bluuebunny/crossref_metadata_2025 huggingface dataset:
Event-Based Optical Flow Estimation with Spatio-Temporal Backpropagation Trained Spiking Neural Network
Authors: Zhang Yisa , Lv Hengyi , Zhao Yuchen , Feng Yang , Liu Hailong , Bi Guoling
Published: January 2023

The advantages of an event camera, such as low power consumption, large dynamic range, and low data redundancy, enable it to shine in extreme environments where traditional image sensors are not competent, especially in high-speed moving target capture and extreme lighting conditions. Optical flow reflects the target’s movement information, and the target’s detailed movement can be obtained using the event camera’s optical flow information. However, the existing neural network methods for optical flow prediction of event cameras has the problems of extensive computation and high energy consumption in hardware implementation. The spike neural network has spatiotemporal coding characteristics, so it can be compatible with the spatiotemporal data of an event camera. Moreover, the sparse coding characteristic of the spike neural network makes it run with ultra-low power consumption on neuromorphic hardware. However, because of the algorithmic and training complexity, the spike neural network has not been applied in the prediction of the optical flow for the event camera. For this case, this paper proposes an end-to-end spike neural network to predict the optical flow of the discrete spatiotemporal data stream for the event camera. The network is trained with the spatio-temporal backpropagation method in a self-supervised way, which fully combines the spatiotemporal characteristics of the event camera while improving the network performance. Compared with the existing methods on the public dataset, the experimental results show that the method proposed in this paper is equivalent to the best existing methods in terms of optical flow prediction accuracy, and it can save 99% more power consumption than the existing algorithm, which is greatly beneficial to the hardware implementation of the event camera optical flow prediction., laying the groundwork for future low-power hardware implementation of optical flow prediction for event cameras.
On-Chip Free-Flow Measurement Revealed Possible Depletion of Macrophages by Indigestible PM2.5 within a Few Hours by the Fastest Intervals of Serial Phagocytosis
Authors: Horonushi Dan , Furumoto Yuya , Nakata Yoshiki , Azuma Toshiki , Yoshida Amane , Yasuda Kenji
Published: January 2023

To understand the influence of indigestible particles like particulate matter 2.5 (PM2.5) on macrophages, we examined the time course of the series phagocytosis of indigestible 2 μm polystyrene spheres (PS). Five kinds of antigens were used as samples for phagocytosis; Zymosan, non-coated 2 μm PS, bovine serum albumin (BSA)-coated PS (BSA-PS), IgG-coated PS (IgG-PS), and IgG-BSA-coated PS (IgG/BSA-PS). To keep the surrounding concentration of antigens against single macrophages constant, antigens flowed at a continuous rate of 0.55 μm/s within a culture dish as a free-flow measurement assay (on-chip free-flow method). The interval of series phagocytosis for IgG/BSA-PS was the shortest among five samples; it was six times faster than Zymosan in terms of engulfment frequency, and up to 50 particles were engulfed within two hours, maintaining constant intervals until reaching the maximum number. The rate of increase in the total number of phagocytozed IgG/BSA-PS over time was constant, at 1.5 particles/min, in series phagocytosis with a 33-cell population, indicating that the phagocytosis rate constant remained constant independent of the number of phagocytoses. Reaction model fitting of the results showed that IgG/BSA-PS had the highest efficiency in terms of the phagocytosis rate constant, 2.3 × 10−2 particles/min, whereas those of IgG-PS, BSA-PS, PS, and Zymosan were 1.4 × 10−2, 1.1 × 10−2, 4.2 × 10−3, and 3.6 × 10−3 particles/min, respectively. One-by-one feeding of IgG/BSA-PS with optical tweezers was examined to confirm the phagocytosis intervals, and we found that the intervals remained constant until several times before the maximum number of antigens for engulfment, also indicating no change in the phagocytosis rate constant regardless of the history of former phagocytosis and phagocytosis number. Simultaneous phagocytosis of two IgG-BSA-decorated microneedle engulfments also showed that the initiation and progress of two simultaneous engulfments on the two different places on a cell were independent and had the same elongation velocity. Therefore, each phagocytosis of indigestible antigens does not affect both in series or in simultaneous subsequent phagocytosis until reaching the maximum capacity of the phagocytosis number. The results suggest (1) no change in the phagocytosis rate constant regardless of the history of phagocytosis numbers and attachment timing and positions, and (2) IgG-BSA decoration of indigestible microparticles in blood accelerates their engulfment faster, resulting in a severe shortage of macrophages within the shortest time.
Fabrication and DC-Bias Manipulation Frequency Characteristics of AlN-Based Piezoelectric Micromachined Ultrasonic Transducer
Authors: Li Tao , Zhang Le , Geng Wenping , He Jian , Rao Yongkang , Huo Jiabing , Yan Kunxian , Chou Xiujian
Published: January 2023

Due to their excellent capabilities to generate and sense ultrasound signals in an efficient and well-controlled way at the microscale, piezoelectric micromechanical ultrasonic transducers (PMUTs) are being widely used in specific systems, such as medical imaging, biometric identification, and acoustic wireless communication systems. The ongoing demand for high-performance and adjustable PMUTs has inspired the idea of manipulating PMUTs by voltage. Here, PMUTs based on AlN thin films protected by a SiO2 layer of 200 nm were fabricated using a standard MEMS process with a resonant frequency of 505.94 kHz, a −6 dB bandwidth (BW) of 6.59 kHz, and an electromechanical coupling coefficient of 0.97%. A modification of 4.08 kHz for the resonant frequency and a bandwidth enlargement of 60.2% could be obtained when a DC bias voltage of −30 to 30 V was applied, corresponding to a maximum resonant frequency sensitivity of 83 Hz/V, which was attributed to the stress on the surface of the piezoelectric film induced by the external DC bias. These findings provide the possibility of receiving ultrasonic signals within a wider frequency range, which will play an important role in underwater three-dimensional imaging and nondestructive testing.
A Calibration Method for the Resolution of 2D TPP Laser Direct Writing
Authors: Xie Yu , Chen Yixiong , Xu Hang , Chen Jianxiong
Published: January 2023

To improve the fabrication efficiency of the two-photon polymerization (TPP) laser direct writing, the TPP exposure process was set to complete by a single-line scan, which was named 2D TPP. The voxel of the 2D TPP should be large enough to cross the photoresist and the underlayer. To explore the resolution limit of the 2D TPP considering the thickness of the photoresist, a new method named the 45° scanning method was proposed. Meanwhile, a two-photon micro-nano fabrication platform was developed. A group of experiments based on the orthogonal decomposition method was carried out to analyze the width and length of the voxel on the S1805 photoresist under different laser power and processing speed. To confirm whether the resolution of the micro-nano structures fabricated by 2D TPP is consistent with the width of the voxel, aluminum wire grids were fabricated through the 2D TPP and the metal lift-off process. A second-order regression equation of the machining resolution and input parameters of the 2D TPP is deduced. The correlation coefficient between the width of the voxel and the aluminum wire grids is 0.961, which means a significant positive correlation between them. Finally, the second-order regression model derived from the fabrication resolution of the 2D TPP was validated by experiments. Full 2D grids were fabricated using 2D TPP and mental lift-off process. This paper provides a convenient, low-cost, and high-efficiency method for calibrating the fabrication resolution of 2D TPP on various photoresists.
A Derating-Sensitive Tantalum Polymer Capacitor’s Failure Rate within a DC-DC eGaN-FET-Based PoL Converter Workbench Study
Authors: Butnicu Dan
Published: January 2023

Many recent studies have revealed that PoL (Point of Load) converters’ output capacitors are a paramount component from a reliability point of view. To receive the maximum degree of reliability in many applications, designers are often advised to derate this capacitor—as such, a careful comprehending of it is required to determine the converter’s overall parameters. PoL converters are commonly found in many electronic systems. Their most important requirements are a stable output voltage with load current variation, good temperature stability, low output ripple voltage, and high efficiency and reliability. If the electronic system in question must be portable, a small footprint and volume are also important considerations—both of which have recently been well accomplished in eGaN transistor technologies. This paper provides details on how derating an output capacitor—specifically, a conductive tantalum polymer surface-mount chip, as this type of capacitor represented a step forward in miniaturization and reliability over previously existing wet electrolytic capacitors—used within a discrete eGaN-FET-based PoL buck converter determines the best performance and the highest MTBF. A setup based on an EPC eGaN FET transistor enclosed in a 9059/30 V evaluation board with a 12 V input voltage/1.2 V output voltage was tested in order to achieve the study’s main scope. Typical electrical performance and reliability data are often provided for customers by manufacturers through technical papers; this kind of public data is often selected to show the capacitors in a favorable light—still, they provide much useful information. In this paper, the capacitor derating process was presented to give a basic overview of the reliability performance characteristics of tantalum polymer capacitor when used within a DC–DC buck converter’s output filter. Performing calculations of the capacitor’s failure rate based on taking a thermal scan of the capacitor’s capsule surface temperature, the behavior of the PoL converter was evaluated.
Structural Design of a Special Machine Tool for Internal Cylindrical Ultrasonic-Assisted Electrochemical Grinding
Authors: Ma Xiaosan , Jiao Feng , Bie Wenbo , Niu Ying , Chu Shuaizhen , Hu Zhanzhan , Yang Xiaohong
Published: January 2023

During the process of internal cylindrical ultrasonic-assisted electrochemical grinding (ICUAECG), both the workpiece and the conductive grinding wheel are rotating, the machining space is closed and narrow, the electrolyte is difficult to spray into the machining area, and the insulation between the workpiece and the machine bed is challenging. According to the machining characteristics of ICUAECG, the structure of a special machine tool was designed to mitigate these problems. In particular, the rotation, electrolyte supply, electric connection, and insulation modes of the workpiece clamping parts were studied, yielding a novel workpiece clamping- and rotating-device design. This structure can fully use the internal space of the hollow spindle of the machine tool, effectively reduce the external moving parts, and achieve the appropriate liquid injection angle of the electrolyte. The ultrasonic vibration system and its installation mechanism, the dressing device of the conductive grinding wheel, and the electric grinding spindle-mounting and -fixing device were analyzed in detail. Then, a special machine tool for ICUAECG was designed, the operability and feasibility of which were verified by experiments involving conductive grinding wheel dressing and ICUAECG.
Monodisperse Micro-Droplet Generation in Microfluidic Channel with Asymmetric Cross-Sectional Shape
Authors: Cho Youngseo , Kim Jungwoo , Park Jaewon , Kim Hyun Soo , Cho Younghak
Published: January 2023

Micro-droplets are widely used in the fields of chemical and biological research, such as drug delivery, material synthesis, point-of-care diagnostics, and digital PCR. Droplet-based microfluidics has many advantages, such as small reagent consumption, fast reaction time, and independent control of each droplet. Therefore, various micro-droplet generation methods have been proposed, including T-junction breakup, capillary flow-focusing, planar flow-focusing, step emulsification, and high aspect (height-to-width) ratio confinement. In this study, we propose a microfluidic device for generating monodisperse micro-droplets, the microfluidic channel of which has an asymmetric cross-sectional shape and high hypotenuse-to-width ratio (HTWR). It was fabricated using basic MEMS processes, such as photolithography, anisotropic wet etching of Si, and polydimethylsiloxane (PDMS) molding. Due to the geometric similarity of a Si channel and a PDMS mold, both of which were created through the anisotropic etching process of a single crystal Si, the microfluidic channel with the asymmetric cross-sectional shape and high HTWR was easily realized. The effects of HTWR of channels on the size and uniformity of generated micro-droplets were investigated. The monodisperse micro-droplets were generated as the HTWR of the asymmetric channel was over 3.5. In addition, it was found that the flow direction of the oil solution (continuous phase) affected the size of micro-droplets due to the asymmetric channel structures. Two kinds of monodisperse droplets with different sizes were successfully generated for a wider range of flow rates using the asymmetric channel structure in the developed microfluidic device.
Lightweight Fan-Beam Microstrip Grid Antenna for Airborne Microwave Interferometric Radiometer Applications
Authors: Gu Chunwang , Liu Hao , Yi Min
Published: January 2023

The microwave interferometric radiometer (MIR) uses aperture synthesis technology to equate multiple small-aperture antennas into a large-aperture antenna to improve spatial resolution. At present, MIR antennas that operate at frequencies above the C-band mostly use horn antennas, waveguide slot antennas, etc., which have the disadvantages of a high profile and large mass. In this paper, a new type of miniaturized, low-profile, and lightweight K-band fan-beam microstrip grid antenna is designed for the airborne campaign of the K-band one-dimensional MIR of a Microwave Imager Combined Active and Passive (MICAP) onboard a Chinese Ocean Salinity Mission (COSM). With a limited size constraint (12.33 mm) on the antenna width, a fan-beam shape antenna pattern was achieved with a 5.34° 3-dB beamwidth in the narrow beam direction and up to a 55° 3-dB beamwidth in the fan-beam direction. A periodic structural unit is proposed in this paper to reduce the design complexity of Taylor weighting, achieving desirable performances on gain (19.1 dB) and sidelobe level (<−20 dB) in the H-plane. Four antenna elements were fabricated and arranged in a non-redundant sparse array. The performance of the four-element sparse array was evaluated with a simulation and real measurement in an anechoic chamber. The coupling between antenna elements was less than −25 dB, and the consistency of phase patterns was better than 3.4°. These results verify the feasibility of the proposed K-band microstrip grid antenna for airborne MIR applications.
Diagnosis and Stratification of COVID-19 Infections Using Differential Plasma Levels of D-Dimer: A Two-Center Study from Saudi Arabia
Authors: Alsrhani Abdullah , Alshomar Ahmad , Elderdery Abozer Y , Rasheed Zafar , Farhana Aisha
Published: January 2023

Background: D-dimer, generated upon the degradation of fibrin, is extensively used to detect thrombosis in various diseases. It is also explored as a marker for thrombosis in cases with COVID-19 disease. Few studies have confirmed its utility as a marker for assessing disease severity. Objectives: The current research was undertaken to determine the role of D-dimer in patients with COVID-19 and to investigate any association with the progression and severity of the disease in the Saudi population. Methods: Clinical indices in confirmed COVID-19 patients were collected from tertiary care hospitals in Aljouf and Qassim regions. The plasma D-dimer levels were quantified directly in the samples collected from COVID-19 patients (n = 148) using an immunofluorescence assay, and the data were presented in Fibrinogen Equivalent Units (mg/L). The collected data of D-dimer were analyzed based on COVID-19 severity, age, and the gender of patients. Results: The findings show that the plasma D-dimer concentrations were significantly (p = 0.0027) elevated in COVID-19 cases (n = 148), compared to in the normal healthy uninfected controls (n = 309). Moreover, the D-dimer levels were analyzed according to the severity of the disease in the patients. The data revealed that D-dimer concentrations were significantly increased in patients with mild infection to moderate disease, and the levels were the highest in patients with severe COVID-19 disease (p < 0.05). Our analysis demonstrates that the D-dimer levels have no association with the age or gender of COVID-19 patients (p > 0.05) in the study population. Conclusions: D-dimer can serve as a biomarker not only for the detection of COVID-19 infection, but also for determining the severity of infection of COVID-19 disease.
Response of Intestinal Microbiota of Tiger Puffer (Takifugu rubripes) to the Fish Oil Finishing Strategy
Authors: Kong Yaoyao , Liao Zhangbin , Ma Xiuhua , Liang Mengqing , Xu Houguo , Mai Kangsen , Zhang Yanjiao
Published: January 2023

The fish oil finishing (FOF) strategy, that is, re-feeding fish with fish oil (FO)-based diet after a certain period of feeding with alternative lipid source-based diets. On tiger puffer, the present study investigated the response of intestinal microbiota to FOF. Fish were fed four diets based on FO, soybean oil, palm oil and beef tallow as lipid sources, respectively, firstly for 50 days (growing-out period), and then fed the FO-based diet for 30 more days (FOF period). The results showed that dietary terrestrially sourced oils impaired the intestinal function in the growing-out period. However, the activities of amylase, trypsin and anti-oxidative enzymes (SOD, CAT, T-AOC), as well as gene expression of inflammatory cytokines (IL-1β, TNF-α, TGF-β) and tight junction protein (Claudin4, Claudin7, Claudin18, JAM, ZO-1) in the intestine were significantly recovered by FOF. The 16S rDNA sequencing analysis showed that FOF improved the similarity of bacterial community among the groups. The MetaStat analysis confirmed that FOF regulated the abundance of butyric acid-producing bacteria (Lachnospiraceae, Eubacterium, Butyricicoccus, Clostridium and Roseburia) and bacteria related to digestion and absorption (Sphingomonas, Romboutsia and Brevibacillus). In conclusion, FOF can recover the intestine function. The intestinal microbiota probably participated in and played a key role in the recovery process.
A Search for Tick-Associated, Bronnoya-like Virus Spillover into Sheep
Authors: Bratuleanu Bianca Elena , Raileanu Cristian , Chrétien Delphine , Guardado-Calvo Pablo , Bigot Thomas , Savuta Gheorghe , Temmam Sarah , Eloit Marc
Published: January 2023

Tick-borne diseases are responsible for many vector-borne diseases within Europe. Recently, novel viruses belonging to a new viral family of the order Bunyavirales were discovered in numerous tick species. In this study, we used metatranscriptomics to detect the virome, including novel viruses, associated with Ixodes ricinus collected from Romania and France. A bunyavirus-like virus related to the Bronnoya virus was identified for the first time in these regions. It presents a high level of amino-acid conservation with Bronnoya-related viruses identified in I. ricinus ticks from Norway and Croatia and with the Ixodes scapularis bunyavirus isolated from a tick cell line in Japan in 2014. Phylogenetic analyses revealed that the Bronnoya viruses’ sub-clade is distinct from several Bunyavirales families, suggesting that it could constitute a novel family within the order. To determine if Bronnoya viruses could constitute novel tick-borne arboviruses, a Luciferase immunoprecipitation assay for detecting antibodies in the viral glycoprotein of the Romanian Bronnoya virus was used to screen sera from small ruminants exposed to tick bites. No positive serum was detected, suggesting that this virus is probably not able to infect small ruminants. This study represents the first serological investigation of mammalian infections with a Bronnoya-like virus and an initial step in the identification of potential new emergences of tick-borne arboviruses.
Duration of Immunity Induced after Vaccination of Cattle with a Live Attenuated or Inactivated Lumpy Skin Disease Virus Vaccine
Authors: Haegeman Andy , De Leeuw Ilse , Mostin Laurent , Van Campe Willem , Philips Wannes , Elharrak Mehdi , De Regge Nick , De Clercq Kris
Published: January 2023

Vaccines have proven themselves as an efficient way to control and eradicate lumpy skin disease (LSD). In addition to the safety and efficacy aspects, it is important to know the duration for which the vaccines confer protective immunity, as this impacts the design of an efficient control and eradication program. We evaluated the duration of immunity induced by a live attenuated vaccine (LSDV LAV) and an inactivated vaccine (LSDV Inac), both based on LSDV. Cattle were vaccinated and challenged after 6, 12 and 18 months for LSDV LAV or after 6 and 12 months for the LSDV Inac. The LSDV LAV elicited a strong immune response and protection for up to 18 months, as no clinical signs or viremia could be observed after a viral LSDV challenge in any of the vaccinated animals. A good immune response and protection were similarly seen for the LSDV Inac after 6 months. However, two animals developed clinical signs and viremia when challenged after 12 months. In conclusion, our data support the annual booster vaccination when using the live attenuated vaccine, as recommended by the manufacturer, which could potentially even be prolonged. In contrast, a bi-annual vaccination seems necessary when using the inactivated vaccine.
Human Papillomavirus in Non-Small Cell Lung Carcinoma: Assessing Virus Presence in Tumor and Normal Tissues and Its Clinical Relevance
Authors: Tsyganov Matvey M. , Ibragimova Marina K. , Rodionov Evgeniy O. , Cheremisina Olga V. , Miller Sergei V. , Tuzikov Sergei A. , Litvyakov Nikolai V.
Published: January 2023

The significance of the role of human papillomavirus (HPV) in the development of lung cancer remains an open question. The data from the literature do not provide conclusive evidence of HPV being involved in the pathogenesis of lung cancer. The aim of this work was to detect the presence of HPV infections with a high carcinogenic risk in patients with non-small cell lung cancer (NSCLC). Materials and methods: the study involved 274 patients with stage IIA–IIIB non-small cell lung cancer. We analyzed normal and tumor tissues as well as blood from each patient. DNA was extracted from patients’ specimens, and HPV detection and genotyping was carried out using commercially available kits by PCR. Results: HPV was detected in 12.7% of the patients (35/274 of all cases). We detected nine different types of human papillomavirus in the patients, namely, types 16, 18, 31, 35, 45, 51, 52, 56, and 59. The HPV-positive samples had a clinically insignificant viral load and were predominantly integrated. The relationship between the presence of HPV and its virological parameters and the clinical and pathological parameters of the patients was established. A metastatic-free survival analysis showed that all patients with HPV in the tumor tissue had a higher 5-year survival rate (94%) compared with the HPV-negative patients (78%). The result was not statistically significant (p = 0.08). Conclusions: data showing a 12.7% human papillomavirus representation among patients with non-small cell lung cancer were obtained. The presence/absence of a viral component in patients with lung cancer was a clinically significant parameter. HPV types 16, 18, and 56, which are the most oncogenic, were most often detected.
Susceptibility and Virulence of Enterobacteriaceae Isolated from Urinary Tract Infections in Benin
Authors: Assouma Funkè F. , Sina Haziz , Adjobimey Tomabu , Noumavo Agossou Damien Pacôme , Socohou Akim , Boya Bawa , Dossou Ange D. , Akpovo Lauriane , Konmy Basile Boni Saka , Mavoungou Jacques F. , Adjanohoun Adolphe , Baba-Moussa Lamine
Published: January 2023

Enterobacteriaceae represent one of the main families of Gram-negative bacilli responsible for serious urinary tract infections (UTIs). The present study aimed to define the resistance profile and the virulence of Enterobacteriaceae strains isolated in urinary tract infections in Benin. A total of 390 urine samples were collected from patients with UTIs, and Enterobacteriaceae strains were isolated according to standard microbiology methods. The API 20E gallery was used for biochemical identification. All the isolated strains were subjected to antimicrobial susceptibility testing using the disc diffusion method. Extended-spectrum beta-lactamase (ESBL) production was investigated using a double-disc synergy test (DDST), and biofilm production was quantified using the microplate method. Multiplex PCR was used to detect uro-virulence genes, namely: PapG, IronB, Sfa, iucD, Hly, FocG, Sat, FyuA and Cnf, using commercially designed primers. More than 26% (103/390) of our samples were contaminated by Enterobacteriaceae strains at different levels. Thus, E. coli (31.07%, 32/103), Serratia marcescens (11.65%, 12/103), Klebsiella ornithinolytica (8.74%, 9/103), Serratia fonticola (7.77%, 8/103) and Enterobacter cloacae (6.80%, 7/103) were identified. Among the isolated strains, 39.81% (41/103) were biofilm-forming, while 5.83% (6/103) were ESBL-producing. Isolates were most resistant to erythromycin, cefixime, ceftriaxone and ampicillin (≥90%) followed by ciprofloxacin, gentamycin, doxycycline and levofloxacin (≥50%), and least resistant to imipenem (27.18%). In regard to virulence genes, Sfa was the most detected (28.15%), followed by IronB (22.23%), iucD (21.36%), Cnf (15.53%), PapG (9.71%), FocG (8.74%), Sat (6.79%), FyuA (5.82%) and Hyl (2.91%). These data may help improve the diagnosis of uropathogenic strains of Enterobacteriaceae, but also in designing effective strategies and measures for the prevention and management of severe, recurrent, or complicated urinary tract infections in Benin.
RyhB Paralogs Downregulate the Expressions of Multiple Survival-Associated Genes and Attenuate the Survival of Salmonella Enteritidis in the Chicken Macrophage HD11
Authors: Meng Xia , He Mengping , Chen Binjie , Xia Pengpeng , Wang Jinqiu , Zhu Chunhong , Wang Heng , Zhu Guoqiang
Published: January 2023

RyhB-1 and RyhB-2 are small non-coding RNAs in Salmonella that act as regulators of iron homeostasis by sensing the environmental iron concentration. Expressions of RyhB paralogs from Salmonella Typhimurium are increased within microphages. RyhB paralogs restrain the growth of S. Typhimurium in RAW264.7 macrophages by modulating the expression of Salmonella pathogenicity island 1 (SPI-1) genes sicA and rtsB. However, little is known about the regulatory role of RyhBs and their virulence-associated targets in Salmonella Enteritidis. We studied candidate targets of RyhB paralogs via RNA-Seq in conditions of iron limitation and hypoxia. RyhB paralogs were expressed when the S. Enteritidis strain CMCC(B)50336 (SE50336) interacted with the chicken macrophage line HD11. We analyzed gene expression associated with Salmonella survival and replication in macrophages in wild-type strain SE50336 and the RyhB deletion mutants after co-incubation with HD11 and screened out targets regulated by RyhBs. The expressions of both RyhB-1 and RyhB-2 were increased after co-incubation with HD11 for 8 h and several survival-associated genes within macrophages, such as ssaI, sseA, pagC, sodC, mgtC, yaeB, pocR, and hns, were upregulated in the ryhB-1 deletion mutant. Specifically, ssaI, the type-three secretion system 2 (T3SS-2) effector encoded by SPI-2, which promoted the survival of Salmonella in macrophages, was upregulated more than 3-fold in the ryhB-1 deletion mutant. We confirmed that both RyhB-1 and RyhB-2 downregulated the expression of ssaI to repress its mRNA translation by directly interacting with its coding sequence (CDS) region via an incomplete complementary base-pairing mechanism. The SPI-2 gene sseA was indirectly modulated by RyhB-1. The survival assays in macrophages showed that the ability of intracellular survival of ryhB-1 and/or ryhB-2 deletion mutants in HD11 was higher than that of the wild-type strain. These results indicate that RyhB paralogs downregulate survival-related virulence factors and attenuate the survival of S. Enteritidis inside chicken macrophage HD11.
Application of Cloning-Free Genome Engineering to Escherichia coli
Authors: Romeo Lucia , Esposito Antonia , Bernacchi Alberto , Colazzo Daniele , Vassallo Alberto , Zaccaroni Marco , Fani Renato , Del Duca Sara
Published: January 2023

The propagation of foreign DNA in Escherichia coli is central to molecular biology. Recent advances have dramatically expanded the ability to engineer (bacterial) cells; however, most of these techniques remain time-consuming. The aim of the present work was to explore the possibility to use the cloning-free genome editing (CFGE) approach, proposed by Döhlemann and coworkers (2016), for E. coli genetics, and to deepen the knowledge about the homologous recombination mechanism. The E. coli auxotrophic mutant strains FB182 (hisF892) and FB181 (hisI903) were transformed with the circularized wild-type E. coli (i) hisF gene and hisF gene fragments of decreasing length, and (ii) hisIE gene, respectively. His+ clones were selected based on their ability to grow in the absence of histidine, and their hisF/hisIE gene sequences were characterized. CFGE method allowed the recombination of wild-type his genes (or fragments of them) within the mutated chromosomal copy, with a different recombination frequency based on the fragment length, and the generation of clones with a variable number of in tandem his genes copies. Data obtained pave the way to further evolutionary studies concerning the homologous recombination mechanism and the fate of in tandem duplicated genes.
Characterization of Phytopythium Species Involved in the Establishment and Development of Kiwifruit Vine Decline Syndrome
Authors: Prencipe Simona , Schiavon Giada , Rosati Marco , Nari Luca , Schena Leonardo , Spadaro Davide
Published: January 2023

Since 2012, the kiwifruit vine decline syndrome (KVDS) has progressively compromised Italian kiwifruit orchards. Different abiotic and biotic factors have been associated with the establishment and development of KVDS. During monitoring of orchards affected by KVDS in north-western Italy during 2016–2019, 71 Phytopythium spp. were isolated. Based on maximum likelihood concatenated phylogeny on the ITS1-5.8S-ITS2 region of the rDNA, large subunit rDNA, and cytochrome oxidase I, isolates were identified as P. vexans (52), P. litorale (10), P. chamaehyphon (7) and P. helicoides (2). Phytopythium litorale and P. helicoides are reported for the first time as agents of KVDS in Italy. To demonstrate pathogenicity and fulfil Koch’s postulates, representative isolates of P. vexans, P. litorale, P. chamaehyphon and P. helicoides were inoculated in potted plants. In these trials, waterlogging was applied to stress plant with a temporary anoxia and to favour the production of infective zoospores by the oomycetes. In experiments in vitro, the four species showed the highest growth at 25–30 °C, depending on the media used. P. helicoides was able to grow also at 40 °C. The four species were able to grow in vitro at a pH ranging from 5.0 to 8.0, showing that pH had less effect on growth than temperature. The present study suggests a strong role of different species of Phytopythium in the establishment and development of KVDS. Phytopythium spp. could be favoured by the average increase in soil temperatures during summer, associated with global warming.
Evaluation of Serratia marcescens Adherence to Contact Lens Materials
Authors: Pifer Reed , Harris Valerie , Sanders Deaja , Crary Monica , Shannon Paul
Published: January 2023

Bacterial keratitis is a risk associated with the use of contact lenses for cosmetic purposes or vision correction. In this in vitro experimental study, we examined the ability of the ocular pathogen Serratia marcescens to adhere to monthly or biweekly replacement contact lenses. We performed quantitative adhesion assays to evaluate the adherence of S. marcescens to seven contact lens materials: comfilcon A, senofilcon A, omafilcon B, fanfilcon A, balafilcon A, senofilcon C, and lehfilcon A. Lehfilcon A is a newly marketed silicon hydrogel contact lens with a surface modification of poly-(2-methacryloyloxyethyl phosphorylcholine) (PMPC). PMPC has previously been demonstrated to be an effective anti-biofouling treatment for numerous surfaces. We observed low S. marcescens adherence to lehfilcon A compared to other materials. We demonstrate the use of the fluorescent dye 5(6)-Carboxytetramethylrhodamine succinimidyl ester to covalently stain live cells prior to material adhesion studies.
TORC1 Signaling in Fungi: From Yeasts to Filamentous Fungi
Authors: Wang Yuhua , Zheng Xi , Li Guohong , Wang Xin
Published: January 2023

Target of rapamycin complex 1 (TORC1) is an important regulator of various signaling pathways. It can control cell growth and development by integrating multiple signals from amino acids, glucose, phosphate, growth factors, pressure, oxidation, and so on. In recent years, it has been reported that TORC1 is of great significance in regulating cytotoxicity, morphology, protein synthesis and degradation, nutrient absorption, and metabolism. In this review, we mainly discuss the upstream and downstream signaling pathways of TORC1 to reveal its role in fungi.
Microbial Dynamics of a Specialty Italian Raw Ewe’s Milk Cheese Curdled with Extracts from Spontaneous and Cultivated Onopordum tauricum Willd
Authors: Rampanti Giorgia , Belleggia Luca , Cardinali Federica , Milanović Vesna , Osimani Andrea , Garofalo Cristiana , Ferrocino Ilario , Aquilanti Lucia
Published: January 2023

Milk coagulants prepared by maceration of flowers harvested from both spontaneous and cultivated Onopordum tauricum Willd. and a commercially available coagulant obtained from Cynara cardunculus L. (control) were assayed for small-scale manufacturing of Caciofiore, an Italian specialty raw ewe’s milk cheese produced in a family run dairy farm located in the Marche region (Central Italy). The microbiota of the three thistle-based milk coagulants and their effect on the microbial dynamics of raw milk cheeses during fermentation and maturation (from day 0 up until day 60) were investigated through a combined approach based on viable counting and Illumina DNA sequencing. In both the control and experimental cheeses, despite the slight differences emerged depending on the coagulant used, Lactococcus lactis and Debaryomyces hansenii were the prevalent species among bacteria and fungi, respectively. Moreover, raw ewe’s milk was the main factor affecting the evolution of both the bacterial and fungal microbiota in all cheeses. The overall similarities between control and experimental cheeses herein analyzed supports the exploitation of Onopordum tauricum Willd. as an alternative milk coagulating agent for production of Caciofiore and, more in general, raw ewe’s milk cheeses.
Biodegradation of Iprodione and Chlorpyrifos Using an Immobilized Bacterial Consortium in a Packed-Bed Bioreactor
Authors: Levío-Raimán Marcela , Bornhardt Cristian , Diez M. Cristina
Published: January 2023

This work provides the basis for implementing a continuous treatment system using a bacterial consortium for wastewater containing a pesticide mixture of iprodione (IPR) and chlorpyrifos (CHL). Two bacterial strains (Achromobacter spanius C1 and Pseudomonas rhodesiae C4) isolated from the biomixture of a biopurification system were able to efficiently remove pesticides IPR and CHL at different concentrations (10 to 100 mg L−1) from the liquid medium as individual strains and free consortium. The half-life time (T1/2) for IPR and CHL was determined for individual strains and a free bacterial consortium. However, when the free bacterial consortium was used, a lower T1/2 was obtained, especially for CHL. Based on these results, an immobilized bacterial consortium was formulated with each bacterial strain encapsulated individually in alginate beads. Then, different inoculum concentrations (5, 10, and 15% w/v) of the immobilized consortium were evaluated in batch experiments for IPR and CHL removal. The inoculum concentration of 15% w/v demonstrated the highest pesticide removal. Using this inoculum concentration, the packed-bed bioreactor with an immobilized bacterial consortium was operated in continuous mode at different flow rates (30, 60, and 90 mL h−1) at a pesticide concentration of 50 mg L−1 each. The performance in the bioreactor demonstrated that it is possible to efficiently remove a pesticide mixture of IPR and CHL in a continuous system. The metabolites 3,5-dichloroaniline (3,5-DCA) and 3,5,6-trichloro-2-pyridinol (TCP) were produced, and a slight accumulation of TCP was observed. The bioreactor was influenced by TCP accumulation but was able to recover performance quickly. Finally, after 60 days of operation, the removal efficiency was 96% for IPR and 82% for CHL. The findings of this study demonstrate that it is possible to remove IPR and CHL from pesticide-containing wastewater in a continuous system.
HIV and Drug-Resistant Subtypes
Authors: Nastri Bianca Maria , Pagliano Pasquale , Zannella Carla , Folliero Veronica , Masullo Alfonso , Rinaldi Luca , Galdiero Massimiliano , Franci Gianluigi
Published: January 2023

Acquired Immunodeficiency Syndrome (AIDS) is a human viral infectious disease caused by the positive-sense single-stranded (ss) RNA Human Immunodeficiency Virus (HIV) (Retroviridae family, Ortervirales order). HIV-1 can be distinguished into various worldwide spread groups and subtypes. HIV-2 also causes human immunodeficiency, which develops slowly and tends to be less aggressive. HIV-2 only partially homologates to HIV-1 despite the similar derivation. Antiretroviral therapy (ART) is the treatment approved to control HIV infection, based on multiple antiretroviral drugs that belong to different classes: (i) NNRTIs, (ii) NRTIs, (iii) PIs, (iv) INSTIs, and (v) entry inhibitors. These drugs, acting on different stages of the HIV life cycle, decrease the patient’s total burden of HIV, maintain the function of the immune system, and prevent opportunistic infections. The appearance of several strains resistant to these drugs, however, represents a problem today that needs to be addressed as best as we can. New outbreaks of strains show a widespread geographic distribution and a highly variable mortality rate, even affecting treated patients significantly. Therefore, novel treatment approaches should be explored. The present review discusses updated information on HIV-1– and HIV-2–resistant strains, including details on different mutations responsible for drug resistance.
Microbial-Based Products to Control Soil-Borne Pathogens: Methods to Improve Efficacy and to Assess Impacts on Microbiome
Authors: Ptaszek Magdalena , Canfora Loredana , Pugliese Massimo , Pinzari Flavia , Gilardi Giovanna , Trzciński Paweł , Malusà Eligio
Published: January 2023

Microbial-based products (either as biopesticide or biofertilizers) have a long history of application, though their use is still limited, mainly due to a perceived low and inconsistent efficacy under field conditions. However, their efficacy has always been compared to chemical products, which have a completely different mechanism of action and production process, following the chemical paradigm of agricultural production. This paradigm has also been applied to regulatory processes, particularly for biopesticides, making the marketing of microbial-based formulations difficult. Increased knowledge about bioinocula behavior after application to the soil and their impact on soil microbiome should foster better exploitation of microbial-based products in a complex environment such as the soil. Moreover, the multifunctional capacity of microbial strains with regard to plant growth promotion and protection should also be considered in this respect. Therefore, the methods utilized for these studies are key to improving the knowledge and understanding of microbial-based product activity and improving their efficacy, which, from farmers’ point of view, is the parameter to assess the usefulness of a treatment. In this review, we are thus addressing aspects related to the production and formulation process, highlighting the methods that can be used to evaluate the functioning and impact of microbial-based products on soil microbiome, as tools supporting their use and marketing.
Colletotrichum truncatum Causing Anthracnose of Tomato (Solanum lycopersicum L.) in Malaysia
Authors: Shahriar Saleh Ahmed , Husna Asmaul , Paul Terna Tersoo , Eaty Most. Nurjahan Khatun , Quamruzzaman Md , Siddique Abu Bakar , Rahim Md Abdur , Ahmmed Abu Noman Faruq , Uddain Jasim , Siddiquee Shafiquzzaman
Published: January 2023

Tomato (Solanum lycopersicum L.) is a popular nutritious vegetable crop grown in Malaysia and other parts of the world. However, fungal diseases such as anthracnose pose significant threats to tomato production by reducing the fruit quality and food value of tomato, resulting in lower market prices of the crop globally. In the present study, the etiology of tomato anthracnose was investigated in commercial tomato farms in Sabah, Malaysia. A total of 22 fungal isolates were obtained from anthracnosed tomato fruits and identified as Colletotrichum species, using morphological characteristics. The phylogenetic relationships of multiple gene sequence alignments such as internal transcribed spacer (ITS), β-tubulin (tub2), glyceraldehyde 3-phosphate dehydrogenase (gapdh), actin (act), and calmodulin (cal), were adopted to accurately identify the Colletotrichum species as C. truncatum. The results of pathogenicity tests revealed that all C. truncatum isolates caused anthracnose disease symptoms on inoculated tomato fruits. To our knowledge, the present study is the first report of tomato anthracnose caused by C. truncatum in Malaysia. The findings of this study will be helpful in disease monitoring, and the development of strategies for effective control of anthracnose on tomato fruits.
Acknowledgment to the Reviewers of Microorganisms in 2022
Authors:   Microorganisms Editorial Office
Published: January 2023

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Chemical Composition, Antibacterial and Antioxidant Activities of Essential Oil from Centipeda minima
Authors: Su Fan , Yang Gan , Hu Datong , Ruan Chen , Wang Jing , Zhang Yingying , Zhu Qingjun
Published: January 2023

This study elucidated the chemical composition of essential oil from Centipeda minima (EOCM) and its antibacterial and antioxidant activities with two chemical monomers thymol and carvacrol. The main chemical composition of EOCM, analyzed by GC-MS, were trans-chrysanthenyl acetate, thymol, aromadendrene and β-caryophyllene. In the screening of antibacterial activity against S. aureus, two monomers with antibacterial activity were obtained: thymol and carvacrol. The MIC of EOCM, thymol and carvacrol were 0.625 mg/mL, 0.156 mg/mL and 0.156 mg/mL, respectively. The experimental results were shown that three drugs could inhibit the growth of S. aureus and inhibit the formation of biofilm by changing the permeability of cell membrane and interfering with the metabolic activities in bacteria. The scavenging effects of the three drugs on DPPH radical and hydroxyl radical showed that the antioxidant effect of the three drugs was EOCM > carvacrol > thymol.
Explaining Accurate Predictions of Multitarget Compounds with Machine Learning Models Derived for Individual Targets
Authors: Lamens Alec , Bajorath Jürgen
Published: January 2023

In drug discovery, compounds with well-defined activity against multiple targets (multitarget compounds, MT-CPDs) provide the basis for polypharmacology and are thus of high interest. Typically, MT-CPDs for polypharmacology have been discovered serendipitously. Therefore, over the past decade, computational approaches have also been adapted for the design of MT-CPDs or their identification via computational screening. Such approaches continue to be under development and are far from being routine. Recently, different machine learning (ML) models have been derived to distinguish between MT-CPDs and corresponding compounds with activity against the individual targets (single-target compounds, ST-CPDs). When evaluating alternative models for predicting MT-CPDs, we discovered that MT-CPDs could also be accurately predicted with models derived for corresponding ST-CPDs; this was an unexpected finding that we further investigated using explainable ML. The analysis revealed that accurate predictions of ST-CPDs were determined by subsets of structural features of MT-CPDs required for their prediction. These findings provided a chemically intuitive rationale for the successful prediction of MT-CPDs using different ML models and uncovered general-feature subset relationships between MT- and ST-CPDs with activities against different targets.
Interaction of Chondroitin and Hyaluronan Glycosaminoglycans with Surfaces of Carboxylated Carbon Nanotubes Studied Using Molecular Dynamics Simulations
Authors: Panczyk Tomasz , Plazinski Wojciech , Dupradeau François-Yves , Brzyska Agnieszka , Wolski Pawel
Published: January 2023

Interaction of β-D-glucopyranuronic acid (GlcA), N-acetyl-β-D-glucosamine (GlcNAc), N-acetyl-β-D-galactosamine (GalNAc) and two natural decameric glycosaminoglycans, hyaluronic acid (HA) and Chondroitin (Ch) with carboxylated carbon nanotubes, were studied using molecular dynamics simulations in a condensed phase. The force field used for carbohydrates was the GLYCAM-06j version, while functionalized carbon nanotubes (fCNT) were described using version two of the general amber force field. We found a series of significant differences in carbohydrate-fCNT adsorption strength depending on the monosaccharide molecule and protonation state of surface carboxyl groups. GlcNAc and GalNAc reveal a strong adsorption on fCNT with deprotonated carboxyl groups, and a slightly weaker adsorption on the fCNT with protonated carboxyl groups. On the contrary, GlcA weakly adsorbs on fCNT. The change in protonation state of surface carboxyl groups leads to the reversal orientation of GlcNAc and GalNAc in reference to the fCNT surface, while GlcA is not sensitive to that factor. Adsorption of decameric oligomers on the surface of fCNT weakens with the increasing number of monosaccharide units. Chondroitin adsorbs weaker than hyaluronic acid and incorporation of four Ch molecules leads to partial detachment of them from the fCNT surface. The glycan–fCNT interactions are strong enough to alter the conformation of carbohydrate backbone; the corresponding conformational changes act toward a more intensive contact of glycan with the fCNT surface. Structural and energetic features of the adsorption process suggest the CH-π interaction-driven mechanism.