An ultrasonic resonance targeting sterilization method and system based on microbial structure fingerprint

Abstract

The application discloses an ultrasonic resonance targeted sterilization method and system based on microbial structure fingerprint. The method collects individual structure characteristic parameter set of target microorganisms, calculates inherent mechanical resonance frequency based on a thin shell resonance model, and generates frequency band grouping and strategy based on the same. The method applies ultrasonic waves with frequency matching and acoustic pressure of 0.05-0.4 MPa to liquid food, so that resonance rupture of microbial cell walls or envelopes occurs. The method adopts coarse channel and thin film gap double-mode channel to adapt acoustic penetration characteristics of different frequency bands, and configures a frequency band division transducer array, an acoustic matching layer, cavitation suppression and frequency self-locking control. The energy consumption of full-band continuous flow processing is about 55%-90% of that of pasteurization, the temperature rise is less than 5 DEG C, and the immunoglobulin G retention rate is greater than 80%. The system comprises a fingerprint database, a frequency calculation module, an ultrasonic processing and control module, can realize GHz frequency band targeted removal of phages, and supports a combined process in series with ceramic membrane microfiltration.

Description

Technical Field

[0001] This invention belongs to the field of food science and technology, specifically relating to an ultrasonic resonance targeted sterilization method based on the structural characteristic parameters of individual microorganisms, and the method and equipment for implementing the system, which is particularly suitable for selective microbial control processing of liquid dairy products. Background Technology

[0002] Milk is one of the most important nutritional foods in the world, and its safety is directly related to public health. Currently, the global dairy industry generally uses high-temperature short-time pasteurization (HTST, 72°C / 15 seconds) or ultra-high temperature instantaneous sterilization (UHT, 135-150°C / 2-5 seconds) to eliminate pathogenic microorganisms in milk.

[0003] However, the above heat treatment method has the following inherent drawbacks: (1) Severe nutritional damage: Heat treatment causes irreversible denaturation of many bioactive substances in milk. Studies have shown that pasteurization can cause a loss of more than 90% of immunoglobulin G (IgG), a reduction of more than 80% in lactoferrin activity, and varying degrees of heat denaturation of α-lactalbumin and β-lactoglobulin. Vitamin C is lost by about 20-30%, and vitamin B12 is reduced by 10-15%.

[0004] (2) Huge energy consumption: Pasteurization requires heating milk from 4°C to 72°C and maintaining it for 15 seconds, then cooling it back to 4°C. Based on the industrial HTST process including heat recovery, the total energy consumption for the entire process is approximately 252,000 J / L (0.07 kWh / L). Against the backdrop of continuously rising global energy prices and the increasingly urgent goal of carbon neutrality, high energy consumption has become a key factor restricting the sustainable development of the dairy industry.

[0005] (3) Lack of selectivity: Traditional heat treatment cannot distinguish between pathogenic bacteria and beneficial bacteria. In the production of fermented dairy products such as yogurt and cheese, heat treatment kills all lactic acid bacteria (LAB) while eliminating pathogenic bacteria, requiring subsequent re-inoculation of fermentation bacteria, which increases the complexity and cost of the process.

[0006] (4) The problem of bacteriophages cannot be solved: Bacteriophage contamination leading to fermentation failure during dairy fermentation is a century-old problem in the global dairy industry, causing hundreds of millions of dollars in losses every year. Traditional methods are almost ineffective against bacteriophages.

[0007] In recent years, ceramic membrane microfiltration (MF) technology has been introduced into the dairy processing field as a "cold sterilization" solution. This technology utilizes α-alumina (α-Al₂O₃) ceramic membranes with a pore size of 0.8-1.4 μm to physically sieve and trap bacteria and spores at a low temperature of 5-15°C. Combined with subsequent mild heat treatment (60-65°C), it can produce ESL (Extended Shelf Life) milk with a shelf life of 30-45 days. This technology has already been industrialized by equipment manufacturers such as Tetra Pak and SPX Flow.

[0008] However, ceramic microfiltration "cold sterilization" technology has the following inherent limitations: (a) Inability to remove bacteriophages and viruses: Common bacteriophages in dairy fermentation (groups 936, c2, and p335) have a size of 30-80 nm, which is much smaller than the minimum pore size of microfiltration membranes (0.8 μm) (a difference of more than 10,000 times), and MF cannot intercept them at all. Similarly, viruses with a size in the range of 20-300 nm cannot be removed by MF.

[0009] (b) Non-selective: Based on the principle of size sieving, MF indiscriminately retains all microorganisms larger than the pore size, and cannot distinguish between pathogenic bacteria and beneficial bacteria. In fermented milk production, the target lactic acid bacteria are also retained in the retentate, and the fermentation bacteria must be re-inoculated.

[0010] (c) Removal without killing, concentrate requires additional treatment: MF separates microorganisms from the permeate and enriches them in the retentate, which contains a high concentration of viable bacteria (up to 10). 7 -10 8 (CFU / mL) requires additional heat treatment or chemical sterilization for safe discharge.

[0011] (d) Membrane fouling leads to flux decline: Proteins (especially casein) and fats in milk deposit on the membrane surface to form a filter cake layer, which leads to a continuous decline in permeate flux. Frequent CIP (Clean-In-Place) acid-base cycle cleaning is required, which increases equipment investment and operating costs.

[0012] (e) Further heat treatment is still required: MF itself cannot meet commercial sterility requirements and must be combined with subsequent mild pasteurization (60-65°C / 15-20s) or ultrapasteurization of the retentate to meet food safety standards. Therefore, MF is not a true "cold sterilization" solution, but a combination of "cold sterilization + mild heat sterilization", and there is still a loss of heat-sensitive nutrients.

[0013] The comparison of the three core technical indicators is as follows:

[0014] Regarding the sterilization principle: pasteurization (HTST) uses heat-induced death; ceramic microfiltration (MF) + mild heat treatment uses physical sieving removal combined with mild heat treatment; the present invention (ultrasonic resonance targeting) uses frequency-matched resonance destruction.

[0015] Regarding operating temperature: pasteurization is 72°C for 15 seconds; MF+ mild heat treatment is 5-15°C plus 60-65°C for 15 seconds; the present invention is 4 plus or minus 2°C.

[0016] In terms of energy consumption: pasteurization is about 252,000 J / L; MF+ mild heat treatment is about 80,000 J / L; the present invention is described in Example 5 (the design sound intensity is about 55%-90% of the pasteurization value, which is further reduced after threshold optimization).

[0017] Regarding IgG retention rate: pasteurization is less than 10%; MF+ mild heat treatment is about 80%-90%; this invention is higher than 80%.

[0018] Regarding phage clearance capability: Pasteurization can clear phages through heat treatment; MF membranes, with a pore size of 0.8-1.4 micrometers, are much larger than the phage size of 30-80 nanometers and cannot intercept phages; this invention uses GHz-band ultrasound to target and clear phages.

[0019] In terms of selective sterilization: pasteurization is non-selective; MF indiscriminately retains all microorganisms larger than the pore size; this invention can selectively kill microorganisms according to frequency band.

[0020] Regarding membrane fouling and consumables: pasteurization has no membrane fouling problem; MF faces serious membrane fouling and requires frequent CIP cleaning; the transducer of this invention is installed without contact, so there is no consumables fouling problem.

[0021] Regarding the treatment of concentrated waste liquid: pasteurization produces no concentrated waste liquid; MF generates a retrieval concentrate containing a high concentration of live bacteria (10^7-8 CFU / mL), which requires additional treatment; this invention is an in-situ sterilization method, producing no concentrated waste liquid.

[0022] The application of ultrasonic technology in food processing has been extensively studied. In existing technologies, ultrasonic sterilization primarily relies on acoustic cavitation—that is, ultrasonic waves generate tiny bubbles in a liquid, which then collapse instantaneously, releasing localized high temperatures (approximately 5000K), high pressures (approximately 50MPa), and shock waves to destroy microorganisms. This non-selective mechanical destruction method is essentially still a "broad-spectrum, violent sterilization" approach, unable to distinguish between pathogenic and beneficial bacteria, and exhibits extremely low efficiency in killing certain drug-resistant bacteria (such as Staphylococcus aureus).

[0023] Ultrasonic resonance inactivation is a completely different physical mechanism: when the frequency of ultrasound precisely matches the inherent mechanical resonance frequency of the microbial cell wall / capsule, it can induce cell wall resonant rupture at sound pressure levels far below the cavitation threshold, achieving selective killing. However, to achieve targeted resonance inactivation, it is essential to accurately determine the individual resonance frequency of the target microorganism.

[0024] Globally, no research has reported methods for calculating resonance frequencies based on individual microbial structural parameters, nor are there any methods or systems for targeted sterilization of multiple microorganisms across the entire spectrum of milk. The main reasons are: (a) the lack of a systematic database of microbial structural parameters; (b) the lack of reliable physical models from microstructure to macroscopic resonance frequencies; and (c) the lack of engineered multi-band resonance processing schemes.

[0025] Therefore, there is an urgent need in this field for a method and system that can accurately calculate the resonance frequency of microorganisms based on their individual structural characteristics and implement selective ultrasonic resonance targeted sterilization accordingly. Summary of the Invention

[0026] The technical problem to be solved by the present invention is: how to provide a non-thermal, low-energy, selective milk sterilization method and system that can effectively eliminate pathogenic microorganisms while retaining the bioactive nutrients of milk to the greatest extent, and can selectively retain or remove specific microbial groups according to processing needs.

[0027] To address the aforementioned technical problems, this invention provides an ultrasonic resonance-based targeted sterilization method based on microbial structural fingerprints, comprising the following steps: Step S1: Obtaining microbial structural parameters For each microorganism in the target microbial community, obtain its individual structural feature parameter set, which includes at least: (a) Cell geometric parameters: equivalent radius R, cell shape factor (sphere / rod / filament); (b) Cell wall / membrane parameters: wall thickness h, number of wall layers, elastic modulus E of each layer, Poisson's ratio ν; (c) Outer membrane characteristic parameters (Gram-negative bacteria): outer membrane integrity index, lipopolysaccharide layer thickness, and porin density; (d) Special structural parameters: presence of spores and number of shell layers, capsid symmetry (for bacteriophages), and capsule thickness.

[0028] Preferably, the set of structural feature parameters constitutes a unique structural fingerprint vector for the microorganism; the dimensions of the fingerprint vector and the specific meaning of each dimension can be adjusted according to the microorganism category and processing requirements, and are not limited to the dimensions and number listed in the embodiments.

[0029] Step S2: Resonance Frequency Calculation Based on the thin-shell resonance physical model, the intrinsic mechanical resonance frequency of each microorganism is calculated according to the individual structural feature parameter set obtained in step S1. The formula for calculating the respiratory modal resonance frequency of the thin-shell resonance model is as follows: f = (1 / 2πR) · √(E / ρ) · √(2h / (R·(1-ν))) Where: R is the equivalent radius, E is the wall elastic modulus, ρ is the wall material density, h is the wall thickness, and ν is Poisson's ratio.

[0030] For microorganisms with multi-layered wall structures, the equivalent elastic modulus method is used for calculation: E_eff = Σ(Eᵢ · hᵢ) / Σ(hᵢ) Where Eᵢ and hᵢ are the elastic modulus and thickness of the i-th layer wall, respectively.

[0031] For Gram-negative bacteria, an outer membrane correction factor γ_outer is introduced: f_G- = f_base · γ_outer The value of γ_outer is determined by the outer membrane integrity index, the lipopolysaccharide layer thickness, and the porin density, and ranges from 0.6 to 1.0.

[0032] For bacteriophages, a protein capsid ultrathin shell model is adopted, and a capsid symmetry correction factor σ_capsid is introduced: f_phage = (1 / 2πR) · √(E_capsid / ρ_capsid) · √(2h / (R·(1-ν)))· σ_capsid The value of σ_capsid is determined by the type of capsid symmetry: 1.0 for icosahedral symmetry, 0.85 for spiral symmetry, and 0.72 for compound symmetry.

[0033] Step S3: Frequency Band Grouping and Processing Strategy Generation Based on the resonance frequencies of each microorganism calculated in step S2, the target microbial community is stratified and grouped according to frequency: - Low frequency band (50-200 kHz): Gram-negative thin-walled bacteria; - Mid-frequency band (200-500 kHz): Gram-positive bacilli and spores; - High frequency band (1-5 MHz): Gram-positive cocci and some small eukaryotic microorganisms; - Ultra-high frequency band (5-10 MHz): Yeast (of which Saccharomyces cerevisiae is about 3.4 MHz, located at the upper end of the high frequency band, and can be incorporated into the ultra-high frequency band for unified processing during implementation). - Microwave band (2-7 GHz): bacteriophage.

[0034] Based on the frequency resolution Δf of each group of microorganisms, determine whether the selectivity condition is met: Δf = |f_target - f_protected| / f_target × 100% Under normal operating conditions, when the physical quality factor Q≈5, the full width at half maximum (FWHM) of the resonance peak is approximately 20%, and the minimum resolvable frequency separation is approximately 20%. When the high-Q narrowband frequency locking method (frequency fine-tuning accuracy ±0.1%) described in Embodiment 11 of this invention is used, the effective resolution can be improved to approximately 5%. Accordingly: When Δf ≥ 5% and frequency locking is enabled, a targeted frequency can be applied directly to achieve selective killing. When Δf < 5%, a time-sequential separation strategy is adopted—first, broad-spectrum resonance is applied to kill the target bacteria, and after cooling, the beneficial bacteria that need to be protected are inoculated for subsequent fermentation.

[0035] Step S4: Ultrasonic Resonance Treatment Based on the processing strategy generated in step S3, multi-band ultrasonic resonance treatment is applied to the milk: (a) Sound pressure control: The working sound pressure is set to 0.05-0.4 MPa, preferably 0.1-0.25 MPa, to ensure that it is within the range of the microbial cell wall resonance destruction threshold and lower than the acoustic cavitation threshold of the milk system (the cavitation threshold in milk is about 0.5 MPa). (b) Frequency application: According to the frequency band grouping in step S3, apply ultrasonic waves of the corresponding frequency to each frequency band sequentially or simultaneously; (c) Temperature control: The process is carried out at 0-10°C, preferably 2-6°C, and the temperature rise of the milk during the process does not exceed 5°C.

[0036] Preferably, the ultrasonic resonance treatment in step S4 employs different fluid channel modes based on the microbial resonance frequency: - Coarse channel mode (suitable for microorganisms with resonant frequency ≤1 MHz): Milk flows through a cylindrical processing chamber with a diameter of 3-5 cm, and ultrasound is applied radially from the tube wall for continuous flow processing; - Thin film slit mode (suitable for microorganisms with resonant frequency >1 MHz): Milk flows through a flat slit treatment chamber with a thickness of 0.8-1.5 mm, and ultrasound is applied from both sides of the slit in a multi-stage series treatment.

[0037] The coarse channel mode and the thin film slit mode can be used in series to achieve continuous flow processing of microorganisms across the entire frequency band.

[0038] Step S5: Effect Verification and Feedback The treated milk is subjected to microbial testing, and the actual kill rate is compared with the expected kill rate. If the deviation exceeds the preset threshold, the structural characteristic parameters of the corresponding microorganisms are corrected according to the deviation data, the resonance frequency calculation results are updated, and the treatment parameters are adjusted before reprocessing.

[0039] This invention also provides an ultrasonic resonance targeted sterilization system based on microbial structural fingerprints, comprising: Fingerprint database module: Stores the individual structural feature parameter set of each microorganism in the target microbial community. The structural feature parameter set includes cell geometric parameters, cell wall / membrane parameters, outer membrane feature parameters, and special structural parameters. Frequency calculation module: It has a built-in thin-shell resonance physical model, which is used to calculate the inherent mechanical resonance frequency of each microorganism based on the set of structural feature parameters in the fingerprint database module, as well as to perform frequency separation analysis and frequency band grouping; Ultrasonic processing module: includes a multi-band transducer array and a fluid processing chamber, wherein the fluid processing chamber comprises a coarse channel processing unit (diameter 3-5 cm, suitable for ≤1 MHz frequency band) and a thin film slit processing unit (slit thickness 0.8-1.5 mm, suitable for >1 MHz frequency band). Control module: Communicates with fingerprint database module, frequency calculation module and ultrasonic processing module, and is used to automatically control the working frequency, sound pressure, processing time and temperature parameters of ultrasonic processing module according to frequency calculation results.

[0040] Preferably, the multi-band transducer array includes: - Low-frequency transducer array (50-200 kHz), mounted on the outer wall of the coarse channel processing unit; - Intermediate frequency transducer array (200-500 kHz), mounted on the outer wall of the coarse channel processing unit; - High-frequency transducer array (1-5 MHz), installed on both sides of the thin-film gap treatment unit; - Ultra-high frequency transducer array (5-10 MHz), installed on both sides of the thin film gap treatment unit.

[0041] Preferably, the thin film gap treatment unit is a multi-stage series structure, with each stage having a gap thickness of 0.8-1.5 mm, and a cooling section between stages, with a total of 2-6 stages.

[0042] Preferably, the system further includes a temperature control unit for maintaining the milk temperature within the range of 0-10°C during processing.

[0043] This invention also provides the application of the above-described method and system in dairy product processing, the application including: (1) Cold sterilization of drinking milk: selectively kills pathogenic microorganisms in milk (Escherichia coli O157:H7, Salmonella, Staphylococcus aureus, Listeria monocytogenes, Mycobacterium tuberculosis, etc.), while retaining bioactive components such as IgG, lactoferrin, α-lactalbumin, and β-lactoglobulin to the greatest extent. (2) Selective pretreatment of fermented milk: In the production of fermented dairy products, pathogenic Gram-negative bacteria and spore-forming bacteria are selectively killed, while specific lactic acid bacteria are retained or utilized as the starting microbiota for fermentation. (3) Phage-targeted removal: In the production of fermented dairy products, GHz band ultrasound is used to target and remove contaminated phages, solving the problem of fermentation failure in the dairy industry.

[0044] Compared with the prior art, the present invention has the following beneficial effects: (1) Non-thermal sterilization with near-zero nutrient loss: Based on the physical mechanism of ultrasonic resonance, the working sound pressure (0.05-0.4MPa) is far below the cavitation threshold (0.5 MPa), the temperature rise during the process does not exceed 5°C, the IgG retention rate in milk can reach more than 80% (pasteurization retention rate <10%), and the lactoferrin retention rate >85%, thus achieving "cold sterilization".

[0045] (2) Full-band continuous flow processing with controllable and optimizable energy consumption: The energy consumption of the full-band continuous flow processing of this invention is approximately 55%-90% of that of pasteurization (0.07 kWh / L), depending on the sound intensity optimization level (approximately 0.063 kWh / L when the design sound intensity is 2-3 W / cm², and approximately 0.038 kWh / L when the sound intensity is optimized to near the resonance threshold); it can be further reduced by combining intermittent excitation and narrowband frequency locking. The theoretical minimum acoustic energy for single-band, single-target processing is approximately 6,000 J / L.

[0046] (3) Selective killing: By calculating the structural characteristic parameters, the precise resonance frequency matching of individual microorganisms can be achieved. Pathogenic microorganisms can be selectively killed while beneficial bacteria are retained according to processing requirements, providing a more flexible and precise means of microbial control for products such as fermented milk and probiotic milk.

[0047] (4) Phage solution: The method of using GHz band ultrasound to target and remove phages was proposed for the first time, providing a new idea for solving the century-old problem of dairy fermentation failure.

[0048] (5) Adaptive optimization: Through effect verification and feedback steps, the structural characteristic parameters and resonant frequency can be continuously corrected to improve the accuracy and reliability of the system in long-term operation.

[0049] (6) Essential advantages of ceramic microfiltration "cold sterilization" technology: Compared with the existing ceramic membrane microfiltration (MF) cold sterilization process, the present invention has fundamental differences in the following aspects: (a) Phage removal ability - MF pore size is 0.8-1.4 μm, while the size of phage is only 30-80 nm (a difference of more than 10,000 times), MF cannot intercept it at all, while the present invention can target and remove phage with GHz band ultrasound; (b) Selectivity - MF is based on size sieving and indiscriminately retains all microorganisms larger than the pore size, and cannot distinguish between pathogenic bacteria and beneficial bacteria, while the present invention can selectively kill them according to frequency band; (c) In-situ killing - MF only physically separates microorganisms from the permeate to the retrieval liquid (the concentrate contains 10 7 -10 8 (cFU / mL live bacteria), this invention breaks down cell walls in situ in milk, eliminating the need to treat concentrated waste liquid; (d) No membrane fouling - MF faces serious protein / fat membrane fouling problems, requiring frequent CIP cleaning. The transducer of this invention is non-contact installed, eliminating consumable pollution problems; (e) Engineered multi-band transducer and impedance matching design - Special piezoelectric materials and acoustic matching layers are used for different frequency bands to ensure efficient energy injection into the milk system (see Examples 8 and 9 for details).

[0050] (7) Synergistic effect with ceramic microfiltration process: The ultrasonic resonance treatment system of the present invention can be integrated in series with the existing ceramic microfiltration production line. MF is used as the front-end pre-filtration to remove most of the bacterial load and somatic cells, and ultrasonic resonance is used for precise targeting at the back end (killing bacteriophages that escaped from MF, selective sterilization, and replacing the subsequent mild pasteurization step of MF). Since MF has removed more than 99.99% of the bacterial load, the back-end ultrasound only needs to be supplemented under low load conditions. The total energy consumption of the combined process can be controlled at about 20,000 J / L (MF about 14,000 J / L + low load ultrasonic resonance about 6,000 J / L), which is 75% more energy-efficient than the "MF + pasteurization" combination (about 80,000 J / L), and at the same time completely eliminates the damage of heat-sensitive nutrients such as IgG to subsequent heat treatment. Attached Figure Description Appendix Figure 1 Method Flowchart Appendix Figure 2 Resonance frequency distribution diagram of 57 microorganisms Appendix Figure 3 Schematic diagram of dual-mode fluid processing channel structure Appendix Figure 4 Flowchart of the time-separation strategy Appendix Figure 5 Overall architecture diagram of full-band continuous flow equipment Detailed Implementation

[0051] The present invention will be further described in detail below with reference to specific embodiments. These embodiments are for illustrative purposes only and are not intended to limit the scope of the invention. Example 1: Construction of a Milk Microbial Structural Fingerprint Database

[0052] 1.1 Microbial species coverage This invention covers all 57 microorganisms detected in milk, including: (a) 39 types of bacteria: 12 types of pathogenic bacteria (Escherichia coli O157:H7, Salmonella, Staphylococcus aureus, Listeria monocytogenes, Mycobacterium tuberculosis, Brucella, Clostridium perfringens, Clostridium botulinum, Enterococcus faecalis, Streptococcus agalactiae, Campylobacter jejuni, Mycobacterium avium paratuberculosis species MAP), 15 types of putrefactive bacteria, 6 types of thermostable bacteria, and 6 types of beneficial bacteria (Lactobacillus acidophilus, Lactobacillus bulgaricus, Lactococcus lactis, Streptococcus thermophilus, Lactobacillus helveticus, Lactobacillus casei). (b) 12 bacteriophages (covering common bacteriophage groups in dairy fermentation such as 936, c2, and P335, with measured / calculated resonance frequencies ranging from approximately 2.2 to 6.9 GHz). (c) Six types of yeast (Kluyveromyces martensii, Yersinia lipolytica, Hansenula barry yeast, Candida famaria, Saccharomyces cerevisiae, Geotrichum candida, with a resonant frequency of approximately 3.4-7.9 MHz).

[0053] 1.2 Structural Parameter Acquisition For each microorganism, the following structural parameters were obtained from authoritative microbiology databases and literature: The structural parameter ranges for the three types of microorganisms are as follows:

[0054] Bacteria: equivalent radius R is 0.3-1.5 micrometers, wall thickness h is 2-40 nanometers, elastic modulus E is 30-350 kPa, density rho is 1050-1100 kg / m3, and Poisson's ratio nu is 0.25-0.35.

[0055] Bacteriophages: equivalent radius R is 15-40 nm, wall thickness h is 2-4 nm, elastic modulus E is 1-2 GPa, density rho is 1300-1400 kg / m3, and Poisson's ratio nu is 0.30-0.40.

[0056] Yeast: Equivalent radius R is 2-3.5 micrometers, wall thickness h is 100-200 nanometers, elastic modulus E is 20-80 MPa, density rho is 1050-1100 kg / m3, and Poisson's ratio nu is 0.25-0.35.

[0057] 1.3 Fingerprint Vector Definition Each microorganism possesses a unique structural fingerprint vector composed of a set of structural feature parameters, including but not limited to: equivalent radius, cell shape factor, cell wall thickness, number of cell walls, elastic modulus of each layer, cell wall density, Poisson's ratio, outer membrane / capsule composite feature parameters, and spore / special structural parameters. These parameters can be further normalized to a unified dimension for ease of calculation and storage. Different microorganisms possess different fingerprint vectors, from which different resonance frequencies are calculated.

[0058] The number of dimensions and the specific meaning of each dimension of the fingerprint vector mentioned above are determined by those skilled in the art based on the target microorganism category, measurable parameters and processing accuracy requirements, and are not necessary limitations of this invention. Example 2: Resonance Frequency Calculation Results

[0059] Using the structural fingerprint database constructed in Example 1, the resonance frequencies of all 57 microorganisms were calculated according to the thin-shell resonance model of the present invention. The results are distributed by frequency band as follows: The calculated resonance frequencies of representative microorganisms are as follows:

[0060] (1) Escherichia coli O157:H7: equivalent radius R is 0.65 μm, wall thickness h is 8 nm, elastic modulus E is 50 kPa, and predicted resonance frequency is 99 kHz.

[0061] (2) Salmonella: R is 0.60 micrometers, h is 8 nanometers, E is 48 kPa, and the frequency is 70 kHz.

[0062] (3) Fluorescent Pseudomonas: R is 0.65 μm, h is 10 nm, E is 35 kPa, and frequency is 70 kHz.

[0063] (4) Bacillus cereus vegetative body: R is 0.75 micrometers, h is 25 nanometers, E is 120 kPa, and the frequency is 355 kHz.

[0064] (5) Clostridium perfringens: R is 0.70 micrometers, h is 30 nanometers, E is 100 kPa, and the frequency is 242 kHz.

[0065] (6) Listeria monocytogenes: R is 0.50 micrometers, h is 20 nanometers, E is 180 kPa, and the frequency is 1.23 MHz.

[0066] (7) MAP Mycobacterium avium paratuberculosis: R is 0.40 μm, h is 35 nm, E is 250 kPa, and frequency is 1.62 MHz.

[0067] (8) Staphylococcus aureus: R is 0.50 micrometers, h is 25 nanometers, E is 350 kPa, and the frequency is 2.35 MHz.

[0068] (9) Enterococcus faecalis: R is 0.55 micrometers, h is 22 nanometers, E is 320 kPa, and the frequency is 4.47 MHz.

[0069] (10) Lactococcus: R is 0.50 micrometers, h is 23 nanometers, E is 340 kPa, and frequency is 2.40 MHz.

[0070] (11) Lactobacillus bulgaricus: R is 0.45 micrometers, h is 18 nanometers, E is 200 kPa, and the frequency is 221 kHz.

[0071] (12) Saccharomyces cerevisiae: R is 3.5 micrometers, h is 150 nanometers, E is 50 MPa, and the frequency is 3.39 MHz.

[0072] (13) Lactic acid bacteria phage group 936: R is 30 nm, h is 3 nm, E is 1.5 GPa, and the frequency is 2.99 GHz.

[0073] Frequency stratification pattern: - Gram-negative bacteria (thin-walled, outer membrane modified) → Low frequency band 50-200 kHz - Gram-positive bacilli and spores (medium-thick wall) → Mid-frequency band 200-500 kHz - Gram-positive cocci (thick-walled, high-modulus) → High-frequency band 1-5 MHz - Yeast (large size, eukaryotic thick-walled) → Mainly distributed in 5-10 MHz, with Saccharomyces cerevisiae at approximately 3.4 MHz, located at the upper end of the high-frequency band. During implementation, it can be integrated into the ultra-high frequency band for unified processing. - Bacteriophage (nanoscale protein capsid) → Microwave band 2-7 GHz Frequency separation analysis: Cross-species frequency resolution analysis showed that the minimum frequency resolution between different groups of microorganisms was 1.4% (Lactobacillus brevis 348.24 kHz vs. Lactobacillus acidophilus 343.37 kHz), and the minimum resolution within the same group was 2% (Staphylococcus aureus 2.35 MHz vs. Lactococcus lactis 2.40 MHz). With a conventional physical quality factor Q≈5, the resonance peak half-width was approximately 20%, and the minimum resolvable frequency resolution was approximately 20%. After enabling the high-Q narrowband frequency locking described in Example 11 of this invention, the effective resolution can be improved to approximately 5%. Therefore, for target-protective bacteria pairs with a resolution <5%, a time-series separation strategy (broad-spectrum sterilization first, followed by inoculation with protective bacteria after cooling) is adopted instead of a frequency-selective strategy.

[0074] 3.1 Processing Unit Configuration According to a specific embodiment of the system of the present invention, an ultrasonic resonance processing device is constructed: (a) Coarse channel processing unit: stainless steel cylindrical cavity, inner diameter 4 cm, length 100 cm, with annular transducer bracket installed on the outer wall, which can replace transducers of different frequency bands. (b) Thin film gap treatment unit: flat cavity, gap width 30 cm, gap thickness 1.2 mm, planar transducer arrays installed on both sides, 4 stages in series, with cooling water jackets between stages; (c) Temperature control system: circulating cooling water jacket to maintain milk temperature at 4±2°C; (d) Peristaltic pump: Control the flow rate to ensure that the processing residence time of each frequency band is ≥15 s.

[0075] 3.2 Processing Parameter Settings Recommended processing parameters (flow rate 5 L / min) for each frequency band are as follows:

[0076] Low frequency band (50-200 kHz): Target microorganisms are Escherichia coli, Salmonella, and Pseudomonas. A coarse channel (4 cm in diameter and 1 m in length) is used, with a sound pressure of 0.1-0.2 MPa and a residence time of about 15 seconds.

[0077] Mid-frequency band (200-500 kHz): Target microorganisms are Bacillus cereus and Clostridium perfringens. A coarse channel (4 cm in diameter and 1 m in length) is used, with a sound pressure of 0.1-0.2 MPa and a residence time of about 15 seconds.

[0078] High frequency band (1-5 MHz): Target microorganisms are Listeria, MAP, and Staphylococcus aureus. A thin film slit is used (slit thickness 1.2 mm), sound pressure is 0.1-0.25 MPa, and the residence time is about 0.9 seconds / level multiplied by 4 levels, which is about 3.5 seconds in total.

[0079] Ultra-high frequency band (5-10 MHz): The target microorganism is yeast. A thin film slit (slit thickness 1.2 mm) is used, with a sound pressure of 0.1-0.2 MPa and a residence time of about 0.9 seconds per level, multiplied by 4 levels, for a total of about 3.5 seconds.

[0080] Note: The residence time of a single stage in the high-frequency / ultra-high-frequency band at a flow rate of 5 L / min is approximately 0.9 s; if a longer residence time is required, it can be achieved by increasing the number of cascade stages (up to 20 stages) or by reducing the flow rate.

[0081] 3.3 Implementation of the Time Separation Strategy For scenarios where the frequency separation between Staphylococcus aureus (2.35 MHz) and Lactococcus lactis (2.40 MHz) is only 2%, a timing separation strategy is adopted: Phase 1 (Broad-spectrum sterilization): Low-frequency → medium-frequency → high-frequency ultrasonic resonance treatment is applied to raw milk in sequence to eliminate all pathogenic microorganisms (including Staphylococcus aureus). Second stage (cooling): Cool the pasteurized milk to 4°C; The third stage (inoculation and fermentation): Inoculate with target lactic acid bacteria (Lactococcus lactis, Streptococcus thermophilus, etc.), raise the temperature to the fermentation temperature (37-43°C), do not apply ultrasound, and allow natural fermentation.

[0082] This strategy physically avoids the problem of insufficient frequency selectivity while retaining all the nutritional advantages of "cold sterilization". Example 4: Acoustic Penetration Characteristics of Milk and Engineering Adaptation

[0083] 4.1 Milk Acoustic Model Whole milk, as a complex multiphase system (fat globules + casein micelles + whey protein + lactose + minerals + water), exhibits frequency-dependent attenuation characteristics to ultrasound. This invention establishes the following four-component attenuation model: α(f) = α_base · f^1.2 + α_fat · f 4 + α_casein · f^1.5 + α_protein · f^1.1 Where: α_base is the basic absorption coefficient (whey / aqueous phase), and α_fat is the Rayleigh scattering coefficient of fat globules (fat globules have a radius of ~2μm, much smaller than the wavelength of the processing frequency band, and the scattering intensity f is...). 4 α_casein is the micelle absorption coefficient of casein, and α_protein is the absorption coefficient of whey protein. The measured coefficients for whole milk (4% fat, 2.6% casein, 0.6% whey protein) are: α_base ≈ 2.0 dB / (cm·MHz^1.2), α_fat ≈ 0.8 dB / (cm·MHz). 4 (Normalized to 0.5MHz), α_casein≈0.4 dB / (cm·MHz^1.5), α_protein≈0.3 dB / (cm·MHz^1.1).

[0084] Calculation results: - 100 kHz → Attenuation approximately 0.16 dB / cm → 20 dB penetration depth > 100 cm - 500 kHz → Attenuation approximately 1.95 dB / cm → 20 dB penetration depth approximately 10 cm - 1 MHz → Attenuation approximately 15.5 dB / cm → 20 dB penetration depth approximately 1.3 cm - 2.35 MHz → Attenuation of approximately 399 dB / cm (high-frequency limit, dominated by fat globule scattering) → Must be treated with thin-film slits 4.2 Dual-mode processing channel engineering adaptation Based on the above acoustic characteristics, this invention adopts a dual-mode processing channel design: Mode A (coarse channel, ≤1 MHz): 4 cm diameter cylindrical cavity, continuous flow processing. Negligible attenuation at 100 kHz, 20 dB penetration depth of approximately 10 cm at 500 kHz, far exceeding the tube diameter, acoustic energy transfer efficiency >95%. Suitable for all Gram-negative bacteria and spore-forming bacteria.

[0085] Mode B (Thin-film slit, >1 MHz): 1.2 mm slit × 30 cm width, 4 stages in series. Each stage ensures that ultrasound can penetrate the entire slit thickness (1.2 mm << penetration depth), and inter-stage cooling prevents temperature rise accumulation. The 4 stages in series ensure that the total treated dose is ≥ equivalent to 6 dB of power transfer. Suitable for Gram-positive cocci and yeasts.

[0086] 5.1 Energy Consumption Comparison The energy consumption comparison between this invention and pasteurization is as follows:

[0087] Regarding sterilization energy consumption: pasteurization is approximately 252,000 J / L (0.07 kWh / L); the full-frequency design sound intensity of this invention is approximately 225,700 J / L (0.063 kWh / L), and after sound intensity optimization, it is approximately 137,500 J / L (0.038 kWh / L).

[0088] In terms of the theoretical minimum energy consumption per frequency band: the present invention has a power consumption of approximately 6,000 J / L.

[0089] Regarding temperature rise: pasteurization is 68°C (from 4°C to 72°C); the present invention is below 1.5°C.

[0090] Regarding the source of energy consumption: pasteurization is achieved through electric heating and cooling; the present invention utilizes the power consumption of an ultrasonic transducer.

[0091] The full-band energy consumption is calculated based on the total acoustic power and flow rate of the three frequency bands in series (low frequency + mid frequency + high frequency): When the design sound intensity is 2.0-3.0 W / cm², the total acoustic power is approximately 11.3 kW, which translates to approximately 0.063 kWh / L based on a flow rate of 5 L / min; after optimizing the sound intensity to near the resonance threshold (approximately 1.3 W / cm²), the energy consumption is approximately 0.038 kWh / L. The theoretical minimum acoustic energy for single-band, single-target processing (e.g., killing only E. coli) is approximately 6,000 J / L.

[0092] 5.2 Nutritional Retention Prediction The following is a comparison of the retention of key nutrients in milk between this invention and pasteurization:

[0093] Immunoglobulin G (IgG): Pasteurization retention rate is less than 10%, while the retention rate predicted by this invention is greater than 80%. The difference is due to the thermal denaturation temperature of about 65°C and the temperature rise of resonance treatment being less than 2°C.

[0094] Lactoferrin: The retention rate after pasteurization is less than 20%, while the retention rate predicted in this invention is higher than 85%. The difference is due to the fact that heat-sensitive proteins maintain their active conformation after low-temperature treatment.

[0095] Alpha-lactalbumin: The pasteurization retention rate is about 50%, and the retention rate predicted in this invention is higher than 95%. The difference mechanism is that the heat denaturation temperature is about 62°C.

[0096] beta-lactoglobulin: The pasteurization retention rate is about 30%, and the present invention predicts that the retention rate is higher than 90%. The difference mechanism is that the heat denaturation temperature is about 78°C, but prolonged heat exposure leads to inactivation.

[0097] Vitamin C: Pasteurization retention rate is about 70-80%, and the retention rate predicted by this invention is higher than 98%. The difference is due to minimal oxidation loss without heat treatment.

[0098] Vitamin B12: Pasteurization retention rate is about 85-90%, and the present invention predicts a retention rate of over 98%, the difference mechanism being non-heat treatment.

[0099] Minerals (Ca, P, Mg): Pasteurization retention rate is about 100%, and the retention rate predicted by this invention is about 100%. The difference mechanism is that ultrasound does not destroy inorganic ions. Example 6: Phage-targeted clearance

[0100] Bacteriophage contamination is a major cause of fermentation failure in dairy fermentation. Due to their nanoscale size (30-80 nm) and protein capsid structure, bacteriophages have a resonance frequency in the GHz band (2-7 GHz), which is more than 1000 times higher than that of lactic acid bacteria (MHz band) (frequency separation of about 99.9%), enabling completely selective removal.

[0101] Specific implementation measures: (a) Integrating a GHz-band planar transducer in the fermenter's air / feed pipeline; (b) Apply 2-7 GHz pulsed ultrasound to the fermentation medium or feed solution (pulse duty cycle 10-30%, to avoid overheating of the liquid); (c) Sound pressure 0.05-0.15 MPa, processing time 5-15 s; (d) Lactic acid bacteria (MHz band) are not affected by the GHz band and their activity is fully preserved.

[0102] This solution is the first to achieve selective treatment at the physical level, which "kills bacteriophages and preserves lactic acid bacteria," fundamentally solving the problem of fermentation failure in the dairy industry.

[0103] This embodiment provides a combined process that integrates ceramic membrane microfiltration (MF) with ultrasonic resonance targeted sterilization. It aims to combine the high-throughput advantage of MF pre-filtration and the selective advantage of ultrasonic resonance back-end precise targeting to achieve a milk processing solution with lower energy consumption and higher nutrient retention than the existing "MF + light pasteurization" process.

[0104] 7.1 Process Flow Step 1 (MF Pre-filtration): After standardization (centrifugation to remove fat, fat content <0.1%), raw milk enters the ceramic membrane microfiltration unit. An α-Al₂O₃ ceramic membrane with a pore size of 1.4 μm is used, operating at 6-10°C, with a transmembrane pressure difference ≤0.3 MPa and a tangential flow rate ≥3 m / s. The total bacterial count in the MF permeate is reduced by ≥4 log₂, and the spore removal rate is ≥99.9%.

[0105] Step Two (Precise Targeting with Ultrasonic Resonance): The MF permeating fluid directly enters the ultrasonic resonance processing system of this invention, and passes through the following stages in sequence: (a) Low-frequency processing (50-200 kHz, coarse channel mode): kill Gram-negative bacteria (E. coli, Salmonella, etc.) that escaped the MF filter, sound pressure 0.15 MPa, dwell time 15 s; (b) Mid-frequency processing (200-500 kHz, coarse channel mode): kill spore-forming bacteria and thermostable bacteria that escaped the MF filter, sound pressure 0.15 MPa, dwell time 15 s; (c) High-frequency processing (1-5 MHz, thin film slit mode): kill Gram-positive cocci (Listeria, Staphylococcus aureus, etc.) that escaped the MF filter, sound pressure 0.2 MPa, 1.2 mm slit × 4 stages in series; (d) GHz band processing (2-7 GHz, pulse mode): targeted removal of bacteriophages, pulse duty cycle 20%, sound pressure 0.1MPa, processing time 10 s.

[0106] Since MF has removed more than 99.99% of the bacterial load, the ultrasonic resonance in step two only operates under extremely low residual load, and its power can be significantly reduced to near the ideal level of a single frequency band.

[0107] Step 3 (Fat Refilling and Standardization): The skim milk treated with ultrasonic resonance is mixed with separately pasteurized light cream to achieve the target fat content (3.5% whole fat or 1.5% low fat) and then filled.

[0108] 7.2 Comparison with existing processes A comprehensive comparison of the three ESL milk processing schemes is as follows:

[0109] Option A (Traditional ESL process): The front end uses MF 1.4 micron filtration, and the back end uses mild pasteurization at 65°C / 15s. The total energy consumption is about 80,000 J / L, the IgG retention rate is about 80-90%, it cannot remove bacteriophages, and the shelf life is 30-45 days.

[0110] Option B (the combined process of the present invention): The front end uses MF 1.4 micron filtration, and the back end uses low-load ultrasonic resonance targeted sterilization. The total energy consumption is about 20,000 J / L, the IgG retention rate is higher than 95%, it can remove bacteriophages, and the shelf life is 30-60 days.

[0111] Option C (Independent process of this invention): No MF front-end treatment, and full-band ultrasonic resonance sterilization at the back end. The total energy consumption is about 137,000-226,000 J / L (see Example 5). The IgG retention rate is higher than 80%, which can remove bacteriophages. The quality is expected to be verified by experiments.

[0112] 7.3 Technical Advantages of Combined Processes (a) Front-end MF reduces back-end load: After MF removes more than 99.99% of bacteria and spores, the ultrasonic resonance system only needs to handle a very low residual load, allowing for higher flow rates and shorter residence times, further increasing production capacity; (b) Replacement of post-processing heat treatment: Ultrasonic resonance completely replaces the mild pasteurization step (65°C / 15s) following MF, eliminating the thermal damage to IgG (denaturation temperature ~65°C) and lactoferrin (denaturation temperature ~68°C) caused by this step, thereby increasing the IgG retention rate of the final product from 80-90% to >95%; (c) Filling the functional gaps of MF: The GHz band of ultrasonic resonance targets and removes bacteriophages that MF cannot intercept, and selectively kills specific pathogens that MF cannot distinguish, fundamentally filling the two biggest defects of the MF process; (d) Significantly reduced total energy consumption: The total energy consumption of the combined process is approximately 20,000 J / L, which is 75% more energy-efficient than the traditional "MF + Pasteur" process of 80,000 J / L; (e) Compatible with existing production line upgrades: The ultrasonic resonance module can be connected as a back-end upgrade kit for existing MF production lines without the need to remove existing MF equipment, thus reducing the capital threshold for dairy companies to upgrade their technology. Example 8: Selection of Multi-band Transducer Array

[0113] To address the acoustic characteristics and power requirements of different frequency bands, this invention employs a dedicated piezoelectric transducer selection for each frequency band: The transducer array selection (corresponding to a production capacity of 5 L / min and three-band series connection) is as follows:

[0114] Low frequency band (200 kHz): PZT-8 piezoelectric material is used, with a ring unit (40 mm in diameter), a single unit power of 150 W, a total acoustic power of 2,513 W, and a total of 17 units.

[0115] Mid-frequency band (1 MHz): Uses PZT-4 piezoelectric material, flat panel unit (diameter 20 mm), single unit power 60W, total acoustic power 1,571 W, a total of 27 units.

[0116] High frequency band (2.5 MHz): PZT-5A piezoelectric material is used, thin film patch unit form (diameter 10 mm), single unit power is 25 W, total acoustic power is 7,200 W, and there are a total of 288 units.

[0117] Selection criteria: Low-frequency band requires high-power radial focusing (PZT-8, high mechanical quality factor, low loss); mid-frequency band requires sound pressure stability (PZT-4, high electromechanical coupling coefficient); high-frequency band requires bandwidth and thin-film structure (PZT-5A, high sensitivity). Transducers for each frequency band are installed on the outer wall or sides of the corresponding processing unit, using non-contact installation, eliminating consumable pollution. Example 9: Sound Matching Layer Design

[0118] To achieve efficient injection of ultrasound from the transducer into the milk system, this invention incorporates a 1 / 4 wavelength acoustic matching layer between the processing cavity wall and the transducer. Z_match = √(Z_PZT × Z_milk) Where Z_PZT is the acoustic impedance of the piezoelectric material (PZT-4 is approximately 33 MRayl), and Z_milk is the acoustic impedance of milk (approximately 1.56 MRayl). The calculated acoustic impedance of the matching layer is approximately 7.16 MRayl. The matching layer material is selected as epoxy resin + tungsten powder composite or epoxy glass microspheres (acoustic impedance 6-8 MRayl), and the thickness is taken as 1 / 4 wavelength of the corresponding frequency. - Low frequency band 200 kHz: thickness approximately 1.888 mm - Mid-frequency band 1 MHz: thickness approximately 0.378 mm - High-frequency band 2.5 MHz: thickness approximately 0.151 mm The acoustic matching layer significantly reduces the reflection loss at the PZT↔ milk interface and greatly improves the acoustic energy injection efficiency, which is the key to ensuring the power transfer of the thin-film slot mode in the high-frequency band (>1 MHz) of this device. Example 10: Cavitation Suppression-Resonance Preservation Synergistic Module

[0119] To avoid acoustic cavitation in the milk system under high-frequency sound intensity (cavitation is an indiscriminate broad-spectrum destruction, which contradicts the selective resonance objective), the processing system of this invention is equipped with a cavitation suppression module: (a) Overpressure device: Apply a static pressure of 1.5-3 bar (preferably 2 bar) to the processing chamber to increase the liquid cavitation threshold and create a wider operating window for the system between the resonance failure threshold (0.2 MPa) and the cavitation threshold (0.5 MPa); (b) Degassing membrane: Milk is pre-degassed using a hollow fiber degassing membrane before processing to reduce dissolved gas content and inhibit cavitation nuclei formation; (c) Frequency Lock-in Linkage: The control module ensures that the operating frequency is always locked at the center of the target resonance peak rather than the cavitation noise band based on the spectrum monitored by the online hydrophone.

[0120] This collaborative module enables the system to operate stably within a window where "resonance destruction is dominant and cavitation is suppressed," which is fundamentally different from existing cavitation sterilization routes. Example 11: Resonance Frequency Self-calibration and Lock-in Control

[0121] To achieve frequency stability and selectivity accuracy during long-term operation, the control module of this invention adopts the following closed-loop strategy: (a) Online peak finding: Broadband hydrophones are placed at the inlet and outlet of the processing chamber to collect ultrasonic spectrum in real time and identify the position of the target bacteria resonance peak; (b) Frequency fine-tuning: When a peak offset is detected (such as due to temperature or composition fluctuations), the control module automatically fine-tunes the drive frequency with a locking accuracy of ±0.1%; (c) Sound pressure-temperature dual closed loop: The temperature rise is limited to ≤2°C by fiber optic temperature measurement feedback, and the driving power is adjusted according to the sound pressure feedback of the hydrophone. (d) Endpoint criterion: The rate of change of bacterial community peaks before and after treatment was compared online and used as the endpoint criterion for bactericidal effect.

[0122] This locking control improves the effective resolution of a conventional wide-peak system with Q≈5 to about 5%, supporting direct frequency-selective killing in scenarios with Δf≥5%. Example 12: CIP in-situ cleaning

[0123] The processing system of this invention supports in-situ cleaning (CIP), and the cleaning procedure is as follows: Circulate 1% NaOH solution at 70°C for 20 min → Rinse with clean water → Circulate 1% HNO3 solution at 60°C for 20 min → Rinse with sterile water until neutral.

[0124] Since the transducers are all non-contact externally mounted, there is no need to disassemble the treatment chamber during the CIP process; the cleaning fluid only comes into contact with the stainless steel chamber and the gap channel, and does not affect the life of the piezoelectric element.

Claims

1. A method for targeted sterilization using ultrasonic resonance based on microbial structural fingerprints, characterized in that, Includes the following steps: S1: Obtain the individual structural feature parameter set of each microorganism in the target microbial community, wherein the structural feature parameter set includes at least cell geometry parameters, cell wall parameters, and wall material parameters; S2: Based on the thin-shell resonance model, calculate the intrinsic mechanical resonance frequency of each microorganism according to the set of structural feature parameters; S3: Group the target microorganisms into frequency bands according to the resonant frequency and generate a frequency application strategy; S4: According to the frequency application strategy, apply ultrasonic waves with a frequency matching the resonant frequency to the liquid food, and control the sound pressure within the range of 0.05-0.4 MPa to cause the cell wall or capsid of the target microorganism to resonate and rupture, while keeping the acoustic cavitation threshold of the liquid food from being exceeded.

2. The method according to claim 1, characterized in that, The set of structural feature parameters mentioned in step S1 includes: Equivalent radius R, cell shape factor, wall thickness h, number of wall layers and elastic modulus of each layer E, wall surface density ρ, Poisson's ratio ν, outer membrane integrity index.

3. The method according to claim 1, characterized in that, Step S2 calculates the inherent mechanical resonance frequency based on the respiratory mode of the thin-shell resonance model. The respiratory mode resonance frequency satisfies the thin-shell resonance relationship with the equivalent radius, wall elastic modulus, wall density, wall thickness, and Poisson's ratio. For microorganisms with multi-layer wall structures, the weighted average of the elastic modulus and thickness of each layer is used as the equivalent elastic modulus for calculation.

4. The method according to claim 1, characterized in that, In step S2, for Gram-negative bacteria, an outer membrane correction factor γ_outer is introduced to correct the calculated frequency, with the value of γ_outer ranging from 0.6 to 1.0; for bacteriophages, a capsid symmetry correction factor σ_capsid is introduced to correct the calculated frequency.

5. The method according to claim 1, characterized in that, The frequency band grouping mentioned in step S3 includes: The low-frequency band (50-200 kHz) corresponds to Gram-negative thin-walled bacteria, the mid-frequency band (200-500 kHz) corresponds to Gram-positive bacilli and spores, the high-frequency band (1-5 MHz) corresponds to Gram-positive cocci, the ultra-high-frequency band (5-10 MHz) corresponds to yeast, and the microwave band (2-7 GHz) corresponds to bacteriophages.

6. The method according to claim 1, characterized in that, Step S3 also includes: Calculate the frequency separation Δf between the target microorganism and the microorganism to be protected: Δf = |f_target - f_protected| / f_target × 100%; When Δf ≥ 5%, a frequency selection strategy is adopted to directly apply the target frequency to kill the target microorganism while preserving the microorganism to be protected; When Δf < 5%, a time-series separation strategy is adopted, first applying broad-spectrum resonance treatment to kill the target microorganism, and then inoculating the microorganism to be protected after cooling.

7. The method according to claim 1, characterized in that, The application of ultrasound in step S4 employs different fluid channel modes depending on the resonant frequency: When the resonant frequency is ≤1 MHz, the coarse channel mode is adopted, and the liquid food flows through a cylindrical processing chamber with a diameter of 3-5 cm. When the resonant frequency is >1 MHz, a thin-film slit mode is adopted, and the liquid food flows through a flat processing cavity with a slit thickness of 0.8-1.5 mm.

8. The method according to claim 7, characterized in that, The coarse channel mode and the thin film slit mode are used in series and combined, with liquid food flowing sequentially through the coarse channel processing unit and the thin film slit processing unit to achieve continuous flow processing of microorganisms across the entire frequency band.

9. The method according to claim 1, characterized in that, The processing temperature in step S4 is controlled at 0-10°C, and the temperature rise of the liquid food during the processing does not exceed 5°C.

10. The method according to claim 1, characterized in that, It also includes step S5: Microbial testing is performed on the processed liquid food, and the actual kill rate is compared with the expected kill rate. If the deviation exceeds the preset threshold, the structural characteristic parameters of the corresponding microorganisms are corrected and the resonance frequency is recalculated.

11. The method according to any one of claims 1-10, characterized in that, The liquid food is milk or dairy products, and the target microbial community includes pathogenic bacteria in milk, including one or more of Escherichia coli O157:H7, Salmonella, Staphylococcus aureus, and Listeria monocytogenes.

12. The method according to claim 11, characterized in that, The target microbial community also includes bacteriophages in milk. Ultrasonic waves in the 2-7 GHz band are applied to the bacteriophages for targeted elimination while preserving the activity of lactic acid bacteria.

13. An ultrasonic resonance targeted sterilization system based on microbial structural fingerprinting, characterized in that, include: The fingerprint database module stores the set of individual structural feature parameters for each microorganism in the target microbial community; The frequency calculation module has a built-in thin-shell resonance model, which is used to calculate the inherent mechanical resonance frequency of each microorganism based on the set of structural feature parameters, and to perform frequency band grouping and frequency separation analysis. The ultrasonic processing module includes a multi-band transducer array and a fluid processing chamber, wherein the fluid processing chamber comprises a coarse channel processing unit with a diameter of 3-5 cm and a thin film slit processing unit with a slit thickness of 0.8-1.5 mm. The control module is communicatively connected to the fingerprint database module, frequency calculation module, and ultrasonic processing module, and is used to automatically control the working frequency, sound pressure, and processing time of the ultrasonic processing module based on the frequency calculation results.

14. The system according to claim 13, characterized in that, The multi-band transducer array includes: The low-frequency transducer group operates at a frequency of 50-200 kHz, uses PZT-8 piezoelectric material, and is installed on the outer wall of the coarse channel processing unit. The intermediate frequency transducer group operates at a frequency of 200-500 kHz, uses PZT-4 piezoelectric material, and is installed on the outer wall of the coarse channel processing unit. The high-frequency transducer group operates at a frequency of 1-5 MHz and uses PZT-5A piezoelectric material. It is installed on both sides of the thin film gap treatment unit. The ultra-high frequency transducer array, operating at a frequency of 5-10 MHz, is installed on both sides of the thin-film gap treatment unit.

15. The system according to claim 13, characterized in that, It also includes an acoustic matching layer, which is disposed between the transducer and the liquid food processing chamber. The acoustic impedance of the acoustic matching layer is located near the geometric median of the acoustic impedance of the piezoelectric material and the acoustic impedance of the liquid food, and the thickness is one-quarter wavelength of the corresponding operating frequency.

16. The system according to claim 13, characterized in that, It also includes a cavitation suppression module, which includes an overpressure device that applies a static pressure of 1.5-3 bar to the processing chamber and / or a hollow fiber degassing membrane for pre-degassing the liquid food, in order to increase the cavitation threshold of the liquid food, suppress the cavitation effect, and retain the resonance damage effect.

17. The system according to claim 13, characterized in that, The control module also includes a frequency locking submodule, which includes a broadband hydrophone installed at the inlet and / or outlet of the processing chamber for real-time acquisition of ultrasonic spectrum and identification of target microbial resonance peaks, and automatic locking of the working frequency at the center of the resonance peaks with a frequency locking accuracy of ±0.1%.

18. The system according to claim 13, characterized in that, The thin film gap treatment unit is a multi-stage series structure, with each stage having a gap thickness of 0.8-1.5 mm and a cooling section between stages, for a total of 2-6 stages.

19. The system according to claim 13, characterized in that, It also includes a temperature control unit to maintain the temperature of liquid food within the range of 0-10°C during processing.

20. The system according to claim 13, characterized in that, The system supports in-situ cleaning (CIP). The cleaning procedure is as follows: alkaline washing with 1% NaOH solution at 70°C for 20 min, followed by water washing; acid washing with 1% HNO3 solution at 60°C for 20 min, followed by water washing until neutral. The transducer is installed externally without contact.

21. The application of the method according to any one of claims 1-12 or the system according to any one of claims 13-20 in dairy processing.

22. The application according to claim 21, characterized in that, The application is the cold sterilization process of drinking milk, which kills pathogenic microorganisms while maintaining a retention rate of >80% of immunoglobulin G in milk.

23. The application according to claim 21, characterized in that, The application is a selective pretreatment of fermented milk, which uses a time-sequential separation strategy to kill pathogenic Gram-negative bacteria and spore-forming bacteria, followed by inoculation with lactic acid bacteria for fermentation.

24. The application according to claim 21, characterized in that, The application involves targeted removal of bacteriophages during dairy fermentation, applying 2-7 GHz frequency ultrasound to contaminated bacteriophages while preserving the activity of lactic acid bacteria.

25. The method according to any one of claims 1-12, characterized in that, It also includes a pre-filtration ceramic membrane step: Before step S1, the liquid food is microfiltered through a ceramic membrane with a pore size of 0.8-1.4 μm. Microorganisms and somatic cells larger than the membrane pore size are physically retained at 5-15°C to obtain the microfiltration permeate. The ultrasonic resonance treatment in step S4 is applied to the microfiltration permeate to kill residual microorganisms that have escaped the microfiltration and to target and remove bacteriophages that the microfiltration cannot intercept.

26. The method according to claim 25, characterized in that, The microfiltration permeate is treated with ultrasonic resonance and then mixed with heat-treated light cream for fat standardization. The ultrasonic resonance treatment completely replaces the pasteurization step following microfiltration in the traditional ESL process, keeping the total energy consumption of the process below approximately 20,000 J / L and achieving an immunoglobulin G retention rate of >95%.

27. A combined system of ceramic membrane microfiltration and ultrasonic resonance targeted sterilization, characterized in that, include: The ceramic membrane microfiltration unit, comprising an α-alumina ceramic membrane with a pore size of 0.8-1.4 μm and an operating temperature of 5-15°C, is used for pre-filtration of liquid foods. An ultrasonic resonance targeted processing unit, connected in series downstream of the permeate outlet of the ceramic membrane microfiltration unit, includes a fingerprint database module, a frequency calculation module, a multi-band ultrasonic processing module, and a control module as described in any one of claims 13-20, for applying frequency-matched ultrasonic resonance processing to the microfiltration permeate. The fat backfilling mixing unit is connected downstream of the outlet of the ultrasonic resonance targeted processing unit and is used to mix the processed skim milk with heat-treated cream.

28. A selective protective sterilization method, characterized in that, Includes the following steps: Obtain the individual structural characteristic parameter sets of the target microorganism and the microorganism to be protected; The inherent mechanical resonance frequencies of the target microorganism and the microorganism to be protected were calculated based on the thin-shell resonance model. Calculate the frequency separation between the two: Δf = |f_target - f_protected| / f_target × 100%; When Δf ≥ 5%, apply ultrasound at the target microorganism's resonant frequency to kill the target microorganism while preserving the microorganism to be protected; When Δf < 5%, apply a broad-spectrum resonance treatment covering the frequency of the target microorganism first, and then inoculate the microorganism to be protected after cooling.

29. The method according to claim 28, characterized in that, The target microorganism is a pathogenic Gram-negative bacterium and / or a spore-forming bacterium, and the microorganism to be protected is a lactic acid bacterium. The method is used for the pretreatment of fermented dairy products. After treatment, the survival rate of the lactic acid bacteria to be protected is >90% and the inactivation amount of pathogenic microorganisms is ≥5 log.

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