Fast Thermocycling in Custom Microfluidic Cartridge for Rapid Single-Molecule Droplet PCR.

The microfluidic droplet polymerase chain reaction (PCR), which enables simultaneous DNA amplification in numerous droplets, has led to the discovery of various applications that were previously deemed unattainable. Decades ago, it was demonstrated that the temperature holding periods at the denaturation and annealing stages in thermal cycles for PCR amplification could be essentially eliminated if a rapid change of temperature for an entire PCR mixture was achieved. Microfluidic devices facilitating the application of such fast thermocycling protocols have significantly reduced the time required for PCR. However, in microfluidic droplet PCR, ensuring successful amplification from single molecules within droplets has limited studies on accelerating assays through fast thermocycling. Our developed microfluidic cartridge, distinguished for its convenience in executing single-molecule droplet PCR with common laboratory equipment, features droplets positioned on a thin glass slide. We hypothesized that applying fast thermocycling to this cartridge would achieve single-molecule droplet PCR amplification. Indeed, the application of this fast protocol demonstrated successful amplification in just 22 min for 30 cycles (40 s/cycle). This breakthrough is noteworthy for its potential to expedite microfluidic droplet PCR assays, ensuring efficient single-molecule amplification within a remarkably short timeframe.

Revolutionizing Patient Monitoring in Age-Related Macular Degeneration: A Comparative Study on the Necessity and Efficiency of the AMD VIEWER.

(1) Background: Age-related Macular Degeneration (AMD) is a critical condition leading to blindness, necessitating lifelong clinic visits for management, albeit with existing challenges in monitoring its long-term progression. This study introduced and assessed an innovative tool, the AMD long-term Information Viewer (AMD VIEWER), designed to offer a comprehensive display of crucial medical data-including visual acuity, central retinal thickness, macular volume, vitreous injection treatment history, and Optical Coherent Tomography (OCT) images-across an individual eye's entire treatment course. (2) Methods: By analyzing visit frequencies of patients with a history of invasive AMD treatment, a comparative examination between a Dropout group and an Active group underscored the clinical importance of regular visits, particularly highlighting better treatment outcomes and maintained visual acuity in the Active group. (3) Results: The efficiency of AMD VIEWER was proven by comparing it to manual data input by optometrists, showing significantly faster data display with no errors, unlike the time-consuming and error-prone manual entries. Furthermore, an elicited Net Promoter Score (NPS) of 70 from 10 ophthalmologists strongly endorsed AMD VIEWER's practical utility. (4) Conclusions: This study underscores the importance of regular clinic visits for AMD patients. It suggests the AMD VIEWER as an effective tool for improving treatment data management and display.

Branch retinal artery occlusion with unruptured retinal arterial macroaneurysm post-SARS-CoV-2 vaccination: A case report.

INTRODUCTION: Many adverse occurrences in the eye have been reported after severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) vaccination. This is the first report of a patient with an unruptured retinal arterial macroaneurysm (RAM) who developed branch retinal artery occlusion (BRAO) one day after SARS-CoV-2 vaccination (BNT162b2 by Pfizer-BioNTech). PATIENT REPORT: A 75-year-old man with a pertinent history of type-2 diabetes mellitus visited the hospital complaining of sudden visual loss in his right eye 1 day after receiving the fourth dose of the SARS-CoV-2 mRNA vaccine; his best-corrected visual acuity (BCVA) decreased from 1.0 to 0.7 (Snellen decimal). The patient had previously been diagnosed with an unruptured RAM and superior paracentral acute middle maculopathy in the same eye. Fundus examination showed increased sheathing of blood vessels. Indocyanine green showed a hyperfluorescent area suggestive of RAM on the right eye disc. Fluorescein angiography and optical coherence tomography angiography revealed arterial obstruction findings in the upper retinal area. DIAGNOSIS AND INTERVENTION: The patient was diagnosed with BRAO with RAM and was followed up without any additional treatment. Follow-up examination after 4 months did not show any improvement in BCVA value. CONCLUSION: This case suggested that BRAO could develop after SARS-CoV-2 vaccination in patients with unruptured RAM; however, more research is required to investigate the causes.

Patterning Perovskite Quantum Dots Using Photopolymerization.

All-inorganic cesium lead halide perovskite quantum dots (QDs) have several potential applications, owing to their unique optical and electronic properties. However, patterning perovskite QDs using conventional methods is difficult because of the ionic nature of QDs. Here, we demonstrate a unique approach, in which perovskite QDs are patterned in polymer films through the photocuring of monomers under patterned light illumination. The pattern illumination creates the transient polymer concentration difference, which drives the QDs to form patterns; hence controlling polymerization kinetics is essential for the generation of the QD pattern. For the patterning mechanism, a light projection system equipped with a digital micromirror device (DMD) is developed; thus, light intensity, an important factor to determine polymerization kinetics, is precisely controlled per position on the photocurable solution, resulting in the understanding of the mechanism and the formation of distinct QD patterns. The demonstrated approach assisted by the DMD-equipped projection system can form desired perovskite QD patterns solely by patterned light illumination, paving the way for the development of patterning methods for perovskite QDs and other nanocrystals.

Effects of Antihistamine-Releasing Contact Lenses on Severe Allergic Conjunctivitis.

Half of the patients who used CL daily did not wish to switch to spectacles even with ocular-allergic symptoms and findings. Therefore, continued CL use with suppressed ocular allergy can improve productivity and quality of life for CL users. This study described the clinical courses of CL users with ocular-allergic conjunctivitis who started ARCL trials on their eyes. Diagnoses of the seven understudied cases were AKC in 3, VKC in 2, SAC in 1, and CLPC in 1 case. All seven cases had myopia. Following the use of ARCL, six patients except for case 7 were satisfied, and objective findings were judged to be well-controlled. ARCL can be a useful tool for CL users with allergic conjunctivitis. However, ARCL should be introduced after allergic conjunctivitis is controlled or becomes asymptomatic. Furthermore, ARCL should be immediately discontinued if ocular-allergic symptoms flare after introducing ARCL.

Auxin-Induced WUSCHEL-RELATED HOMEOBOX13 Mediates Asymmetric Activity of Callus Formation upon Cutting.

Plants have the regenerative ability to reconnect cut organs, which is physiologically important to survive severe tissue damage. The ability to reconnect organs is utilized as grafting to combine two different individuals. Callus formation at the graft junction facilitates organ attachment and vascular reconnection. While it is well documented that local wounding signals provoke callus formation, how callus formation is differentially regulated at each cut end remains elusive. Here, we report that callus formation activity is asymmetrical between the top and bottom cut ends and is regulated by differential auxin accumulation. Gene expression analyses revealed that cellular auxin response is preferentially upregulated in the top part of the graft. Disruption of polar auxin transport inhibited callus formation from the top, while external application of auxin was sufficient to induce callus formation from the bottom, suggesting that asymmetric auxin accumulation is responsible for active callus formation from the top end. We further found that the expression of a key regulator of callus formation, WUSCHEL-RELATED HOMEOBOX 13 (WOX13), is induced by auxin. The ectopic callus formation from the bottom end, which is triggered by locally supplemented auxin, requires WOX13 function, demonstrating that WOX13 plays a pivotal role in auxin-dependent callus formation. The asymmetric WOX13 expression is observed both in grafted petioles and incised inflorescence stems, underscoring the generality of our findings. We propose that efficient organ reconnection is achieved by a combination of local wounding stimuli and disrupted long-distance signaling.

Branch retinal vein occlusion post severe acute respiratory syndrome coronavirus 2 vaccination.

In this article, we report two patients who experienced the first onset of branch retinal vein occlusion (BRVO) 3 days after the administration of the BNT162b2 (Pfizer-BioNTech) severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) vaccine. Case 1: A 50-year-old woman without any history of retinal disease developed vision loss in her right eye 3 days after receiving the first dose of the SARS-CoV-2 mRNA vaccine. Case 2: A 56-year-old woman without any history of retinal disease developed vision loss in her right eye 3 days after receiving the first dose of the SARS-CoV-2 mRNA vaccine. Case 1: Temporal superior BRVO and secondary macular edema (ME) were observed in the patient's right eye. Her best-corrected visual acuity (BCVA) was 20/25. Case 2: Temporal inferior BRVO and secondary ME were observed in the patient's right eye. Her BCVA was 13/20. Case 1: Three doses of intravitreal ranibizumab (IVR) were administered. Case 2: Three doses of IVR were administered. Case 1: ME resolved and BCVA improved to 20/20. Case 2: ME resolved and BCVA improved to 20/20. Both the cases showed a possible association between the SARS-CoV-2 vaccination and the first onset of BRVO.

Wound-inducible WUSCHEL-RELATED HOMEOBOX 13 is required for callus growth and organ reconnection.

Highly efficient tissue repair is pivotal for surviving damage-associated stress. Plants generate callus upon injury to heal wound sites, yet regulatory mechanisms of tissue repair remain elusive. Here, we identified WUSCHEL-RELATED HOMEOBOX 13 (WOX13) as a key regulator of callus formation and organ adhesion in Arabidopsis (Arabidopsis thaliana). WOX13 belongs to an ancient subclade of the WOX family, and a previous study shows that WOX13 orthologs in the moss Physcomitrium patens (PpWOX13L) are involved in cellular reprogramming at wound sites. We found that the Arabidopsis wox13 mutant is totally defective in establishing organ reconnection upon grafting, suggesting that WOX13 is crucial for tissue repair in seed plants. WOX13 expression rapidly induced upon wounding, which was partly dependent on the activity of an AP2/ERF transcription factor, WOUND-INDUCED DEDIFFERENTIATION 1 (WIND1). WOX13 in turn directly upregulated WIND2 and WIND3 to further promote cellular reprogramming and organ regeneration. We also found that WOX13 orchestrates the transcriptional induction of cell wall-modifying enzyme genes, such as GLYCOSYL HYDROLASE 9Bs, PECTATE LYASE LIKEs and EXPANSINs. Furthermore, the chemical composition of cell wall monosaccharides was markedly different in the wox13 mutant. These data together suggest that WOX13 modifies cell wall properties, which may facilitate efficient callus formation and organ reconnection. Furthermore, we found that PpWOX13L complements the Arabidopsis wox13 mutant, suggesting that the molecular function of WOX13 is partly conserved between mosses and seed plants. This study provides key insights into the conservation and functional diversification of the WOX gene family during land plant evolution.

Patterning Silver Nanowires by Inducing Transient Concentration Gradients in Reaction Mixtures.

Patterning nanocrystals in polymer films is essential for the widespread use of nanocrystals in various fields from optics to electronics; therefore, the development of patterning methods for nanocrystals is an important task. Here, we report a unique approach for patterning silver nanowires (AgNWs) using a thermodynamic driving force induced by transient concentration gradients in reaction mixtures. The procedure starts with the preparation of a photocurable monomer solution containing homogeneously dispersed AgNWs. Ultraviolet illumination through a straight-line mask reduces the polymerization rate of monomers in the masked area, decreasing the polymer concentration in comparison with that in the unmasked area. Such transient polymer concentration gradients yield imbalances in the chemical potentials of AgNWs, inducing the migration of AgNWs to form a straight-line pattern of AgNWs. The pattern of AgNWs was visualized via photoluminescence imaging under a laser scanning confocal microscope and compared with the light patterns applied to the mixture. These observations revealed that the magnitude of the AgNW migration is enhanced as the transient concentration gradient increases by thickening the mask to decrease the intensity of light passing through the mask. The structural features of the AgNW pattern were reproduced using numerical simulations based on a set of reaction-diffusion equations, which suggested the key role of the polymerization kinetics characterized by the Trommsdorff-Norrish effect. Moreover, as the AgNW pattern becomes clearer, the electrical resistance along the patterns decreases and more complex patterns can be produced, indicating the potential of the method. Overall, the present patterning method constitutes a simple approach that only requires illumination through a mask to generate the AgNW pattern, which renders it a promising alternative for patterning nanocrystals in polymer films.

Exacerbation of branch retinal vein occlusion post SARS-CoV2 vaccination: Case reports.

RATIONALE: In this paper, we report on 2 patients who developed branch retinal vein occlusion (BRVO) exacerbation 1 day after administration of the BNT162b2 (Pfizer-BioNTech) SARS-CoV-2 vaccine. PATIENT CONCERNS: Case 1: A 71 year-old female developed vision loss in her left eye 1 day after receiving a second dose of the SARS-CoV-2 mRNA vaccine. This patient was diagnosed with temporal inferior BRVO and secondary macular edema (ME) in her left eye. ME resolved after 3 doses of intravitreal aflibercept (IVA). After treatment, no recurrence of ME was observed.Case 2: A 72 year-old man developed vision loss in his right eye 1 day after receiving the first dose of the SARS-CoV-2 mRNA vaccine. This patient was diagnosed with temporal superior BRVO in the right eye without ME. The patient was followed up and did not undergo any additional treatment. DIAGNOSES: Case1: Temporal superior BRVO and secondary ME were observed in the left eye. Her best-corrected visual acuity (BCVA) was 20/30.Case2: Temporal superior BRVO recurrence and secondary ME were observed in the right eye. BCVA was 20/25. INTERVENTIONS: Case1: Additional dose of IVA was administered. Case2: Two times of Intravitreal ranibizumab was administered twice. OUTCOMES: Case1: Subsequently, ME resolved BCVA was 20/20. Case2: Subsequently, ME resolved BCVA was 20/25. LESSONS: Both cases showed a possible association between SARS-CoV-2 vaccination and the exacerbation of BRVO.

Heterocypris spadix sp. nov. (Crustacea: Ostracoda: Cypridoidea) from Japan, with Information on Its Reproductive Mode.

We describe the cypridoidean ostracod Heterocypris spadix sp. nov. from brackish water on Okinawa Island, Japan. The species closely resembles Heterocypris salina (Brady, 1868) but differs in that (1) the marginal infolds on valves are less developed, (2) the tubercles on the anterior margin of the right valve are completely covered by the selvage and invisible in inner view, and (3) the calcified inner lamella on the ventral margin of the left and right valves is scarcely evident in inner view, as the ventral margins of the valves bend inwardly. We determined partial sequences for the cytochrome c oxidase subunit I (COI; cox1) and 18S rRNA genes in H. spadix for future DNA barcoding and phylogenetic analyses. Our sample contained only females. A breeding experiment revealed that H. spadix females reproduce parthenogenetically. Another experiment showed that H. spadix has low tolerance to desiccation, with all individuals at 25°C dying between 1-2 hours after removal from water. We amplified and sequenced a partial 16S rRNA sequence for the endosymbiotic bacterium Cardinium from H. spadix. Infection by Cardinium may be related to the parthenogenetic reproductive mode we observed in H. spadix.

A New Species of Ostracod (Crustacea) Associated with a Feather Star: First Report of Ostracoda from Crinoidea.

We describe Obesostoma crinophilum sp. nov. (Ostracoda: Podocopida: Paradoxostomatidae) obtained from the body surface of the feather star Antedon serrata A. H. Clark, 1908 (Crinoidea: Comatulida: Antedonidae). This is the first report of Ostracoda associated with Crinoidea. None of the highly specialized appendages and/or carapace that are related to a commensal lifestyle were observed in O. crinophilum sp. nov. Therefore, the relationship between O. crinophilum sp. nov. and A. serrata must be transient rather than obligatory. However, O. crinophilum sp. nov. has a more developed hook-like distal claw on the antenna in comparison with four previously known Obesostoma species. The relatively well-developed distal claw of the antenna in O. crinophilum sp. nov. should indicate its intimate association with feather stars, though the feeding habit is still unknown.

Pontopolycope orientalis sp. nov. (Crustacea: Ostracoda: Polycopidae), the First Report of a Living Species of the Genus from the Indo-Pacific Region.

The present study describes Pontopolycope orientalis sp. nov. (Polycopidae), which was collected from the shallow sandy bottom off of Nagannu Island, Okinawa Prefecture, southwestern Japan. This new species is the first living representative of Pontopolycope from the Indo-Pacific region. We also provided a key to identifying five living congeners throughout the world's oceans. Since Pontopolycope is defined by the morphological characteristics of the carapace, the following five fossil and subfossil (empty valve) species of Polycope (Polycopidae) are moved to Pontopolycope: Pontopolycope luxuriosa (Herrig, 1964) comb. nov., P. sanctacatherinae (Whatley and Downing, 1983) comb. nov., P. krauseae (Herrig, 1994) comb. nov., P. proboscidea (Herrig, 1994) comb. nov., and P. isolata (Whatley, Jones and Wouters, 2000) comb. nov. Finally, the nucleotide sequence data of three genes (18S rRNA, 28S rRNA, and mitochondrial cytochrome c oxidase subunit 1) of the new species are provided for future systematic and phylogenetic studies.

Concealment of trauma and occupational accidents among Fukushima nuclear disaster decontamination workers: A case report.

OBJECTIVES: Limited information exists concerning occupational risks in decontamination work after the Fukushima Daiichi Nuclear Power Plant (FDNPP) accident. Workers involved tend to be migrant workers, face various health risks, and are usually from a low socioeconomic background and generally have difficulty in finding employment. We report a specific case to illustrate the way these workers tend to get injured during working hours and draw attention to the problems arising. CASE PRESENTATION: A 59-year-old Japanese male decontamination worker was referred to our emergency department after a fall while he was working in an Exclusion Zone surrounding the FDNPP. He was blind in his right eye. He was diagnosed with traumatic multiple rib fractures and a tube thoracostomy was performed. He was discharged from hospital after 7 days. Payment has been changed from "occupational accident," which is required to be reported to the Local Labor Standards Office, to "general medical treatment" which is no obligation. CONCLUSION: Trauma or physical injury of any kind is an occupational hazard for workers, especially those operating in the chaotic and unpredictable environments following any disasters. Companies employing such workers and owners of any facilities or locations in which they may be working are responsible for the safety of their workers. They should provide appropriate training and should comply with all prevailing Employment Laws and follow mandatory safety regulations. If companies and authorities are in breach of any laws, ignore their responsibilities, or jeopardize the health of their workers, they should be held accountable.

Molecular and Morphological Diversity of Heterodesmus Brady and Its Phylogenetic Position within Cypridinidae (Ostracoda).

Ostracod genus Heterodesmus Brady, 1866 is known thus far to contain only three species: H. adamsii Brady, 1866; H. apriculus Hiruta, 1992; and H. naviformis (Poulsen, 1962). This genus has been recorded from the Sea of Japan, and the coastal areas of Thailand and Vietnam. The main generic character is the presence of antero-dorsal and postero-dorsal tube-like processes on the rostrum on both valves. The three species mostly differ in the shell lateral projections. Their relationship and the position of Heterodesmus within family Cypridinidae are poorly understood, partly due to the lack of publication of DNA data so far. We study Heterodesmus collected from several localities in the Northwest Pacific, namely Tsushima and Iki Islands in Japan and Maemul Island in Korea. Besides morphological characters, we also use two mitochondrial markers (16S rRNA and mtCOI) and three nuclear regions (18S rRNA, 28S rRNA, and internal transcribed spacer - ITS) in the samples to detect the biodiversity of this genus. Our phylogenetic tree based on molecular data coupled with morphology reveals the presence of two species, H. adamsii and H. apriculus. We report on their morphological variability, molecular diversity, and phylogenetic position within Cypridinidae based on 16S, 28S and 18S rRNAs, and provide a taxonomic key for all living genera of this family. For the first time, we give an overview of the intrageneric and intrafamily DNA distances of the above markers for the entire subclass Myodocopa.

Nanocrystals Assembled by the Chemical Reaction of the Dispersion Solvent.

The development of methods to pattern nanocrystals with different sizes and shapes remains a challenge. In this study, we demonstrate a unique class of bottom-up approaches to assemble nanocrystals into patterns. Our approach for patterning nanocrystals focuses on the utilization and control of the chemical reaction of solvents surrounding nanocrystals. The photopolymerization of solvent molecules through a photomask creates time-dependent concentration gradients of the solvents. Dispersed nanocrystals such as silver nanowires (AgNWs) migrate and are gradually organized and integrated into the polymerizing films based on the concentration gradients. The AgNW-embedded film properties are determined by the organized AgNW structures and include light transmission and electrical conductivity. Overall, the demonstrated method is very simple, widely applicable to various nanocrystals and solvents, and can thus contribute to the development of a new class of nanocrystal patterning methods.

Predominance of methicillin-resistant Staphylococcus aureus SCCmec type II-CC5 and SCCmec type IV-CC1/CC8 among companion animal clinical isolates in Japan: Findings from phylogenetic comparison with human clinical isolates.

OBJECTIVES: To characterise the genotypic profiles of methicillin-resistant Staphylococcus aureus (MRSA) clinical isolates from companion animals and to investigate their association with those from humans in Japan. METHODS: Non-duplicated MRSA clinical isolates recovered between July 2016 and January 2018 were analysed. The MRSA isolates were typed by polymerase chain reaction (PCR)-based open reading frame (ORF) typing (POT) scores, SCCmec types, multilocus sequence typing, and virulence gene profiles. Phylogenetic comparison of those isolates with previously described human isolates was performed. RESULTS: Among 56 MRSA isolates (33 cats, 20 dogs and three rabbits), 26 isolates with a POT1 score of 93, SCCmec type II mostly belonged to CC5, including ST5. Twenty-six isolates with a POT1 score of 106, SCCmec type IV showed diversity of STs: 15 isolates belonged to CC8, mainly including ST8, and 11 isolates belonged to CC1, including ST1 and newly identified STs 4768, 4775, and 4779. Two cat isolates were ST8-SCCmec type IV possessing pvl/ACME-arcA, presumed to be the hypervirulent community-associated MRSA (CA-MRSA) clone USA300. Notably, all three rabbit isolates belonged to ST4768. The POT1 score 106 CA-MRSA isolates from animals and humans were divided into two large clusters of CC1 and CC8, where host species-specific sub-clusters were not identified within each cluster. A large cluster of POT1 score 93 healthcare-associated MRSA (HA-MRSA) isolates from animals and humans consisted of sub-clusters formed exclusively by the vast majority of human isolates and those formed by animal and human isolates. CONCLUSION: Companion animals could be potential reservoirs and vehicles for the transmission of CA-MRSA to humans, and could transmit companion animal-adaptive HA-MRSA lineages to humans as their second reservoirs.

Detection of Acinetobacter pittii ST220 co-producing NDM-1 and OXA-820 carbapenemases from a hospital sink in a non-endemic country of NDM.

OBJECTIVES: NDM-1 is by far one of the most commonly prevalent carbapenemases in Enterobacteriaceae and Acinetobacter baumannii. This study presented an Acinetobacter pittii (A. pittii) isolate co-harboring blaNDM-1 and blaOXA-820 from a university hospital sink, where New Delhi metallo-ß-lactamase (NDM) producers have not been found in either patients or their environments. METHODS: Whole-genome sequencing was performed on the HiSeq 4000 platform, and the reads were de novo assembled using the A5-miSeq Assembly pipeline. Annotation of the resulting scaffolds were performed by using the DDBJ Fast Annotation and Submission Tool (DFAST). The blaNDM-1-carrying plasmid was determined. RESULTS: The A. pittii ST220 strain SU1805 detected from a sink strainer in the treatment room was resistant to imipenem and meropenem. Antimicrobial resistance genes blaNDM-1, blaOXA-820, blaADC-43, and aphA6 were found in this strain. The blaNDM-1 was found to be located downstream of an ISAba125 element on a plasmid pSU1805NDM with a size of 41,022 bp, and GC content of 38.3% harbouring 48 protein-coding genes. The aphA6 gene was also located upstream of the ISAba125 on the same plasmid. The A. pittii intrinsic blaOXA-213-like gene blaOXA-820 was located between fxsA and yncA genes in the chromosome. The strain also harboured biofilm-associated genes such as ompA, the csu operon and their regulating genes bfmRS. CONCLUSION: This study described the first isolation of NDM-1-producing A. pittii in Japan, and highlighted the importance of proper implementation of measures against AMR for sink drainage systems, since NDM producers may have already been hidden in such environments in a non-endemic country of NDM.