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Abstract Porcine deltacoronavirus (PDCoV), also known as HKU15, is a swine enteropathogenic virus that is believed to have originated in birds. PDCoV belongs to the genus Deltacoronavirus (DCoV), the members of which have mostly been identified in diverse avian species. We recently reported that chicken or porcine aminopeptidase N (APN), the major cellular receptor for PDCoV, can mediate cellular entry via three pseudotyped retroviruses displaying spike proteins from three avian DCoVs (HKU11, HKU13, and HKU17). In the present work, to better understand how avian-origin CoVs may be transmitted to pigs, we investigated the unknown DCoV entry pathway in avian cells. We show that clathrin-mediated endocytosis is involved in the entry of these DCoV pseudoviruses into chicken-origin DF-1 cells. Pseudovirus entry was suppressed by means of pharmacological inhibitors, dominant-negative mutants, and siRNAs targeting various cellular proteins and signalling molecules, suggesting that PDCoV and avian DCoV pseudovirus entry into DF-1 cells depends on clathrin, dynamin-2, cathepsins and a low-pH environment but is independent of caveolae and macropinocytosis. Furthermore, we found that DCoV pseudovirus entry was linked to Rab5- and Rab7-dependent pathways. This is the first report demonstrating that these DCoVs utilize clathrin-mediated endocytosis pathways to enter avian-origin cells, providing new insights into interspecies transmission of DCoVs.

Herein, a lucid meta-holographic approach leveraging geometric phase metasurfaces and photonic crystal surface-emitting lasers is presented. Such a straightforward technique inherently enables polarized structured light projection with opposite circular polarization states, effectively doubling the field of view for depth sensing applications. Furthermore, the innate polarization state of projected light provides enhanced resistance to environmental interference. To demonstrate the practical utility of this method, an integrated depth sensing system based on the geometric phase meta-hologram has been developed and successfully detects targets at an operating distance of 75 cm, paving the way for advanced 3D sensing applications in various fields.

Abstract Free-space optical communications hold promising advantages, including a large bandwidth, access to license-free spectrum, high data rates, quick and simple deployment, low power consumption, and relaxed quality requirements. Nevertheless, key technical challenges remain, such as a higher transmission efficiency, a lower transmission loss, and a smaller form factor of optical systems. Here, we demonstrate the viability of circular-polarization-multiplexed multi-channel optical communication using metasurfaces alongside a photonic-crystal surface-emitting laser (PCSEL) light source at wavelength of 940 nm. Through the light manipulation with metasurface, we split the linearly polarized incidence into left and right circular polarizations with desired diffraction angles. Such orthogonal polarization states provide a paradigm of polarization division multiplexing technique for light communication. The PCSEL light source maintains a low divergence angle of about 0.373 degrees after passing through an ultra-thin metasurface without further bulky collimator or light guide, making end-to-end (E2E) and device-to-device (D2D) communications available in a compact form. Both light source and modulated polarized light exhibit a − 3 dB bandwidth over 500 MHz, with successful 1 Gbit/s transmission demonstrated in eye diagrams. Our results affirm that metasurface effectively boosts transmission capacity without compromising the light source's inherent properties. Future metasurface designs could expand channel capacity, and its integration with PCSEL monolithically holds promise for reducing interface losses, thereby enhancing efficiency.

In recent years, an increasing number of viruses have triggered outbreaks that pose a severe threat to both human and animal life, as well as caused substantial economic losses. It is crucial to understand the genomic structure and epidemiology of these viruses to guide effective clinical prevention and treatment strategies. Nanopore sequencing, a third-generation sequencing technology, has been widely used in genomic research since 2014. This technology offers several advantages over traditional methods and next-generation sequencing (NGS), such as the ability to generate ultra-long reads, high efficiency, real-time monitoring and analysis, portability, and the ability to directly sequence RNA or DNA molecules. As a result, it exhibits excellent applicability and flexibility in virus research, including viral detection and surveillance, genome assembly, the discovery of new variants and novel viruses, and the identification of chemical modifications. In this paper, we provide a comprehensive review of the development, principles, advantages, and applications of nanopore sequencing technology in animal and human virus research, aiming to offer fresh perspectives for future studies in this field.

Compact photonic devices that integrate metasurfaces with light sources have been widely studied. However, experimental demonstrations of a higher efficiency of integration are still lacking. To enhance the efficiency of light sources integrated with metasurfaces, we employed a forward design optimization method and index matching between the light source and metasurface substrate to design metagratings. To optimize the overall diffraction efficiency, we manipulated the degrees of freedom in phase, the lattice constants, and the number of unit cells. The same material was utilized for the nanostructures and substrate (homo-metagrating) for index matching, while Si and GaAs materials were used for working at 1550 and 940 nm, respectively. The experimental homo-metagratings operating at 1550 nm and made of Si exhibited an overall average efficiency of 51.3% at diffraction angles of 60.3°. On the other hand, experimental homo-metagratings operating at 940 nm and made of GaAs exhibited an overall average efficiency of 52.4% at diffraction angles of 49.3°. This suggests that the future integration of metagratings with a polarization-specific laser can further enhance the overall diffraction efficiency.

The global health crisis caused by the COVID-19 pandemic has led to a surge in demand and use of personal protective equipment (PPE) such as masks, putting great pressure on social production and the environment. It is urgent to find an efficient and non-destructive disinfection method for the safe reuse of PPE. This study proposes a PPE disinfection method that uses erythrosine, a U.S. Food and Drug Administration-approved food dye, as photosensitizer to produce singlet oxygen for virus inactivation, and indicates the completion of disinfection by its photobleaching color change. After spraying 100 μL of 10 μM erythrosine on the surface of the mask for 3 times and light exposure for 25 min, the titer of coronavirus decreased by more than 99.999%, and the color of erythrosine on the mask surface disappeared. In addition, the structure of the mask was intact and the filtration efficiency was maintained at > 95% after 10 cycles of erythrosine treatment. Therefore, this disinfection method can provide at least 10 cycles of reuse with the advantages of high safety and convenient, and the completion of disinfection can be indicated by its photobleaching, which is suitable for hospitals and daily life to reduce the consumption of PPE.

Abstract Meta-optics has achieved major breakthroughs in the past decade; however, conventional forward design faces challenges as functionality complexity and device size scale up. Inverse design aims at optimizing meta-optics design but has been currently limited by expensive brute-force numerical solvers to small devices, which are also difficult to realize experimentally. Here, we present a general inverse-design framework for aperiodic large-scale (20k × 20k λ 2) complex meta-optics in three dimensions, which alleviates computational cost for both simulation and optimization via a fast approximate solver and an adjoint method, respectively. Our framework naturally accounts for fabrication constraints via a surrogate model. In experiments, we demonstrate aberration-corrected metalenses working in the visible with high numerical aperture, poly-chromatic focusing, and large diameter up to the centimeter scale. Such large-scale meta-optics opens a new paradigm for applications, and we demonstrate its potential for future virtual-reality platforms by using a meta-eyepiece and a laser back-illuminated micro-Liquid Crystal Display.

Abstract Internet of Things (IoT) technology is prosperous for the betterment of human well-being. With the expeditious needs of miniature functional devices and systems for adaptive optics and light manipulation at will, relevant sensing techniques are thus in the urgent stage of development. Extensive developments in ultrathin artificial structures, namely metasurfaces, are paving the way for the next-generation devices. A bunch of tunable and reconfigurable metasurfaces with diversified catalogs of mechanisms have been developed recently, enabling dynamic light modulation on demand. On the other hand, monolithic integration of metasurfaces and light-emitting sources form ultracompact meta-devices as well as exhibiting desired functionalities. Photon-matter interaction provides revolution in more compact meta-devices, manipulating light directly at the source. This study presents an outlook on this merging paradigm for ultracompact nanophotonics with metasurfaces, also known as metaphotonics. Recent advances in the field hold great promise for the novel photonic devices with light emission and manipulation in simplicity.

Porcine reproductive and respiratory syndrome virus (PRRSV) has caused huge economic losses for the global pig industry, but its origins and evolution remain a mystery. In 2018, the genome sequences of seven arteriviruses isolated from rodents were determined, and here we publish new analysis showing that they may be ancestors of PRRSV. The sequence similarity of these viruses to PRRSV was ~60%, with shared genome organization and other characteristics, such as slippery sequences and C-rich motifs in nsp2, and a transactivated protein sequence in nsp1β. Codon usage basis analysis showed that PRRSV was closer to these rodent arteriviruses than lactate dehydrogenase-elevating virus (LDV) and they were both under pressure of natural selection. Evolutionary analysis revealed that four of the rodent arteriviruses shared the same genus with PRRSV, and were more closely related to PRRSV-2 than PRRSV-1. In addition to this, they all appeared earlier than PRRSV according to evolutionary modeling, and we speculate that they represent an intermediate step in the origin of PRRSV by arterivirus transmission from rodents to swine. Our in-depth analysis furthers our understanding of arteriviruses, and will serve as the basis for subsequent exploration of the evolution of PRRSV and other arteriviruses.

Intestinal microbial metabolites have been increasingly recognized as important regulators of enteric viral infection. However, very little information is available about which specific microbiota-derived metabolites are crucial for swine enteric coronavirus (SECoV) infection in vivo. Using swine acute diarrhea syndrome (SADS)-CoV as a model, we were able to identify a greatly altered bile acid (BA) profile in the small intestine of infected piglets by untargeted metabolomic analysis. Using a newly established ex vivo model-the stem cell-derived porcine intestinal enteroid (PIE) culture-we demonstrated that certain BAs, cholic acid (CA) in particular, enhance SADS-CoV replication by acting on PIEs at the early phase of infection. We ruled out the possibility that CA exerts an augmenting effect on viral replication through classic farnesoid X receptor or Takeda G protein-coupled receptor 5 signaling, innate immune suppression or viral attachment. BA induced multiple cellular responses including rapid changes in caveolae-mediated endocytosis, endosomal acidification and dynamics of the endosomal/lysosomal system that are critical for SADS-CoV replication. Thus, our findings shed light on how SECoVs exploit microbiome-derived metabolite BAs to swiftly establish viral infection and accelerate replication within the intestinal microenvironment.

Although feline coronavirus (FCoV) infection is extremely common in cats, there are currently few effective treatments. A peptide derived from the heptad repeat 2 (HR2) domain of the coronavirus (CoV) spike protein has shown effective for inhibition of various human and animal CoVs in vitro, but further use of FCoV-HR2 in vivo has been limited by lack of practical delivery vectors and small animal infection model. To overcome these technical challenges, we first constructed a recombinant Bacillus subtilis (rBSCotB-HR2P) expressing spore coat protein B (CotB) fused to an HR2-derived peptide (HR2P) from a serotype II feline enteric CoV (FECV). Immunogenic capacity was evaluated in mice after intragastric or intranasal administration, showing that recombinant spores could trigger strong specific cellular and humoral immune responses. Furthermore, we developed a novel mouse model for FECV infection by transduction with its primary receptor (feline aminopeptidase N) using an E1/E3-deleted adenovirus type 5 vector. This model can be used to study the antiviral immune response and evaluate vaccines or drugs, and is an applicable choice to replace cats for the study of FECV. Oral administration of rBSCotB-HR2P in this mouse model effectively protected against FECV challenge and significantly reduced pathology in the digestive tract. Owing to its safety, low cost, and probiotic features, rBSCotB-HR2P is a promising oral vaccine candidate for use against FECV/FCoV infection in cats.

ABSTRACT Hepatitis E virus (HEV) is a major cause of acute viral hepatitis in humans. A convenient small mammalian model for basic research and antiviral testing is still greatly needed. Although a small rodent, the Mongolian gerbil, was reported to be susceptible to swine genotype-4 HEV infection, whether the previous results were reliable and consistent needs to be validated by using biologically pure HEV stocks or infectious RNA. In this study, we revisited this gerbil infection model for human HEV of genotype 1, 3, or 4 (G1, G3, or G4) by HEV reverse genetics. Gerbils inoculated intrahepatically with capped G3 HEV RNA transcripts or intraperitoneally with infectious G3 cloned HEV produced robust infection, as evidenced by presence of HEV in livers, spleens, and feces for up to 7 weeks post inoculation, seroconversion, and pathological liver lesions. Furthermore, the value of the gerbil model in antiviral testing and type I IFN in host defense was assessed. We demonstrated the effectiveness of peg-IFNα-2a and ribavirin in inhibiting HEV replication in gerbils. By treatment with two molecule inhibitors of TBK1, we also revealed a role of RIG-I like receptor-interferon regulatory factor 3 in host anti-HEV innate immune sensing in this in vivo model. Finally, susceptibility of G4 HEV was demonstrated in intrahepatically inoculated gerbils with infectious HEV RNA transcripts, whereas no evidence for G1 HEV susceptibility was found. The availability of the convenient gerbil model will greatly facilitate HEV-specific antiviral development and assess the mechanism of host immune response during HEV infection. IMPORTANCE HEV infects >20 million people annually, causing acute viral hepatitis as well as chronic hepatitis, neurological diseases, and pregnancy-associated high mortality, which require therapeutic intervention. The HEV antiviral research is largely limited by the lack of a convenient small animal model. Here we revisit the Mongolian gerbil model for three genotypes of human HEV by infectious HEV clones and recognized standards of experimental procedures. Fecal virus shedding, seroconversion, and pathological liver lesions could be detected in HEV-inoculated gerbils. We demonstrate the effectiveness and usefulness of this model in testing antiviral drugs, and in assessing the mechanism of host innate immune response upon HEV infection. This conventional rodent model will aid in future antiviral development and delineating mechanism of host immune response.

ABSTRACTPorcine deltacoronavirus (PDCoV) is a newly emerging threat to the global porcine industry. PDCoV has been successfully isolated using various medium additives including trypsin, and although we know it is important for viral replication, the mechanism has not been fully elucidated. Here, we systematically investigated the role of trypsin in PDCoV replication including cell entry, cell-to-cell membrane fusion and virus release. Using pseudovirus entry assays, we demonstrated that PDCoV entry is not trypsin dependent. Furthermore, unlike porcine epidemic diarrhea virus (PEDV), in which trypsin is important for the release of virus from infected cells, PDCoV release was not affected by trypsin. We also demonstrated that trypsin promotes PDCoV replication by enhancing cell-to-cell membrane fusion. Most importantly, our study illustrates two distinct spreading patterns from infected cells to uninfected cells during PDCoV transmission, and the role of trypsin in PDCoV replication in cells with different virus spreading types. Overall, these results clarify that trypsin promotes PDCoV replication by mediating cell-to-cell fusion transmission but is not crucial for viral entry. This knowledge can potentially contribute to improvement of virus production efficiency in culture, not only for vaccine preparation but also to develop antiviral treatments.

Here, the authors demonstrate a multi-momentum transformation metasurface. The orbital angular momentum meta-transformer reconstructs different vortex beams into on-axis distinct patterns, and the linear momentum meta-transformer converts red, green and blue beams to vivid color images.

Coronaviruses (CoVs) are a known global threat, and most recently the ongoing COVID-19 pandemic has claimed more than 2 million human lives. Delays and interference with IFN responses are closely associated with the severity of disease caused by CoV infection. As the most abundant viral protein in infected cells just after the entry step, the CoV nucleocapsid (N) protein likely plays a key role in IFN interruption. We have conducted a comprehensive comparative analysis and report herein that the N proteins of representative human and animal CoVs from four different genera [swine acute diarrhea syndrome CoV (SADS-CoV), porcine epidemic diarrhea virus (PEDV), severe acute respiratory syndrome CoV (SARS-CoV), SARS-CoV-2, Middle East respiratory syndrome CoV (MERS-CoV), infectious bronchitis virus (IBV) and porcine deltacoronavirus (PDCoV)] suppress IFN responses by multiple strategies. In particular, we found that the N protein of SADS-CoV interacted with RIG-I independent of its RNA binding activity, mediating K27-, K48- and K63-linked ubiquitination of RIG-I and its subsequent proteasome-dependent degradation, thus inhibiting the host IFN response. These data provide insight into the interaction between CoVs and host, and offer new clues for the development of therapies against these important viruses.

A PEDV/PDCoV/TGEV/SADS-CoV/XIPC 5-plex real-time RT-PCR was developed and validated for the simultaneous detection and differentiation of four swine enteric coronaviruses (PEDV, PDCoV, TGEV and SADS-CoV) in one PCR reaction (XIPC serves as an exogenous internal positive control). The 5-plex PCR had excellent analytical specificity, analytical sensitivity, and repeatability based on the testing of various viral and bacterial pathogens, serial dilutions of virus isolates, and in vitro transcribed RNAs. The 5-plex PCR had comparable diagnostic performance to a commercial PEDV/TGEV/PDCoV reference PCR, based on the testing of 219 clinical samples. Subsequently, 1807 clinical samples collected from various U.S. states during 2019–2021 were tested by the 5-plex PCR to investigate the presence of SADS-CoV in U.S. swine and the frequency of detecting swine enteric CoVs. All 1807 samples tested negative for SADS-CoV. Among the samples positive for swine enteric CoVs, there was a low frequency of detecting TGEV, an intermediate frequency of detecting PDCoV, and a high frequency of detecting PEDV. Although there is no evidence of SADS-CoV presence in the U.S. at present, the availability of the 5-plex PCR will enable us to conduct ongoing surveillance to detect and differentiate these viruses in swine samples and other host species samples as some of these coronaviruses can cause cross-species infection.

Flat optics, metasurfaces, metalenses, and related materials promise novel on-demand light modulation within ultrathin layers at wavelength scale, enabling a plethora of next-generation optical devices, also known as metadevices. Metadevices designed with different materials have been proposed and demonstrated for different applications, and the mass production of metadevices is necessary for metadevices to enter the consumer electronics market. However, metadevice manufacturing processes are mainly based on electron beam lithography, which exhibits low productivity and high costs for mass production. Therefore, processes compatible with standard complementary metal–oxide–semiconductor manufacturing techniques that feature high productivity, such as i-line stepper and nanoimprint lithography, have received considerable attention. This paper provides a review of current metasurfaces and metadevices with a focus on materials and manufacturing processes. We also provide an analysis of the relationship between the aspect ratio and efficiency of different materials.

Hepatitis E virus (HEV) is one of the major etiological agents responsible for acute hepatitis. Hepatitis E virus does not replicate efficiently in mammalian cell cultures, thus a useful model that mimics persistent HEV replication is needed to dissect the molecular mechanism of pathogenesis. Here we report a genotype-3 HEV RNA replicon expressing an EGFP-Zeocin (EZ) resistant gene (p6-EZ) that persistently self-replicated in cell lines of human (Huh-7-S10-3) or hamster (BHK-21) origin after transfection with in vitro RNA transcripts and subsequent drug screening. Two cell lines, S10-3-EZ and BHK-21-EZ, stably expressed EGFP in the presence of Zeocin during continuous passages. Both genomic and subgenomic HEV RNAs and viral replicase proteins were stably expressed in persistent HEV replicon cells. The values of the cell models in antiviral testing, innate immune RNA sensing and type I IFN in host defense were further demonstrated. We revealed a role of RIG-I like receptor-interferon regulatory factor 3 in host antiviral innate immune sensing during HEV replication. We further demonstrated that treatment with interferon (IFN-α) or ribavirin significantly reduced expression of replicon RNA in a dose-dependent manner. The availability of the models will greatly facilitate HEV-specific antiviral development, and delineate mechanisms of HEV replication.

Orbital angular momentum (OAM) beams with topological charge ℓ are commonly generated and detected by modulating an incoming field with an azimuthal phase profile of the form exp(iℓϕ) by a variety of approaches. This results in unwanted radial modes and reduced power in the desired OAM mode. Here, we show how to enhance the modal purity in the creation and detection of classical OAM beams and in the quantum detection of OAM photons. Classically, we combine holographic and metasurface control to produce high purity OAM modes and show how to detect them with high efficiency, extending the demonstration to the quantum realm with spatial light modulators. We demonstrate ultra-high purity OAM modes in orders as high as ℓ = 100 and a doubling of dimensionality in the quantum OAM spectrum from a spontaneous parametric downconversion source. Our work offers a simple route to increase the channel capacity in classical and quantum communication using OAM modes as a basis.

Porcine torovirus (PToV) is a potential enteric swine pathogen, found at especially high rates in piglets with diarrhea. It was first reported in the Netherlands in 1998 and has emerged in many countries around the world. Infections are generally asymptomatic and have not directly caused large economic losses, though co-infections with other swine pathogens and intertype recombination may lead to unpredictable outcomes. This review introduces progress in PToV research regarding its discovery, relationship with other Toroviruses, virion morphological characteristics, genetic structure and variation, recent epidemiology, diagnostic methods, and possibilities for future research.

To investigate the evolutionary process by which porcine epidemic diarrhea virus (PEDV) in the United States hypothetically descended from strains in China, we analyzed PEDV-positive samples collected in China during January 2012–July 2013. Recombination in 2 strain sublineages was likely associated with identification of PEDV in the United States in 2013.

Porcine deltacoronavirus (PDCoV) causes severe diarrhea and vomiting in affected piglets. The aim of this study was to establish the basic, in vitro characteristics of the life cycle such as replication kinetics, cellular ultrastructure, virion morphology, and induction of autophagy of PDCoV. Time-course analysis of viral subgenomic and genomic RNA loads and infectious titers indicated that one replication cycle of PDCoV takes 5 to 6 h. Electron microscopy showed that PDCoV infection induced the membrane rearrangements with double-membrane vesicles and large virion-containing vacuoles. The convoluted membranes structures described in alpha- and beta-coronavirus were not observed. PDCoV infection also increased the number of autophagosome-like vesicles in the cytoplasm of cells, and the autophagy response was detected by LC3 I/II and p62 Western blot analysis. For the first time, this study presents the picture of the PDCoV infection cycle, which is crucial to help elucidate the molecular mechanism of deltacoronavirus replication.

This paper develops a dielectrophoretic (DEP) chip with multi-layer electrodes and a micro-cavity array for programmable manipulations of cells and impedance measurement. The DEP chip consists of an ITO top electrode, flow chamber, middle electrode on an SU-8 surface, micro-cavity arrays of SU-8 and distributed electrodes at the bottom of the micro-cavity. Impedance sensing of single cells could be performed as follows: firstly, cells were trapped in a micro-cavity array by negative DEP force provided by top and middle electrodes; then, the impedance measurement for discrimination of different stage of bladder cancer cells was accomplished by the middle and bottom electrodes. After impedance sensing, the individual releasing of trapped cells was achieved by negative DEP force using the top and bottom electrodes in order to collect the identified cells once more. Both cell manipulations and impedance measurement had been integrated within a system controlled by a PC-based LabVIEW program. In the experiments, two different stages of bladder cancer cell lines (grade III: T24 and grade II: TSGH8301) were utilized for the demonstration of programmable manipulation and impedance sensing; as the results show, the lower-grade bladder cancer cells (TSGH8301) possess higher impedance than the higher-grade ones (T24). In general, the multi-step manipulations of cells can be easily programmed by controlling the electrical signal in our design, which provides an excellent platform technology for lab-on-a-chip (LOC) or a micro-total-analysis-system (Micro TAS).

Porcine deltacoronavirus (PDCoV), also known as porcine coronavirus HKU15, was first detected in North America in early 2014 and associated with enteric disease in pigs, resulting in an urgent need to further investigate the ecology of this virus. While assays detecting nucleic acids were implemented quickly, assays to detect anti-PDCoV antibodies have not been available. In this study, an indirect anti-PDCoV IgG enzyme-linked immunosorbent assay (ELISA) based on the putative S1 portion of the spike protein was developed and utilized to determine the prevalence of anti-PDCoV IgG in U.S. pigs. The diagnostic sensitivity of the PDCoV ELISA was 91% with a diagnostic specificity of 95%. A total of 968 serum samples were tested including samples with confirmed infection with PDCoV, porcine epidemic diarrhea virus (PEDV), transmissible gastroenteritis virus or porcine respiratory coronavirus. There was no cross-reactivity with any of the other coronaviruses. Among 355 arbitrarily selected serum samples collected in 2014 and originating from 51 farms across 18 U.S. states, anti-PDCoV IgG antibodies were detected in 8.7% of the samples and in 25.5% of the farms whereas anti-PEDV IgG was detected in 22.8% of the samples and in 54.9% of the farms. In addition, anti-PDCoV IgG antibodies were detected in archived samples collected in 2010, perhaps indicating an earlier undetected introduction into the U.S. pig population. Overall, the obtained data suggest that PDCoV seroprevalence in U.S. pigs is lower compared to PEDV and PDCoV may have been introduced to the U.S. prior to PEDV.

ABSTRACT Coronaviruses are known to infect humans and other animals and cause respiratory and gastrointestinal diseases. Here we report the emergence of porcine epidemic diarrhea virus (PEDV) in the United States and determination of its origin, evolution, and genotypes based on temporal and geographical evidence. Histological lesions in small intestine sections of affected pigs and the complete genomic sequences of three emergent strains of PEDV isolated from outbreaks in Minnesota and Iowa were characterized. Genetic and phylogenetic analyses of the three U.S. strains revealed a close relationship with Chinese PEDV strains and their likely Chinese origin. The U.S. PEDV strains underwent evolutionary divergence, which can be classified into two sublineages. The three emergent U.S. strains are most closely related to a strain isolated in 2012 from Anhui Province in China, which might be the result of multiple recombination events between different genetic lineages or sublineages of PEDV. Molecular clock analysis of the divergent time based on the complete genomic sequences is consistent with the actual time difference, approximately 2 to 3 years, of the PED outbreaks between China (December 2010) and the United States (May 2013). The finding that the emergent U.S. PEDV strains share unique genetic features at the 5′-untranslated region with a bat coronavirus provided further support of the evolutionary origin of PEDV from bats and potential cross-species transmission. The data from this study have important implications for understanding the ongoing PEDV outbreaks in the United States and will guide future efforts to develop effective preventive and control measures against PEDV. IMPORTANCE The sudden emergence of porcine epidemic diarrhea virus (PEDV), a coronavirus, for the first time in the United States causes significant economic and public health concerns. Since its recognition in May 2013, PEDV has rapidly spread across the United States, resulting in high mortality in piglets in more than 17 States now. The ongoing outbreaks of Middle East respiratory syndrome coronavirus in humans from countries in or near the Arabian Peninsula and the historical deadly nature of the 2002 outbreaks of severe acute respiratory syndrome coronavirus create further anxiety over the emergence of PEDV in the United States due to the lack of scientific information about the origin and evolution of this emerging coronavirus. Here we report the detailed genetic characterization, origin, and evolution of emergent PEDV strains in the United States. The results provide much needed information to devise effective preventive and control strategies against PEDV in the United States.

Porcine reproductive and respiratory syndrome virus (PRRSV) is one of the most economically important swine pathogens, which causes reproductive failure in sows and respiratory disease in piglets. A major hurdle to control PRRSV is the ineffectiveness of the current vaccines to confer protection against heterologous strains. Since both GP4 and M genes of PRRSV induce neutralizing antibodies, in this study we molecularly bred PRRSV through DNA shuffling of the GP4 and M genes, separately, from six genetically different strains of PRRSV in an attempt to identify chimeras with improved heterologous cross-neutralizing capability. The shuffled GP4 and M genes libraries were each cloned into the backbone of PRRSV strain VR2385 infectious clone pIR-VR2385-CA. Three GP4-shuffled chimeras and five M-shuffled chimeras, each representing sequences from all six parental strains, were selected and further characterized in vitro and in pigs. These eight chimeric viruses showed similar levels of replication with their backbone strain VR2385 both in vitro and in vivo, indicating that the DNA shuffling of GP4 and M genes did not significantly impair the replication ability of these chimeras. Cross-neutralization test revealed that the GP4-shuffled chimera GP4TS14 induced significantly higher cross-neutralizing antibodies against heterologous strains FL-12 and NADC20, and similarly that the M-shuffled chimera MTS57 also induced significantly higher levels of cross-neutralizing antibodies against heterologous strains MN184B and NADC20, when compared with their backbone parental strain VR2385 in infected pigs. The results suggest that DNA shuffling of the GP4 or M genes from different parental viruses can broaden the cross-neutralizing antibody-inducing ability of the chimeric viruses against heterologous PRRSV strains. The study has important implications for future development of a broadly protective vaccine against PRRSV.