An Improved Test Method for Assaying the Inhibition of Bioflavonoids on Xanthine Oxidase Activity in†vitro.

The difference on inhibitory effects of bioflavonoids inhibiting XOD activity assayed by varying test methods cause of us to be further in consideration. The reported test method creating a micro-environment surrounding XOD in the absence of ⋅O2 -, which is seemly different from the assay in vivo. So, the vitro test method for assaying XOD activity is necessary to be improved for selection of potential inhibitors in the presence of ⋅O2 -. The inhibitory results demonstrated that bioflavonoids of MY, DMY, QUE and LUT are capable to be on effective IC50 values, but others are not. As well, their resulting inhibitions determined by the improved test method are much less than that reported in the literature, indicating that their chemical affinities with XOD become weaker. Moreover, DMY assayed on the inhibitions of XOD in the improved test method performs to be a better inhibitor, as compared to the assay of the reported test methods. Abasing on the transformation of DMY into MY in the presence of ⋅O2 -, the good inhibition of DMY on XOD activity can be explained by the synergistic effect of MY.

Camera-in-the-loop based test scenario generation method for pedestrian collision avoidance system.

The Pedestrian Collision Avoidance System (PCAS) of Intelligent Vehicle (IV) can be effective in preventing the occurrence of traffic accidents. However, the complicated operation environments introduce great challenges to the camera used by the PCAS. Therefore, the camera based PCAS should be fully tested and evaluated before deployment. The traditional simulation test for the camera based PCAS attempted to use geometric or physical simulation models, which have low reality and are suitable for the primary stage of the PCAS development. Camera-in-the-Loop (CIL) test is one of Hardware-in-the-Loop methods that embeds the real camera hardware into the virtual simulation system to test the camera. CIL can utilize the real hardware response while overcoming the common simulation weakness of fidelity. In this paper, we construct a CIL test platform, and propose the CIL based test scenarios generation and scenario parameter impact evaluation method for PCAS. First, we construct the CIL test platform whose image quality and functional confidence are both validated to prove CIL credibility. Second, the PCAS under the test is analyzed and the corresponding test scenario parameters are designed. In order to accelerate the test scenario generation, a Greedy Based Combination test method (GBC) based on the CIL is proposed. The Chi-square analysis and two-factor of variance analysis verification methods are used to analyze the influence of individual and multiple scenario parameters on the PCAS performance. The experiment results show that the GBC improves the test speed by 12 times compared to the traversal test, and the frequency ratio of each scenario parameter is no more than 3% different from that of the traversal test. Also, GBC has an equivalent ability to find the PCAS collision scenarios parameter combination to the traversal test.

Influence parental- and child-related factors on the acceptance of SARS-CoV-2 test methods in schools and daycare facilities.

Introduction: Rapid testing for Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) infections was an essential step in reducing the spread of the virus and monitoring pandemic development. Most mandatory standard pandemic testing in Germany has been performed in schools and daycare facilities. We investigated the influence of behavioral and attitudinal characteristics of children and caregivers on their acceptance of (i) antigen-based nasal swab rapid and (ii) oral saliva-based pooled Polymerase Chain Reaction (PCR) tests. Methods: Conducted through a cross-sectional survey between November and December 2021, with 1962 caregivers and 581 children/adolescents participating, the study evaluated the acceptability of each testing method on a six-point scale. Participants scored one test method conducted on their child at one of six levels with 1 and 6 denoting "excellent" (1) and "inadequate" (6), respectively. We considered demographic variables, vaccination status, child mental health (measured by the SDQ-questionnaire), and facility type (kindergarten, primary school, secondary school) as covariates. Results: Results reveal a preference for saliva-based PCR tests over nasal swabs by about one grade, particularly among parents of unvaccinated children, especially if their child expressed future vaccination reluctance. Testing acceptance was lower among children with mental health issues, primary school-aged, and those with less-educated parents. Perception of test accuracy and convenience influenced attitudes, favoring saliva-based PCR tests. Moreover, children with mental health issues felt less secure during testing. Discussion: To our knowledge, this is the first study to investigate the influence of different testing methods on testing acceptance for SARS-CoV-2 in children and caregivers. Our study identifies predictors of lower acceptance of public health surveillance measures and enables the development of educational programs on testing and vaccination tailored to the needs of specific target groups. Moreover, we demonstrate that test acceptance in vulnerable groups can be enhanced by careful choice of an appropriate testing method.

Developing an optimised method for accurate wear testing of dental materials using the 'Rub&Roll' device.

Dental materials are challenged by wear processes in the oral environment and should be evaluated in laboratory tests prior to clinical use. Many laboratory wear-testing devices are high-cost investments and not available for cross-centre comparisons. The 'Rub&Roll' wear machine enables controlled application of force, chemical and mechanical loading, but the initial design was not able to test against rigid antagonist materials. The current study aimed to probe the sensitivity of a new 'Rub&Roll' set-up by evaluating the effect of force and test solution parameters (deionized water; water + abrasive medium; acid + abrasive medium) on the wear behaviour of direct and indirect dental resin-based composites (RBCs) compared with human molars against 3D-printed rod antagonists. Molars exhibited greater height loss than RBCs in all test groups, with the largest differences recorded with acidic solutions. Direct RBCs showed significantly greater wear than indirect RBCs in the groups containing abrasive media. The acidic + abrasive medium did not result in increased wear of RBC materials. The developed method using the 'Rub&Roll' wear machine in the current investigation has provided a sensitive wear test method to allow initial screening of resin-based composite materials compared with extracted human molars under the influence of different mechanical and erosive challenges.

Raman spectroscopy as an alternative rapid microbial bioburden test method for continuous, automated detection of contamination in biopharmaceutical drug substance manufacturing.

AIMS: To investigate an in-line Raman method capable of detecting accidental microbial contamination in pharmaceutical vessels, such as bioreactors producing monoclonal antibodies via cell culture. METHODS AND RESULTS: The Raman method consists of a multivariate model built from Raman spectra collected in-line during reduced-scale bioreactor batches producing a monoclonal antibody, as well as a reduced-scale process with intentional spiking of representative compendial method microorganisms (n = 4). The orthogonal partial least squares regression discriminant analysis model (OPLS-DA) area under the curve (AUC), specificity and sensitivity were 0.96, 0.99, and 0.95, respectively. Furthermore, the model successfully detected contamination in an accidentally contaminated manufacturing-scale batch. In all cases, the time to detection (TTD) for Raman was superior compared to offline, traditional microbiological culturing. CONCLUSIONS: The Raman OPLS-DA method met acceptance criteria for equivalent decision making to be considered a viable alternative to the compendial method for in-process bioburden testing. The in-line method is automated, non-destructive, and provides a continuous assessment of bioburden compared to an offline compendial method, which is manual, results in loss of product, and in practice is only collected once daily and requires 3-5 days for enumeration.

Screening of environmental stimuli for the positive regulation of stomatal aperture in centipedegrass.

Grasslands, the largest carbon pool in China, possess enormous potential for carbon sequestration. Increasing the stomatal aperture to increase the CO2 absorption capacity is a potential method to improve plant photosynthetic efficiency and ultimately enhance the carbon sequestration capacity of grass plants. Research on stomatal aperture regulation has focused mostly on Arabidopsis or crops, while research on grass plants in these areas is scarce, which seriously restricts the implementation of this grassland carbon sequestration strategy. Here, a widely used ecological grass, centipedegrass, was used as the experimental material. First, a convenient method for observing the stomatal aperture was developed. The leaves were floated in a potassium ion-containing open solution (67 mM KCl, pH 6.0) with the adaxial surface rather than the abaxial surface in contact with the solution and were cultivated under light for 1.5 h. Then, nail polish was applied on the adaxial surface, and a large number of open stomata were imprinted. Second, with the help of this improved method, the concentration‒response characteristics of the stomatal aperture to eleven environmental stimuli were tested. The stomatal aperture is dependent on these environmental stimuli in a concentration-dependent manner. The addition of 100 µM brassinolide led to the maximal stomatal aperture. This study provided a technical basis for manipulating stomatal opening to increase the carbon sequestration capacity of centipedegrass.

A Novel image processing technique for weighted particle size distribution assessment.

The objective of the study was to create a reliable method that could be used to evaluate the particle size distribution of samples and pre-mixes in real-world situations, particularly those consisting of typical formulation blends. The goal was to use this method to assess the uniformity of the samples and ensure that they met the required quality standards. The researchers aimed to create a method that could be easily incorporated into the manufacturing process, providing a practical and efficient solution. This study demonstrates the use of ImageJ software to analyze the particle size distribution (PSD) of powders. The technique produces qualitative data from microscopy images and quantitative data from analysis of parameters including average diameter, D10, D50, D90, and standard deviation. The method was tested with various treatments, showing differentiating outcomes in all cases. The alternate technique provides a rapid and cost-effective method for PSD analysis, surpassing the limitations of sieve analysis. Extensive testing of the method, using a variety of sample types, including typical formulation blends, was performed. The results suggest that the method can effectively assess the morphology of changing materials during batch manufacturing and characterize uniformity in blends. The methodology has the capability to identify attributes related to PSD that are typically required to be monitored during manufacturing. The technique allows for accurate and reliable quantification of the attributes through image capture technology. The technique has future potential and has important implications for material science, powder rheology, pharmaceutical formulation development, and continual process monitoring.

[Figure: see text]A Novel Image Processing Technique for Weighted Particle Size Distribution Assessment.

A Novel Temperature Drift Error Estimation Model for Capacitive MEMS Gyros Using Thermal Stress Deformation Analysis.

Because the conventional Temperature Drift Error (TDE) estimation model for Capacitive MEMS Gyros (CMGs) has inadequate Temperature Correlated Quantities (TCQs) and inaccurate parameter identification to improve their bias stability, its novel model based on thermal stress deformation analysis is presented. Firstly, the TDE of the CMG is traced precisely by analyzing its structural deformation under thermal stress, and more key decisive TCQs are explored, including ambient temperature variation ∆T and its square ∆T2, as well its square root ∆T1/2; then, a novel TDE estimation model is established. Secondly, a Radial Basis Function Neural Network (RBFNN) is applied to identify its parameter accurately, which eliminates local optimums of the conventional model based on a Back-Propagation Neural Network (BPNN) to improve bias stability. By analyzing heat conduction between CMGs and the thermal chamber with heat flux analysis, proper temperature control intervals and reasonable temperature control periods are obtained to form a TDE precise test method to avoid time-consuming and expensive experiments. The novel model is implemented with an adequate TCQ and RBFNN, and the Mean Square Deviation (MSD) is introduced to evaluate its performance. Finally, the conventional model and novel model are compared with bias stability. Compared with the conventional model, the novel one improves CMG's bias stability by 15% evenly. It estimates TDE more precisely to decouple Si-based materials' temperature dependence effectively, and CMG's environmental adaptability is enhanced to widen its application under complex conditions.

The inter-laboratory validation study of EpiSensA for predicting skin sensitization potential.

The Epidermal Sensitization Assay (EpiSensA) is a reconstructed human epidermis (RhE)-based gene expression assay for predicting the skin sensitization potential of chemicals. Since the RhE model is covered by a stratified stratum corneum, various kinds of test chemicals, including lipophilic ones and pre-/pro-haptens, can be tested with a route of exposure akin to an in vivo assay and human exposure. This article presents the results of a formally managed validation study of the EpiSensA that was carried out by three participating laboratories. The purpose of this validation study was to assess transferability of the EpiSensA to new laboratories along with its within- (WLR) and between-laboratory reproducibility (BLR). The validation study was organized into two independent stages. As demonstrated during the first stage, where three sensitizers and one non-sensitizer were correctly predicted by all participating laboratories, the EpiSensA was successfully transferred to all three participating laboratories. For Phase I of the second stage, each participating laboratory performed three experiments with an identical set of 15 coded test chemicals resulting in WLR of 93.3%, 93.3%, and 86.7%, respectively. Furthermore, when the results from the 15 test chemicals were combined with those of the additional 12 chemicals tested in Phase II of the second stage, the BLR for 27 test chemicals was 88.9%. Moreover, the predictive capacity among the three laboratories showed 92.6% sensitivity, 63.0% specificity, 82.7% accuracy, and 77.8% balanced accuracy based on murine local lymph node assay (LLNA) results. Overall, this validation study concluded that EpiSensA is easily transferable and sufficiently robust for assessing the skin sensitization potential of chemicals.

Pozzolanic activity experimental dataset of calcined coal gangue.

The coal gangue in this dataset was subjected to a series of processes, including drying, crushing, and milling. Subsequently, the coal gangue powder was subjected to high-temperature calcination in a muffle furnace, with a heating rate of 4 â„ƒ/min. The pozzolanic activity of coal gangue powder was investigated at various calcination temperatures (600 â„ƒ, 700 â„ƒ, 800 â„ƒ, 900 â„ƒ) and different holding times (1h, 2h). Cement mortar specimens containing calcined coal gangue powder were prepared, and their compressive and flexural strengths were tested to evaluate the reactivity of the calcined coal gangue. In addition, the Rapid, Relevant and Reliable (R3) activity test was conducted to test the reactivity. The thermogravimetric analyzer was employed to determine the TG-DTG curves of coal gangue powder. X-ray diffractometer, Fourier infrared spectrometer and scanning electron microscope were utilized to investigate the microstructure of activated coal gangue powder at different temperature ranges. These data can be used for determining the optimal calcination scheme of coal gangue to maximize its potential as a partial cement clinker replacement in cement production, thereby contributing to cost reduction and carbon emission mitigation.

The effective doses of remimazolam besylate in the procedural sedation of endoscopic retrograde cholangiopancreatography.

This study aimed to determine the values of the half-effective dose (ED50) and 95% effective dose (ED95) of remimazolam besylate used in the procedural sedation of endoscopic retrograde cholangiopancreatography (ERCP). Sixty patients who fulfilled the inclusion and exclusion criteria of this study were selected. Sufentanil was administered intravenously and remimazolam besylate was administered 2 min later. ERCP treatment was feasible when the modified alertness/sedation (MOAA/S) score was ≤2. If choking or movement occurred during duodenoscope placement, it was considered as a positive reaction. The dose was increased in the next patient; otherwise, it was considered as a negative reaction, and the dose was reduced in the next patient. The ED50 and ED95 values and 95% confidence interval (CI) of remimazolam besylate were calculated by Probit regression analysis. All 60 patients completed the trial. The ED50 and ED95 values of remimazolam besylate were 0.196 and 0.239 mg/kg, respectively, for the procedural sedation of ERCP. The time of MOAA/S score ≤ 2 was (82.58 ± 21.70) s, and the mean time of awakening was (9.03 ± 5.64) min. Transient hypotension was observed in two patients without medical intervention. The ED50 and ED95 values of remimazolam besylate used in the procedural sedation of ERCP were 0.196 and 0.239 mg/kg, and the dose of the medications has definite efficacy and good safety.

Evaluation of TIB Molbiol LightMix® assays for detection of Mycoplasma genitalium and key resistance mutations for macrolides and fluoroquinolones.

The LightMix® Modular Mycoplasma Macrolide and LightMix® Modular parC Fluoroquinolone Resistance assays (TIB Molbiol) were evaluated using sequential Mycoplasma genitalium positive (n = 125) and negative (n = 93) clinical samples. Results were compared to the results of an established commercial assay (ResistancePlus MG assay, SpeeDx Pty Ltd) or Sanger sequencing (for parC). Detection of M. genitalium by the TIB Molbiol assay had a high agreement with the reference assay, with a positive percent agreement (PPA) of 97.6 [95% confidence interval (CI): 93.1-99.5] and negative percent agreement (NPA) of 95.7 (95% CI: 89.5-98.8). From 105 positive samples, macrolide resistance detection had a PPA of 100% (95% CI: 93.7-100) and NPA of 81.3% (95% CI: 67.4-91.1). For the detection of fluroquinolone resistance mutation G248T/S83I or "other mutation" in the quinolone resistance determinant region, from 95 samples there was 100% (95% CI: 86.3-100) sensitivity and 100% (95% CI: 94.5-100) specificity. The understanding of the basis for fluoroquinolone treatment failure is still developing; it is therefore important to use the output of parC-based resistance assays with caution to avoid the inappropriate use of antibiotic therapies, especially considering the limited number of alternative treatments.

Present-Day Tectonic Stress Evolution in Southern Yunnan Based on Focal Mechanisms.

Tectonic extrusion bypassing the eastern Himalayan syntaxis results in a significant increase in regional stress instability and the associated frequent occurrence of earthquakes in southern Yunnan, China. However, the stress field, and the relationship between the focal mechanism of earthquakes and stress evolution in southern Yunnan, remain enigmatic. In this paper, using a modified grid point test method, we calculated the focal mechanism of ML ≥ 2.5 earthquakes in southern Yunnan (22-25° N, 100-104° E) from January 2009 to June 2023. Utilizing the solutions of historical earthquake focal mechanisms, we obtained the present-day regional tectonic stress field in southern Yunnan via inversion. The results indicate complex and diverse seismic focal mechanisms, and the main types of earthquakes are strike-slip events, followed by normal fault and reverse fault events. The orientations of the maximum and minimum principal stress axes rotate in a clockwise direction from northeast to southwest. The internal stress orientation distribution of the rhombic Sichuan-Yunnan block in the study area is consistent, and the block boundary zone is the site where stress deflection occurs, and the regional tectonic stress field is influenced by the interaction among different blocks. The distribution of R-value in the Lamping-Simao block gradually increases from north to south, indicating that the compressive stress required for material transport becomes relatively small. Combined with the geological and tectonic background of the study area, our results suggest that the speed of block movement gradually decreases from north to south; the distribution of R-value in the South China block is significantly smaller than that of the interior of the Sichuan-Yunnan rhombus, and the proportion of compressive stresses is larger, indicating a stronger extrusion in this region, which may be related to the fact that the Sichuan-Yunnan rhombus is strongly resisted by the South China block in the east.

Investigation and Improvement of Test Methods for Capacitance and DCESR of EDLC Cells.

The quick and accurate characterization of commercial electrochemical double-layer capacitor (EDLC) cells, especially their capacitance and direct-current equivalent series internal resistance (DCESR), is of great significance for the design, maintenance, and monitoring of EDLCs used in areas of energy, sensors, electric power, construction machinery, rail transit, automobile transportation, and military. In this study, the capacitance and DCESR of three commercial EDLC cells with similar performance were determined and compared by following the three commonly-used standards of IEC 62391, Maxwell, and QC/T741-2014, which are significantly different in test procedures and calculation methods. The analysis of the test procedures and results demonstrated that the IEC 62391 standard has the disadvantages of a large testing current, long testing time, and a complex and inaccurate DCESR calculation, whereas the Maxwell standard has the disadvantages of a large testing current, a small capacitance, and large DCESR testing results, and furthermore the QC/T 741 standard has the disadvantages of a high resolution requirement for the equipment and small DCESR results. Therefore, an improved method was proposed to determine the capacitance and DCESR of EDLC cells by short-time constant voltage charging and discharging interruption methods, respectively, with the advantages of high accuracy, low equipment requirements, short testing time, and the easy calculation of DCESR over the original three standards.

Recent Progress in Testing and Characterization of Hardenability of Aluminum Alloys: A Review.

In this paper, the progress of the test methods and characterization approaches of aluminum alloys hardenability was reviewed in detail. The test method mainly included the traditional end-quenching method and the modified method. While the characterization approaches of alloy hardenability consist mainly of ageing hardness curves, solid solution conductivity curves, ageing tensile curves, time temperature transformation (TTT) curves, time temperature properties (TTP) curves, continuous cooling transformation (CCT) curves, and advanced theoretical derivation method have appeared in recent years. The hardenability testing equipment for different tested samples with different material natures, engineering applications properties, and measurement sizes was introduced. Meanwhile, the improvement programmed proposed for shortcomings in the traditional hardenability testing process and the current deficiencies during the overall hardenability testing process were also presented. In addition, the influence factors from the view of composition design applied to the hardenability behaviors of Aluminum alloys were summarized. Among them, the combined addition of micro-alloying elements is considered to be a better method for improving the hardenability of high-strength aluminum alloys.

Chloride Transport and Related Influencing Factors of Alkali-Activated Materials: A Review.

Chloride transport is a vital issue in the research on the durability of alkali-activated materials (AAMs). Nevertheless, due to its miscellaneous types, complex mix proportions, and limitations in testing methods, the reports of different studies are numerous and vary greatly. Therefore, in order to promote the application and development of AAMs in chloride environments, this work systematically reviews the chloride transport behavior and mechanism, solidification of chloride, influencing factors, and test method of chloride transport of AAMs, along with conclusions regarding instructive insights to the chloride transport problem of AAMs in future work.

Quantitation of Mycoplasma genitalium using droplet digital PCR.

Mycoplasma genitalium is a sexually transmitted infection with increasing concerns around antimicrobial resistance. Droplet digital PCR (ddPCR) is a rapid quantification method with high precision that may be useful for absolute quantitation of bacteria in samples. This study aimed to develop a ddPCR assay for the quantification of M. genitalium. ddPCR targeting the gene mgpB was established and analysed using the QX100 ddPCR system. The assay was evaluated against quantitated DNA standards, and then in comparison to an established quantitative PCR performed on the Lightcycler 480 II. DNA template of increasing complexity was used, including synthetic double stranded DNA, DNA extracts from laboratory-cultured M. genitalium strains (n = 17) and DNA from M. genitalium-positive clinical samples (n = 21). There was a strong correlation between ddPCR concentration estimates and measured DNA standards (r2 = 0.997), and between ddPCR and qPCR quantitation for different templates (r2 ranging from 0.953 to 0.997). ddPCR reliably detected template in a range from <10 copies per reaction to >104 copies per reaction and demonstrated linearity over dilution series. Concentration estimates by ddPCR were reproducibly less than those determine by qPCR. ddPCR demonstrated precise and reproducible quantitation of M. genitalium with a variety of templates.

The sensitivity of the zebrafish embryo coiling assay for the detection of neurotoxicity by compounds with diverse modes of action.

In the aim to determine neurotoxicity, new methods are being validated, including tests and test batteries comprising in vitro and in vivo approaches. Alternative test models such as the zebrafish (Danio rerio) embryo have received increasing attention, with minor modifications of the fish embryo toxicity test (FET; OECD TG 236) as a tool to assess behavioral endpoints related to neurotoxicity during early developmental stages. The spontaneous tail movement assay, also known as coiling assay, assesses the development of random movement into complex behavioral patterns and has proven sensitive to acetylcholine esterase inhibitors at sublethal concentrations. The present study explored the sensitivity of the assay to neurotoxicants with other modes of action (MoAs). Here, five compounds with diverse MoAs were tested at sublethal concentrations: acrylamide, carbaryl, hexachlorophene, ibuprofen, and rotenone. While carbaryl, hexachlorophene, and rotenone consistently induced severe behavioral alterations by ~ 30 h post fertilization (hpf), acrylamide and ibuprofen expressed time- and/or concentration-dependent effects. At 37-38 hpf, additional observations revealed behavioral changes during dark phases with a strict concentration-dependency. The study documented the applicability of the coiling assay to MoA-dependent behavioral alterations at sublethal concentrations, underlining its potential as a component of a neurotoxicity test battery.

Derivation of Corrosion Depth Formula According to Corrosion Factors in District Heating Water through Regression Analysis.

In order to predict the corrosion depth of a district heating pipeline, it is necessary to analyze various corrosion factors. In this study, the relationship between corrosion factors such as pH, dissolved oxygen, and operating time and corrosion depth was investigated using the Box-Behnken method within the response surface methodology. To accelerate the corrosion process, galvanostatic tests were conducted in synthetic district heating water. Subsequently, a multiple regression analysis was performed using the measured corrosion depth to derive a formula for predicting the corrosion depth as a function of the corrosion factors. As a result, the following regression formula was derived for predicting the corrosion depth: "corrosion depth (µm) = -133 + 17.1 pH + 0.00072 DO + 125.2 Time - 7.95 pH × Time + 0.002921 DO × Time".

Rapid Screening and Comparison of Chimeric Lysins for Antibacterial Activity against Staphylococcus aureus Strains.

Chimeric lysins composed of various combinations of cell wall-lysing (enzymatic) and cell-wall-binding (CWB) domains of endolysins, autolysins, and bacteriocins have been developed as alternatives to or adjuvants of conventional antibiotics. The screening of multiple chimeric lysin candidates for activity via E. coli expression is not cost effective, and we previously reported on a simple cell-free expression system as an alternative. In this study, we sufficiently improved upon this cell-free expression system for use in screening activity via a turbidity reduction test, which is more appropriate than a colony reduction test when applied in multiple screening. Using the improved protocol, we screened and compared the antibacterial activity of chimeric lysin candidates and verified the relatively strong activity associated with the CHAP (cysteine, histidine-dependent amidohydrolase/peptidase) domain of secretory antigen SsaA-like protein (ALS2). ALS2 expressed in E. coli showed two major bands, and the smaller one (subprotein) was shown to be expressed by an innate downstream promoter and start codon (ATG). The introduction of synonymous mutations in the promoter resulted in clearly reduced expression of the subprotein, whereas missense mutations in the start codon abolished antibacterial activity as well as subprotein production. Interestingly, most of the S. aureus strains responsible for bovine mastitis were susceptible to ALS2, but those from human and chicken were less susceptible. Thus, the simple and rapid screening method can be applied to select functional chimeric lysins and define mutations affecting antibacterial activity, and ALS2 may be useful in itself and as a lead molecule to control bovine mastitis.