurality of the following substances: (a) a living lactobacillus paracasei TY-F15 according to claim 1 or 2; (b) a lysate of the lactobacillus paracasei TY-F15 according to claim 1 or 2; {c} a culture of the lactobacillus paracasei TY-F15 according to claim 1 or 2; and (d} a fermentation broth of the lactobacillus paracasei TY-F15 according to claim 1 or 2.

4. Composition according to claim 3, characterized in that the composition further contains one or a combination of probiotics, prebiotics, dietary fiber and traditional Chinese medicine.

5. Preparation, characterized in that the preparation contains the lactobacillus paracasei TY-F15 according to claim 1 or 2 or the composition according to claim 3 or 4, and a carrier, wherein the carrier is a medicinal carrier or an edible carrier.

Preparation according to claim 5, characterized in that the preparation is tablets, pills, capsules, powders, gels, granules or liquids.

7. Use of lactobacillus paracasei TY-F15 in rot-resistant fungi according to claim 102.

Use according to claim 7, characterized in that the rot-resistant fungi are one or more of penicillium {Penicillium}, aspergillus niger (Aspergillus niger), trichoderma longibrachiatum (Trichoderma longibrachiatum}, aspergillus aculeatus {Aspergillus aculeatus), and aspergillus fumigatus (Aspergillus fumigatus).

Use according to one of claims 7 or 8, characterized in that the use contains one of the following: (a) a use of the lactobacillus paracasei TY-F15 as a fermentation strain in the preparation of fermented milk; (b) an application of the lactobacillus paracasei TY-F15 in combination with another fermentation strain in the preparation of fermented milk as a fermentation strain; (c) a use of the live actobacillus paracasei TY-F15 or the metabolites of the lactobacillus paracasei TY-F15 as additives for pharmaceutical products or foodstuffs.

Use according to claim 9, characterized in that the other fermentation strain contains one or combinations of lactobacillus bulgaricus (Lactobacillus bulgaricus) and philptoctococcus thermophilus (Streptococcus thermophilus).