Correction: Teng et al. Efficacy Assessment of Phage Therapy in Treating Staphylococcus aureus-Induced Mastitis in Mice. Viruses 2022, 14, 620.

In the original publication [...].

Ablation for malignant liver tumor using high-intensity focused ultrasound and radio-frequency: A meta-analysis.

BACKGROUND: Primary liver cancer is a major health issue, so finding the most effective treatment is vital. OBJECTIVE: The present meta-analysis compares high-intensity focused ultrasound (HIFU) to radiofrequency (RF) ablation for primary liver cancer treatment. METHODS: PubMed, MEDLINE, CNKI, VIP, and Wanfang were used to search for English and Chinese papers. After carefully confirming data completeness and applying inclusion and exclusion criteria, RevMan 5.3 was used to evaluate the included literature. Data analysis utilized a fixed-effects model for heterogeneity between 0.1 and 0.5. RESULTS: The meta-analysis included 304 patients: 119 had HIFU and 185 RF ablation. For primary liver cancer, HIFU and RF ablation were equally efficacious (odds ratio 1.02, 95% confidence interval [0.54, 1.92]). Overall survival, disease-free survival, and complications at 1, 2, and 3 years were not significantly different (odds ratio 0.72, 95% confidence range [0.04, 12.79], P= 0.82). CONCLUSION: The meta-analysis shows no significant difference in efficacy, long-term survival rates, or complication rates between HIFU and RF ablation for primary liver cancer, but more large-scale, high-quality randomized clinical trials are needed to prove their equivalence. Both therapy strategies seem promising, but additional information is needed to determine their respective merits.

Improved Performance of Acoustically Actuated Magnetoelectric Antenna with FeGa/FeGaB Bilayer.

Acoustically actuated magnetoelectric (ME) antennas utilize acoustic wave resonance to complete the process of receiving and transmitting signals, which promotes the development of antenna miniaturization technology. This paper presents a bilayer magnetostrictive/AlN ME laminated antenna. The proposed laminated antenna uses the FeGa/FeGaB bilayer materials as magnetostrictive materials, which combine the advantages of soft magnetic properties of FeGa and the low loss of FeGaB. First, multiphysics modeling and analysis are performed for the proposed ME laminated antenna by finite element method (FEM). The positive/inverse ME effects and the influences of the volume ratio of the FeGa/FeGaB bilayer on the antenna performance are studied. The results show that the output voltage and ME coefficient of the FeGa/FeGaB bilayer magnetostrictive material with a volume ratio of 1:1 are 3.97 times and 195.8% higher than that of the single FeGaB layer, respectively. The eddy current loss is 52.08% lower than that of single-layer FeGa. According to the surface equivalence principle, the far-field radiation process is simulated. The results show that the gain of the ME antenna is 15 dB larger than that of the same-size micro-loop antenna, and the gain of the ME antenna is about -44.9 dB. The improved performance and magnetic tunability of the proposed bilayer magnetostrictive materials make ME antennas excellent candidates for portable devices and implantable medical devices.

Whole-exome sequencing revealed novel genetic alterations in patients with tetralogy of Fallot.

Background: The most prevalent cyanotic congenital heart disease (CHD) phenotype is tetralogy of Fallot (TOF). Rare genetic variations have been identified as significant risk factors for CHD. Thus, this research sought to identify the pathogenic variations and molecular etiologies of TOF. Methods: This study employed whole-exome sequencing (WES) and Sanger sequencing to identify pathogenic variations in DNA samples from patients with TOF. The pathogenicity of the variations was predicted using an in-silico approach. Results: We enrolled 17 patients with TOF in this study. Among these patients, 14 had mutations in TOF-related genes, including GJB2, TBX15, CTNS, SPINK1, GATA6, PRIMOL, GDF15, SLC17A9, AIFM1, FOXC2, KLF13, ABCA4, CPA6, FKBP10, ASPA, SBF1, HBA2, IGLL1, GNE, and KLHL10. We also gathered WES data from three participants without TOF, who comprised the control group, but no variations were found in the indicated genes. Further analysis showed that the patients with FKBP10 and GNE variants had more serious clinical symptoms. Sanger sequencing confirmed that the two variants were heterozygous in TOF patients. Conclusions: We identified several genetic variants associated with TOF and confirmed that FKBP10 and GNE variants were associated with TOF severity. The findings of this study help researchers and clinicians on genetic counseling with the verification of the potential of WES in detecting TOF and help implement early interventions for patients with TOF.

Next-generation sequencing technology for the diagnosis of Pneumocystis pneumonia in an immunocompetent female: A case report.

BACKGROUND: Pneumocystis pneumonia (PCP) is a serious fungal infection usually seen in patients with human immunodeficiency virus, and it is more frequently found and has a high fatality rate in immunocompromised people. Surprisingly, it rarely occurs in immunocompetent patients. However, the clinical diagnosis of this pathogen is made more difficult by the difficulty of obtaining accurate microbiological evidence with routine tests. This case reports a PCP patient with normal immune function who was diagnosed through next-generation sequencing (NGS). CASE SUMMARY: A 23-year-old female who had no special disease in the past was admitted to the hospital with a persistent fever and cough. Based on the initial examination results, the patient was diagnosed with bipulmonary pneumonia, and empirical broad-spectrum antibiotic therapy was administered. However, due to the undetermined etiology, the patient's condition continued to worsen. She was transferred to the intensive care unit because of acute respiratory failure. After the diagnosis of Pneumocystis jirovecii infection through NGS in bronchoalveolar lavage fluid and treatment with trimethoprim/sulfamethoxazole and caspofungin, the patient gradually recovered and had a good prognosis. CONCLUSION: This case emphasizes that, for patients with normal immune function the possibility of PCP infection, although rare, cannot be ignored. NGS plays an important role in the diagnosis of refractory interstitial pneumonia and acute respiratory failure.

Development of a colloidal gold immunochromatographic strip with enhanced signal for the detection of bovine parvovirus.

Bovine parvovirus (BPV) is a pathogen responsible for respiratory and digestive tract symptoms in calves and abortion and stillbirth in pregnant cows. In this study, we developed a colloidal gold immunochromatographic (GICG) strip with an enhanced signal for detecting BPV according to the double-antibody sandwich principle and an enzyme-based signal amplification system to amplify the signal. This system utilizes horseradish peroxidase reacting with a substrate solution containing 3,3',5,5'-tetramethylbenzidine and dextran sulfate to obtain insoluble blue products on the test and control lines. We optimized different reaction conditions, including the amount of monoclonal antibodies (mAbs), pH of the colloidal gold solution, coating solution, blocking solution, sample pad treatment solution, antibody concentration in the control line, and antibody concentration in the detection line. The sensitivity of the signal-enhanced GICG strip showed that the minimum amount for detecting BPV was 102 TCID50, 10 times higher than that of the traditional GICG strip. The results of the specificity test showed that the signal-enhanced GICG strip had no cross-reactivity with BRV, BVDV, or BRSV. The results of the repeatability test showed that the coefficient of variation between and within batches was less than 5%, showing good repeatability. Moreover, for validation, PCR and the signal-enhanced GICG strip were used to detect 280 clinical bovine fecal samples. The concordance rate compared with PCR was 99.29%. Hence, the developed strip exhibited high sensitivity and specificity for the detection of BPV. Therefore, this strip could be a rapid, convenient, and effective method for the diagnosis of BPV infection in the field.

A miniaturized transit-time ultrasonic flowmeter based on ScAlN piezoelectric micromachined ultrasonic transducers for small-diameter applications.

Transit-time ultrasonic flowmeters (TTUFs) are among the most widely used devices for flow measurements. However, traditional TTUFs are usually based on a bulk piezoelectric transducer, which limits their application in small-diameter channels. In this paper, we developed a miniaturized TTUF based on scandium-doped aluminum nitride (ScAlN) piezoelectric micromachined ultrasonic transducers (PMUTs). The proposed TTUF contains two PMUT-based transceivers and a π-type channel. The PMUTs contain 13 × 13 square cells with dimensions of 2.8 × 2.8 mm2. To compensate for the acoustic impedance mismatch with liquid, a layer of polyurethane is added to the surface of the PMUTs as a matching layer. The PMUT-based transceivers show good transmitting sensitivity (with 0.94 MPa/V surface pressure) and receiving sensitivity (1.79 mV/kPa) at a frequency of 1 MHz in water. Moreover, the dimensions of the π-type channel are optimized to achieve a measurement sensitivity of 82 ns/(m/s) and a signal-to-noise ratio (SNR) better than 15 dB. Finally, we integrate the fabricated PMUTs into the TDC-GP30 platform. The experimental results show that the developed TTUF provides a wide range of flow measurements from 2 to 300 L/h in a channel of 4 mm diameter, which is smaller than most reported channels. The accuracy and repeatability of the TTUF are within 0.2% and 1%, respectively. The proposed TTUF shows great application potential in industrial applications such as medical and chemical applications.

Stepped-Tube Backside Cavity Piezoelectric Ultrasound Transducer Based on Sc0.2AI0.8N Thin Films.

This paper presents a novel piezoelectric micromachined ultrasonic transducer (PMUT) with theoretical simulation, fabrication, and testing. Conventional methods using a PCB or an external horn to adjust the PMUT acoustic field angle are limited by the need for transducer size. To address this limitation, the stepped-tube (expanded tube) backside cavity PMUT has been proposed. The stepped-tube PMUT and the tube PMUT devices have the same membrane structure, and the acoustic impedance matching of the PMUT is optimized by modifying the boundary conditions of the back cavity structure. The acoustic comparison experiments show that the average output sound pressure of the stepped-tube backside cavity PMUT has increased by 17%, the half-power-beam-width (θ-3db) has been reduced from 55° to 30° with a reduction of 45%, and the side lobe level signal is reduced from 147 mV to 66 mV. In addition, this work is fabricated on an eight-inch wafer. The process is compatible with standard complementary metal oxide semiconductor (CMOS), conditions are stable, and the cost is controllable, plus it facilitates the batch process. These conclusions suggest that the stepped-tube backside cavity PMUT will bring new, effective, and reliable solutions to ranging applications.

Miniature Ultrasonic Spatial Localization Module in the Lightweight Interactive.

The advancement of spatial interaction technology has greatly enriched the domain of consumer electronics. Traditional solutions based on optical technologies suffers high power consumption and significant costs, making them less ideal in lightweight implementations. In contrast, ultrasonic solutions stand out due to their lower power consumption and cost-effectiveness, capturing widespread attention and interest. This paper addresses the challenges associated with the application of ultrasound sensors in spatial localization. Traditional ultrasound systems are hindered by blind spots, large physical dimensions, and constrained measurement ranges, limiting their practical applicability. To overcome these limitations, this paper proposes a miniature ultrasonic spatial localization module employing piezoelectric micromechanical ultrasonic transducers (PMUTs). The module is comprised of three devices each with dimension of 1.2 mm × 1.2 mm × 0.5 mm, operating at a frequency of around 180 kHz. This configuration facilitates a comprehensive distance detection range of 0-800 mm within 80° directivity, devoid of blind spot. The error rate and failure range of measurement as well as their relationship with the SNR (signal-to-noise ratio) are also thoroughly investigated. This work heralds a significant enhancement in hand spatial localization capabilities, propelling advancements in acoustic sensor applications of the meta-universe.

An SOI-Structured Piezoresistive Differential Pressure Sensor with High Performance.

This paper presents a piezoresistive differential pressure sensor based on a silicon-on-insulator (SOI) structure for low pressure detection from 0 to 30 kPa. In the design phase, the stress distribution on the sensing membrane surface is simulated, and the doping concentration and geometry of the piezoresistor are evaluated. By optimizing the process, the realization of the pressure sensing diaphragm with a controllable thickness is achieved, and good ohmic contact is ensured. To obtain higher sensitivity and high temperature stability, an SOI structure with a 1.5 µm ultra-thin monocrystalline silicon layer is used in device manufacturing. The device diaphragm size is 700 µm × 700 µm × 2.1 µm. The experimental results show that the fabricated piezoresistive pressure sensor has a high sensitivity of 2.255 mV/V/kPa and a sensing resolution of less than 100 Pa at room temperature. The sensor has a temperature coefficient of sensitivity (TCS) of -0.221 %FS/°C and a temperature coefficient of offset (TCO) of -0.209 %FS/°C at operating temperatures ranging from 20 °C to 160 °C. The reported piezoresistive microelectromechanical systems (MEMS) pressure sensors are fabricated on 8-inch wafers using standard CMOS-compatible processes, which provides a volume solution for embedded integrated precision detection applications of air pressure, offering better insights for high-temperature and miniaturized low-pressure sensor research.

Bimorph Dual-Electrode ScAlN PMUT with Two Terminal Connections.

This paper presents a novel bimorph Piezoelectric Micromachined Ultrasonic Transducer (PMUT) fabricated with 8-inch standard CMOS-compatible processes. The bimorph structure consists of two layers of 20% scandium-doped aluminum nitride (Sc0.2Al0.8N) thin films, which are sandwiched among three molybdenum (Mo) layers. All three Mo layers are segmented to form the outer ring and inner plate electrodes. Both top and bottom electrodes on the outer ring are electrically linked to the center inner plate electrodes. Likewise, the top and bottom center plate electrodes are electrically connected to the outer ring in the same fashion. This electrical configuration maximizes the effective area of the given PMUT design and improves efficiency during the electromechanical coupling process. In addition, the proposed bimorph structure further simplifies the device's electrical layout with only two-terminal connections as reported in many conventional unimorph PMUTs. The mechanical and acoustic measurements are conducted to verify the device's performance improvement. The dynamic mechanical displacement and acoustic output under a low driving voltage (1 Vpp) are more than twice that reported from conventional unimorph devices with a similar resonant frequency. Moreover, the pulse-echo experiments indicate an improved receiving voltage of 10 mV in comparison with the unimorph counterpart (4.8 mV). The validation of device advancement in the electromechanical coupling effect by using highly doped ScAlN thin film, the realization of the proposed bimorph PMUT on an 8-inch wafer paves the path to production of next generation, high-performance piezoelectric MEMS.

Beyond fundamental resonance mode: high-order multi-band ALN PMUT for in vivo photoacoustic imaging.

This paper reports on an aluminum nitride (AlN) piezoelectric micromachined ultrasound transducer (PMUT) array for photoacoustic (PA) imaging, where the high-order resonance modes of the PMUT are utilized to improve imaging resolution. A flexural vibration mode (FVM) PMUT is fabricated and applied in a photoacoustic imaging (PAI) system. Specifically, the microelectromechanical system (MEMS)-based PMUT is suitable for PA endoscopic imaging of blood vessels and bronchi due to its miniature size and high sensitivity. More importantly, AlN is a nontoxic material, which makes it harmless for biomedical applications. In the PAI system, the AlN PMUT array is used to detect PA signals, and the acousto-mechanical response is designed and optimized at the PMUT's fundamental resonance. In this work, we focus on the high-order resonance performance of the PMUT PAI beyond the fundamental resonance. The acoustic and electrical responses of the PMUT's high-order resonance modes are characterized and analyzed. The fundamental and three high-order resonance bandwidths are 2.2, 8.8, 18.5, and 48.2 kHz. Compared with the resolution at the fundamental resonance mode, the resolutions at third- and fourth-order resonance modes increase by 38.7% and 76.9% in a phantom experiment. The high-order resonance modes of the AlN PMUT sensor array provide higher central frequency and wider bandwidth for PA signal detection, which increase the resolution of PAI compared to the PMUT working at the fundamental resonance mode.

Development of MEMS Airflow Volumetric Flow Sensing System with Single Piezoelectric Micromachined Ultrasonic Transducer (PMUT) Array.

Compared to conventional ultrasonic flowmeters using multiple transducers, this paper reports, for the first time, an airflow volumetric flowmeter using a signal PMUT array to measure the flow rate in a rectangular pipe. The PMUT around 200 kHz is selected to fit the system requirements. All PMUT elements on this single array are then electrically grouped into transmitter and receiver. In order to minimize the crosstalk signal between transmitter and receiver, a phase shift signal is applied at the transmitter to reduce the amplitude of the crosstalk signal by 87.8%, hence, the resultant high sensing resolution. Based on the analog signal extracted from the single PMUT array, a complete flow sensing system is built by using the cross-correlation method and cosine interpolation, whereby the change in flow rate is reflected by the time of flight difference (dTof) recorded at the receiver. Meanwhile, the acoustic path self-calibration is realized by using multiple echoes. Compared with the previously reported MEMS flowmeters with dual or multiple PMUT devices, this paper proposes a single PMUT array flow sensing system, which is able to measure the flow rate changes up to 4 m3/h. With the implementation of a single device, the problem of ultrasound device/reflector misalignment during system setup is completely eradicated.

Nondestructive Wafer Level MEMS Piezoelectric Device Thickness Detection.

This paper introduces a novel nondestructive wafer scale thin film thickness measurement method by detecting the reflected picosecond ultrasonic wave transmitting between different interfacial layers. Unlike other traditional approaches used for thickness inspection, this method is highly efficient in wafer scale, and even works for opaque material. As a demonstration, we took scandium doped aluminum nitride (AlScN) thin film and related piezoelectric stacking layers (e.g. Molybedenum/AlScN/Molybdenum) as the case study to explain the advantages of this approach. In our experiments, a laser with a wavelength of 515 nm was used to first measure the thickness of (1) a single Molybdenum (Mo) electrode layer in the range of 100-300 nm, and (2) a single AlScN piezoelectric layer in the range of 600-1000 nm. Then, (3) the combined stacking layers were measured. Finally, (4) the thickness of a standard piezoelectric composite structure (Mo/AlScN/Mo) was characterized based on the conclusions and derivation extracted from the aforementioned sets of experiments. This type of standard piezoelectric composite has been widely adopted in a variety of Micro-electromechanical systems (MEMS) devices such as the Piezoelectric Micromachined Ultrasonic Transducer (PMUT), the Film Bulk Acoustic Resonator (FBAR), the Surface Acoustic Wave (SAW) and more. A comparison between measurement data from both in-line and off-line (using Scanning Electron Microscope) methods was conducted. The result from such in situ 8-inch wafer scale measurements was in a good agreement with the SEM data.

AlScN Film Based Piezoelectric Micromechanical Ultrasonic Transducer for an Extended Long-Range Detection.

Piezoelectric micromachined ultrasonic transducers (PMUTs) have been widely applied in distance sensing. However, the sensing distance of currently reported miniaturized ultrasonic sensors (e.g., PMUTs or CMUT) is still limited up to a certain range (e.g., ≤5 m) compared to conventional bulk ultrasonic devices. This paper reports a PMUT array design using scandium-doped aluminum nitride (AlScN) as its piezoelectric layer for an extended long-range detection purpose. To minimize air attenuation, our device is resonating at 66 kHz for a high receive sensitivity of 5.7 mV/Pa. The proposed PMUT array can generate a sound pressure level (SPL) as high as 120 dB at a distance of 10 cm without beam forming. This PMUT design is catered for a pin-to-pin replacement of the current commercial bulk ultrasonic ranging sensor and works directly with the conventional range finding system (e.g., TI PGA460). In comparison with the common bulk transducer, the size of our device is 80% smaller. With the identical ranging detection setup, the proposed PMUT array improves the system SNR by more than 5 dB even at a distance as far as 6.8 m. The result of extended sensing distance validates our miniaturized PMUT array as the optimized candidate for most ultrasonic ranging applications. With the progressive development of piezoelectric MEMS, we believe that the PMUT technology could be a game changer in future long-range sensing applications.

Design and 3D FEM Analysis of a Flexible Piezoelectric Micromechanical Ultrasonic Transducer Based on Sc-Doped AlN Film.

In this paper, a flexible piezoelectric micromachined ultrasonic transducer (PMUT) based on Scandium (Sc)-doped Aluminum Nitride (AlN) film was designed and modeled by the three-dimensional finite element method (3D-FEM). The resonant frequency of 218.1 kHz was reported. It was noticeable that a high effective electromechanical coupling coefficient (k2eff) of 1.45% was obtained when a combination of a flexible PI and a thin Si layer was used as the PMUT supporting structure layer. Compared with a pure Si supporting layer counterpart, the coupling coefficient had been improved by 110.68%. Additionally, the increase of Sc doping concentration in AlN film further enhanced the device electromechanical coupling coefficient and resulted in an improvement for transmitting/receiving sensitivity of the proposed flexible PMUT. When the doping concentration of Sc reached 30%, the emission sensitivity was as large as 1.721 µm/V, which was 2.86 times greater than that of conventional AlN film-based PMUT. The receiving sensitivity was found to be 2.11 V/KPa, which was as high as 1.23 times the performance of an undoped device. Furthermore, the bending simulation result showed that the proposed flexible PMUT device can maintain a good mechanical stability when the bending radius is greater than 1.5 mm. The simulation of sound field characteristics demonstrated that the flexible PMUT based on AlScN could receive stable sound pressure signals under the bending radius of 1.5 cm.

[Clinical Analysis of Elderly Patients with AML/High-Risk MDS].

OBJECTIVE: To analyze the clinical features of acute myeloid leukemia (AML)/high-risk myelodysplastic syndrome (MDS) patients aged over 60 years old. METHODS: The clinical data of 61 elderly newly diagnosed patients with AML and high-risk MDS who submitted to the Department of Hematology/Oncology of the First Affiliated Hospital of Tsinghua University from January 2009 to April 16, 2021 were retrospectively analyzed. These patients were divided into chemotherapy group (45 cases) and supportive treatment group (16 cases). The overall survival (OS) was analyzed by Kaplan-Meier method, and the prognostic factors of survival were analyzed by multivariate Cox regression. RESULTS: After 2 cycles of induction chemotherapy, the complete remission (CR) rate was 37.8% (17/45), and overall response rate was 62.2% (28/45) in the chemotherapy group. The median OS in the chemotherapy group and supportive treatment group was 11.3 (0.07-43) and 1.6 (0.33-7.72) months, respectively (P<0.001). The median OS in patients who reached CR or did not reach after 1 cycle of induction chemotherapy was 19.8 (10-30.63) and 8.17 (0.07-43) months, respectively (P<0.05), while after 2 cycles was 22.7 (4.2-43) and 7.26 (0.07-26) months, respectively (P<0.001). Univariate analysis showed that age > 80 years old, CCI score > 2, PS score > 2 and supportive treatment were the adverse prognostic factors for OS. Further multivariate analysis suggested that chemotherapy was the only independent prognostic factor for OS (HR=0.140, 95%CI: 0.048-0.409, P<0.001). In the chemotherapy group, univariate analysis showed that CCI score > 2 and failure to reach CR after induction chemotherapy were poor prognostic factors. Multivariate analysis showed that CCI score > 2 (HR=0.139, 95%CI: 0.050-0.384, P<0.001) and failure to achieve CR after induction chemotherapy (HR=0.103, 95%CI: 0.041-0.259, P<0.001) were the adverse prognostic factors for OS. The patients were tolerant to side-effect of chemotherapy. CONCLUSION: Appropriate chemotherapy can prolong the survival of elderly patients with AML and high-risk MDS.

Whole exome sequencing identifies novel inherited genetic variants in tetralogy of Fallot.

Background: Tetralogy of Fallot (TOF) is the most common neonatal cyanotic heart defect, and genetic variation is an important risk factor for the etiology of TOF. Identifying TOF-associated genetic variants is critical to understanding susceptibility and outcome in patients with TOF and may help delineate pathological mechanisms. Methods: Whole exome sequencing (WES) was performed 19 patients with sporadic TOF and 3 healthy controls. The dbSNP, GnomAD, Denovo-db, and ClinVar databases were used to annotate the mutations. PolyPhen, SIFT, MutationTaster, and FATHMM softwares were used for mutation pathogenicity analysis. Sanger sequencing was used to validate candidate variants. Results: We identified 21 genetic variants involving 16 genes were found in 12 patients with sporadic TOF. The types of mutations were missense and splicing variants. None of these genes were detected in samples from the 3 healthy controls. These variants include 9 pathogenic variants, 6 suspected pathogenic variants, and 6 variants of unknown significance (VUS). Further analysis showed that the patients with apolipoprotein B (APOB) and ring finger protein 135 (RNF135) variants had more serious clinical symptoms. Sanger sequencing confirmed that the two variants were heterozygous in TOF patients. Conclusions: We identified several genetic variants associated with TOF and confirmed that RNF135 and ABOB variants were associated with TOF severity. These findings provide new evidence for exploring the genetic mechanism of TOF.

Lactobacillus casei Ghosts as a Vehicle for the Delivery of DNA Vaccines Mediate Immune Responses.

We developed Lactobacillus casei bacterial ghosts (BGs) as vehicles for delivering DNA vaccines and analyzed their effects on immune responses. Uptake of the plasmids encoding the enhanced green fluorescent protein (pCI-EGFP) and BGs loaded with pCI-EGFP by macrophages was investigated using fluorescence microscopy and flow cytometry. The results showed that pCI-EGFP-loaded L. casei BGs were efficiently taken up by macrophages. Lactobacillus casei BGs loaded with plasmids encoding VP6 protein of PoRV (pCI-PoRV-VP6) significantly upregulated the mRNA expression of interleukin (IL)-1ß, IL-10, tumor necrosis factor (TNF)-α, inducible nitric oxide synthase (iNOS), arginase-1 (Arg-1), Mannose receptor (CD206) toll-like receptor (TLR)-2, TLR4, and TLR9 in macrophages. The levels of markers of M1 polarization (IL-10 and TNF-α) and M2 polarization (Arg-1 and CD206) were increased in macrophages incubated with pCI-PoRV-VP6-loaded BGs compared with the control group. The results of the enzyme-linked immunosorbent assay showed that the secretion of IL-1ß, IL-10, and TNF-α in macrophages was significantly upregulated compared with the control group. Flow cytometry demonstrated that L. casei BGs loaded with pCI-PoRV-VP6 promoted the maturation of dendritic cells (DCs). Following incubation with pCI-PoRV-VP6-loaded BGs, the mRNA expression levels of IL-1ß, IL-6 and interferon (IFN)-γ in DCs were significantly increased. ELISA assay showed the secretion of the IL-1ß, IL-6, IFN-γ IL-10 and TNF-α in DCs were upregulated significantly. Thus, L. casei BGs promoted the maturation and activation of DCs. We analyzed the stimulatory capacity of DCs in a mixed lymphocyte reaction with allogeneic T cells. T cell proliferation increased upon incubation with DCs stimulated by BGs. After immunizing mice with BGs loaded with pCI-PoRV-VP6, the specific IgG levels in the serum were higher than those elicited by BGs loaded with pCI-PoRV-VP6. BGs loaded with pCI-PoRV-VP6 on Th1 and Th2 cytokines polarized T cells into the Th1 type and increased the proportion of CD4+/CD8+ T cells. These results indicate L. casei BGs effectively mediate immune responses and can be used as delivery system for DNA vaccination.

Efficacy Assessment of Phage Therapy in Treating Staphylococcus aureus-Induced Mastitis in Mice.

The primary aim of this study was to evaluate the efficacy of phage against mastitis induced by drug-resistant S. aureus in a mouse model. In this study, five S. aureus phages-4086-1, 4086-2, 4086-3, 4086-4, and 4086-6-were isolated from milk samples secreted by mastitis cows. Transmission electron microscopy showed that all the five phages had icosahedral heads and short non-contractile tails, which are typical characteristics of the family Podoviridae. All these phages were species-specific against S. aureus. The one-step growth curve showed a short latency period (10-20 min) and high burst size (up to 400 PFU/infected cell). To evaluate the effectiveness of the phage 4086-1 in the treatment against mastitis, a mouse model of mastitis was challenged with drug-resistant S. aureus. The results showed the proliferation of S. aureus in the mammary glands was significantly inhibited after treating by phage 4086-1. The concentrations of TNF-α and IL-6 decreased significantly, which demonstrated the phages could effectively alleviate the inflammatory responses. Furthermore, the histopathological analysis showed that inflammatory infiltration in the mammary glands was significantly reduced. These results demonstrate that phage may be a promising alternative therapy against mastitis caused by drug-resistant S. aureus.