Unraveling the assembly mechanism of SADS-CoV virus nucleocapsid protein: insights from RNA binding, dimerization, and epitope diversity profiling.

The swine acute diarrhea syndrome coronavirus (SADS-CoV) has caused significant disruptions in porcine breeding and raised concerns about potential human infection. The nucleocapsid (N) protein of SADS-CoV plays a vital role in viral assembly and replication, but its structure and functions remain poorly understood. This study utilized biochemistry, X-ray crystallography, and immunization techniques to investigate the N protein's structure and function in SADS-CoV. Our findings revealed distinct domains within the N protein, including an RNA-binding domain, two disordered domains, and a dimerization domain. Through biochemical assays, we confirmed that the N-terminal domain functions as an RNA-binding domain, and the C-terminal domain is involved in dimerization, with the crystal structure analysis providing visual evidence of dimer formation. Immunization experiments demonstrated that the disordered domain 2 elicited a significant antibody response. These identified domains and their interactions are crucial for viral assembly. This comprehensive understanding of the N protein in SADS-CoV enhances our knowledge of its assembly and replication mechanisms, enabling the development of targeted interventions and therapeutic strategies. IMPORTANCE: SADS-CoV is a porcine coronavirus that originated from a bat HKU2-related coronavirus. It causes devastating swine diseases and poses a high risk of spillover to humans. The coronavirus N protein, as the most abundant viral protein in infected cells, likely plays a key role in viral assembly and replication. However, the structure and function of this protein remain unclear. Therefore, this study employed a combination of biochemistry and X-ray crystallography to uncover distinct structural domains in the N protein, including RNA-binding domains, two disordered domains, and dimerization domains. Additionally, we made the novel discovery that the disordered domain elicited a significant antibody response. These findings provide new insights into the structure and functions of the SADS-CoV N protein, which have important implications for future studies on SADS-CoV diagnosis, as well as the development of vaccines and anti-viral drugs.

Establishment of replication-competent vesicular stomatitis virus recapitulating SADS-CoV entry.

Zoonotic coronaviruses pose a continuous threat to human health, with newly identified bat-borne viruses like swine acute diarrhea syndrome coronavirus (SADS-CoV) causing high mortality in piglets. In vitro studies indicate that SADS-CoV can infect cell lines from diverse species, including humans, highlighting its potential risk to human health. However, the lack of tools to study viral entry, along with the absence of vaccines or antiviral therapies, perpetuates this threat. To address this, we engineered an infectious molecular clone of Vesicular Stomatitis Virus (VSV), replacing its native glycoprotein (G) with SADS-CoV spike (S) and inserting a Venus reporter at the 3' leader region to generate a replication-competent rVSV-Venus-SADS S virus. Serial passages of rVSV-Venus-SADS S led to the identification of an 11-amino-acid truncation in the cytoplasmic tail of the S protein, which allowed more efficient viral propagation due to increased cell membrane anchoring of the S protein. The S protein was integrated into rVSV-Venus-SADS SΔ11 particles, susceptible to neutralization by sera from SADS-CoV S1 protein-immunized rabbits. Additionally, we found that TMPRSS2 promotes SADS-CoV spike-mediated cell entry. Furthermore, we assessed the serum-neutralizing ability of mice vaccinated with rVSV-Venus-SADS SΔ11 using a prime-boost immunization strategy, revealing effective neutralizing antibodies against SADS-CoV infection. In conclusion, we have developed a safe and practical tool for studying SADS-CoV entry and exploring the potential of a recombinant VSV-vectored SADS-CoV vaccine.IMPORTANCEZoonotic coronaviruses, like swine acute diarrhea syndrome coronavirus (SADS-CoV), pose a continual threat to human and animal health. To combat this, we engineered a safe and efficient tool by modifying the Vesicular Stomatitis Virus (VSV), creating a replication-competent rVSV-Venus-SADS S virus. Through serial passages, we optimized the virus for enhanced membrane anchoring, a key factor in viral propagation. This modified virus, rVSV-Venus-SADS SΔ11, proved susceptible to neutralization, opening avenues for potential vaccines. Additionally, our study revealed the role of TMPRSS2 in SADS-CoV entry. Mice vaccinated with rVSV-Venus-SADS SΔ11 developed potent neutralizing antibodies against SADS-CoV. In conclusion, our work presents a secure and practical tool for studying SADS-CoV entry and explores the promise of a recombinant VSV-vectored SADS-CoV vaccine.

Functional dissection of the spike glycoprotein S1 subunit and identification of cellular cofactors for regulation of swine acute diarrhea syndrome coronavirus entry.

Swine acute diarrhea syndrome coronavirus (SADS-CoV) is a novel porcine enteric coronavirus, and the broad interspecies infection of SADS-CoV poses a potential threat to human health. This study provides experimental evidence to dissect the roles of distinct domains within the SADS-CoV spike S1 subunit in cellular entry. Specifically, we expressed the S1 and its subdomains, S1A and S1B. Cell binding and invasion inhibition assays revealed a preference for the S1B subdomain in binding to the receptors on the cell surface, and this unknown receptor is not utilized by the porcine epidemic diarrhea virus. Nanoparticle display demonstrated hemagglutination of erythrocytes from pigs, humans, and mice, linking the S1A subdomain to the binding of sialic acid (Sia) involved in virus attachment. We successfully rescued GFP-labeled SADS-CoV (rSADS-GFP) from a recombinant cDNA clone to track viral infection. Antisera raised against S1, S1A, or S1B contained highly potent neutralizing antibodies, with anti-S1B showing better efficiency in neutralizing rSADS-GFP infection compared to anti-S1A. Furthermore, depletion of heparan sulfate (HS) by heparinase treatment or pre-incubation of rSADS-GFP with HS or constituent monosaccharides could inhibit SADS-CoV entry. Finally, we demonstrated that active furin cleavage of S glycoprotein and the presence of type II transmembrane serine protease (TMPRSS2) are essential for SADS-CoV infection. These combined observations suggest that the wide cell tropism of SADS-CoV may be related to the distribution of Sia or HS on the cell surface, whereas the S1B contains the main protein receptor binding site. Specific host proteases also play important roles in facilitating SADS-CoV entry.IMPORTANCESwine acute diarrhea syndrome coronavirus (SADS-CoV) is a novel pathogen infecting piglet, and its unique genetic evolution characteristics and broad species tropism suggest the potential for cross-species transmission. The virus enters cells through its spike (S) glycoprotein. In this study, we identify the receptor binding domain on the C-terminal part of the S1 subunit (S1B) of SADS-CoV, whereas the sugar-binding domain located at the S1 N-terminal part of S1 (S1A). Sialic acid, heparan sulfate, and specific host proteases play essential roles in viral attachment and entry. The dissection of SADS-CoV S1 subunit's functional domains and identification of cellular entry cofactors will help to explore the receptors used by SADS-CoV, which may contribute to exploring the mechanisms behind cross-species transmission and host tropism.

NS7a of SADS-CoV promotes viral infection via inducing apoptosis to suppress type III interferon production.

Swine acute diarrhea syndrome coronavirus (SADS-CoV) is a newly discovered swine coronavirus with potential cross-species transmission risk. Although SADS-CoV-induced host cell apoptosis and innate immunity antagonization has been revealed, underlying signaling pathways remain obscure. Here, we demonstrated that infection of SADS-CoV induced apoptosis in vivo and in vitro, and that viral protein NS7a is mainly responsible for SADS-CoV-induced apoptosis in host cells. Furthermore, we found that NS7a interacted with apoptosis-inducing factor mitochondria associated 1 (AIFM1) to activate caspase-3 via caspase-6 in SADS-CoV-infected cells, and enhanced SADS-CoV replication. Importantly, NS7a suppressed poly(I:C)-induced expression of type III interferon (IFN-λ) via activating caspase-3 to cleave interferon regulatory factor 3 (IRF3), and caspase-3 inhibitor protects piglets against SADS-CoV infection in vivo. These findings reveal how SADS-CoV induced apoptosis to inhibit innate immunity and provide a valuable clue to the development of effective drugs for the clinical control of SADS-CoV infection.IMPORTANCEOver the last 20 years, multiple animal-originated coronaviruses, including severe acute respiratory syndrome coronavirus (SARS-CoV), middle east respiratory syndrome coronavirus (MERS-CoV), and SARS-CoV-2, have caused millions of deaths, seriously jeopardized human health, and hindered social development, indicating that the study of animal-originated coronaviruses with potential for cross-species transmission is particularly important. Bat-originated swine acute diarrhea syndrome coronavirus (SADS-CoV), discovered in 2017, can not only cause fatal diarrhea in piglets, but also infect multiple human cells, with a potential risk of cross-species transmission, but its pathogenesis is unclear. In this study, we demonstrated that NS7a of SADS-CoV suppresses IFN-λ production via apoptosis-inducing factor mitochondria associated 1 (AIFM1)-caspase-6-caspase-3-interferon regulatory factor 3 (IRF3) pathway, and caspase-3 inhibitor (Z-DEVD-FMK) can effectively inhibit SADS-CoV replication and protect infected piglets. Our findings in this study contribute to a better understanding of SADS-CoV-host interactions as a part of the coronaviruses pathogenesis and using apoptosis-inhibitor as a drug as potential therapeutic approaches for prevention and control of SADS-CoV infection.

Bat-Origin Swine Acute Diarrhea Syndrome Coronavirus Is Lethal to Neonatal Mice.

Bats are reservoirs for diverse coronaviruses, including swine acute diarrhea syndrome coronavirus (SADS-CoV). SADS-CoV has been reported to have broad cell tropism and inherent potential to cross host species barriers for dissemination. We rescued synthetic wild-type SADS-CoV using one-step assembly of a viral cDNA clone by homologous recombination in yeast. Furthermore, we characterized SADS-CoV replication in vitro and in neonatal mice. We found that SADS-CoV caused severe watery diarrhea, weight loss, and a 100% fatality rate in 7- and 14-day-old mice after intracerebral infection. We also detected SADS-CoV-specific N protein in the brain, lungs, spleen, and intestines of infected mice. Furthermore, SADS-CoV infection triggers excessive cytokine expression that encompasses a broad array of proinflammatory mediators, including interleukin 1ß (IL-1ß), IL-6, IL-8, tumor necrosis factor alpha (TNF-α), C-X-C motif chemokine ligand 10 (CXCL10), interferon beta (IFN-ß), IFN-γ, and IFN-λ3. This study highlights the importance of identifying neonatal mice as a model for developing vaccines or antiviral drugs against SADS-CoV infection. IMPORTANCE SADS-CoV is the documented spillover of a bat coronavirus that causes severe disease in pigs. Pigs are in frequent contact with both humans and other animals and theoretically possess a greater chance, compared to many other species, of promoting cross-species viral transmission. SADS-CoV has been reported to have broad cell tropism and inherent potential to cross host species barriers for dissemination. Animal models are an essential feature of the vaccine design toolkit. Compared with neonatal piglets, the mouse is small, making it an economical choice for animal models for SADS-CoV vaccine design. This study showed the pathology of neonatal mice infected with SADS-CoV, which should be very useful for vaccine and antiviral studies.

N-linked glycoproteins and host proteases are involved in swine acute diarrhea syndrome coronavirus entry.

IMPORTANCE: Gaining insight into the cell-entry mechanisms of swine acute diarrhea syndrome coronavirus (SADS-CoV) is critical for investigating potential cross-species infections. Here, we demonstrated that pretreatment of host cells with tunicamycin decreased SADS-CoV attachment efficiency, indicating that N-linked glycosylation of host cells was involved in SADS-CoV entry. Common N-linked sugars Neu5Gc and Neu5Ac did not interact with the SADS-CoV S1 protein, suggesting that these molecules were not involved in SADS-CoV entry. Additionally, various host proteases participated in SADS-CoV entry into diverse cells with different efficiencies. Our findings suggested that SADS-CoV may exploit multiple pathways to enter cells, providing insights into intervention strategies targeting the cell entry of this virus.

Evaluation of serum Epstein-Barr virus envelope glycoproteins antibodies and their association with systemic autoimmune diseases.

Systemic autoimmune diseases (SADs) are a growing spectrum of autoimmune disorders that commonly affect multiple organs. The role of Epstein-Barr virus (EBV) infection or reactivation as a trigger for the initiation and progression of SADs has been established, while the relationship between EBV envelope glycoproteins and SADs remains unclear. Here, we assessed the levels of IgG, IgA, and IgM against EBV glycoproteins (including gp350, gp42, gHgL, and gB) in serum samples obtained from patients with rheumatoid arthritis (RA) and systemic lupus erythematosus (SLE), and found that RA and SLE patients exhibited a statistically significant increase in the levels of 8 and 11 glycoprotein antibodies, respectively, compared to healthy controls (p < 0.05). The LASSO model identified four factors as significant diagnostic markers for RA: gp350 IgG, gp350 IgA, gHgL IgM, and gp42 IgA; whereas for SLE it included gp350 IgG, gp350 IgA, gHgL IgA, and gp42 IgM. Combining these selected biomarkers yielded an area under the curve (AUC) of 0.749 for RA and 0.843 for SLE. We subsequently quantified the levels of autoantibodies associated with SADs in mouse sera following immunization with gp350. Remarkably, none of the tested autoantibody levels exhibited statistically significant alterations. Elevation of glycoprotein antibody concentration suggests that Epstein-Barr virus reactivation and replication occurred in SADs patients, potentially serving as a promising biomarker for diagnosing SADs. Moreover, the absence of cross-reactivity between gp350 antibodies and SADs-associated autoantigens indicates the safety profile of a vaccine based on gp350 antigen.

Validation of the Individualized Metabolic Surgery score in predicting long-term remission of diabetes after duodenal switch-type procedures.

AIM: To validate the Individualized Metabolic Surgery (IMS) score and assess long-term remission of type 2 diabetes (T2D) after duodenal switch (DS)-type procedures in patients with obesity. In addition, to help guide metabolic procedure selection for those patients categorized as having severe T2D. MATERIALS AND METHODS: This is a retrospective single cohort study of all patients with T2D and severe obesity, who underwent DS-type procedures at a single institution from December 2010 to December 2018. Study endpoints included validating the IMS score in our cohort and evaluating the impact of DS-type procedures on long-term (≥ 5 years) remission of T2D, especially in patients with severe disease. A receiver operator characteristic curve was used to assess the accuracy of the IMS score using the area under the curve (AUC). RESULTS: The study cohort included 30 patients with complete baseline and long-term glycaemic data after their index DS-type surgery. Twelve patients (40%) were classified with severe T2D, and the distribution of IMS-based severity groups was similar between our cohort and the original IMS study (P = .42). IMS scores predicted long-term T2D remission with AUC = 0.77. Patients with IMS-based severe diabetes achieved significantly higher long-term remission after DS-type procedures compared with gastric bypass and/or sleeve gastrectomy from the original IMS study (42% vs. 12%; P < .05). CONCLUSIONS: The IMS score properly classifies the severity of T2D in our study cohort and adequately predicts its long-term remission after DS-type procedures. While T2D remission decreases with more severe IMS scores, long-term remission remains high after DS-type procedures among patients with severe disease.

Quercetin inhibition of porcine intestinal alpha coronavirus in vitro and in vivo.

BACKGROUND: Porcine acute diarrhea syndrome coronavirus (SADS-CoV) is one of the novel pathogens responsible for piglet diarrhea, contributing to substantial economic losses in the farming sector. The broad host range of SADS-CoV raises concerns regarding its potential for cross-species transmission. Currently, there are no effective means of preventing or treating SADS-CoV infection, underscoring the urgent need for identifying efficient antiviral drugs. This study focuses on evaluating quercetin as an antiviral agent against SADS-CoV. RESULTS: In vitro experiments showed that quercetin inhibited SADS-CoV proliferation in a concentration-dependent manner, targeting the adsorption and replication stages of the viral life cycle. Furthermore, quercetin disrupts the regulation of the P53 gene by the virus and inhibits host cell cycle progression induced by SADS-CoV infection. In vivo experiments revealed that quercetin effectively alleviated the clinical symptoms and intestinal pathological damage caused by SADS-CoV-infected piglets, leading to reduced expression levels of inflammatory factors such as TLR3, IL-6, IL-8, and TNF-α. CONCLUSIONS: Therefore, this study provides compelling evidence that quercetin has great potential and promising applications for anti- SADS-CoV action.

Global abundance estimates for 9,700 bird species.

Quantifying the abundance of species is essential to ecology, evolution, and conservation. The distribution of species abundances is fundamental to numerous longstanding questions in ecology, yet the empirical pattern at the global scale remains unresolved, with a few species' abundance well known but most poorly characterized. In large part because of heterogeneous data, few methods exist that can scale up to all species across the globe. Here, we integrate data from a suite of well-studied species with a global dataset of bird occurrences throughout the world-for 9,700 species (∼92% of all extant species)-and use missing data theory to estimate species-specific abundances with associated uncertainty. We find strong evidence that the distribution of species abundances is log left skewed: there are many rare species and comparatively few common species. By aggregating the species-level estimates, we find that there are ∼50 billion individual birds in the world at present. The global-scale abundance estimates that we provide will allow for a line of inquiry into the structure of abundance across biogeographic realms and feeding guilds as well as the consequences of life history (e.g., body size, range size) on population dynamics. Importantly, our method is repeatable and scalable: as data quantity and quality increase, our accuracy in tracking temporal changes in global biodiversity will increase. Moreover, we provide the methodological blueprint for quantifying species-specific abundance, along with uncertainty, for any organism in the world.

Dimensional profiling of psychopathology in children and adolescents based on the K-SADS-PL and an analysis of the construct validity of two ADHD symptom dimensions.

OBJECTIVES: The traditional view on psychiatric disorders as categorical and distinct is being challenged by perspectives emphasizing the relevance of dimensional and transdiagnostic assessment. However, most diagnostic instruments are based on a categorical view with a threshold-approach to disease classification. METHODS: We here describe algorithms for dimensionalizing the psychopathological ratings of the widely used diagnostic interview for children and adolescents, the Kiddie-Schedule for Affective Disorders and Schizophrenia - Present and Lifetime Version (K-SADS-PL). We further evaluate the criterion-related construct validity of the dimensionalized attention-deficit/hyperactivity disorder (ADHD) scales using Rasch models in a sample of 590 children (mean age 10.29 (.36), 49% girls). RESULTS: The algorithms generate scores of current symptom load, i.e., the sum of clinician-rated symptoms within each disorder assessed with the interview. We found support for counting symptoms of inattention and hyperactivity/impulsivity, respectively, but not for a single combined ADHD scale. CONCLUSIONS: The algorithms constitute an initial step in creating a framework for clinician-rated dimensional analyses of symptoms derived from the K-SADS-PL, but future studies are needed to further evaluate the construct validity of the remaining scales and the reliability and clinical utility of the method. We believe that our proposed algorithms offer a novel method of dimensional psychopathological assessment, which can be applied in multiple branches of child and adolescent psychiatry.

Comprehensive Subcellular Localization of Swine Acute Diarrhea Syndrome Coronavirus Proteins.

Bats are reservoirs for diverse coronaviruses, including swine acute diarrhea syndrome coronavirus (SADS-CoV). SADS-CoV was first identified in diarrheal piglets in 2017. As a novel alphacoronavirus, SADS-CoV shares ~95% identity with bat alphacoronavirus HKU2. SADS-CoV has been reported to have broad cell tropism and inherent potential to cross host species barriers for dissemination. Thus far, no effective antiviral drugs or vaccines are available to treat infections with SADS-CoV. Therefore, knowledge of the protein-coding gene set and a subcellular localization map of SADS-CoV proteins are fundamental first steps in this endeavor. Here, all SADS-CoV genes were cloned separately into Flag-tagged plasmids, and the subcellular localizations of viral proteins, with the exception of nsp11, were detected using confocal microscopy techniques. As a result, nsp1, nsp3-N, nsp4, nsp5, nsp7, nsp8, nsp9, nsp10, nsp14, and nsp15 were localized in the cytoplasm and nuclear spaces, and these viral proteins may perform specific functions in the nucleus. All structural and accessory proteins were mainly localized in the cytoplasm. NS7a and membrane protein M colocalized with the Golgi compartment, and they may regulate the assembly of SADS-CoV virions. Maturation of SADS-CoV may occur in the late endosomes, during which envelope protein E is involved in the assembly and release of the virus. In summary, the present study demonstrates for the first time the location of all the viral proteins of SADS-CoV. These fundamental studies of SADS-CoV will promote studies of basic virology of SADS-CoV and support preventive strategies for animals with infection of SADS-CoV. IMPORTANCE SADS-CoV is the first documented spillover of a bat coronavirus that causes severe diseases in domestic animals. Our study is an in-depth annotation of the newly discovered swine coronavirus SADS-CoV genome and viral protein expression. Systematic subcellular localization of SADS-CoV proteins can have dramatic significance in revealing viral protein biological functions in the subcellular locations. Furthermore, our study promote understanding the fundamental science behind the novel swine coronavirus to pave the way for treatments and cures.

Lethal Swine Acute Diarrhea Syndrome Coronavirus Infection in Suckling Mice.

Swine acute diarrhea syndrome coronavirus (SADS-CoV) is a recently emerging bat-borne coronavirus responsible for high mortality rates in piglets. In vitro studies have indicated that SADS-CoV has a wide tissue tropism in different hosts, including humans. However, whether this virus potentially threatens other animals remains unclear. Here, we report the experimental infection of wild-type BALB/c and C57BL/6J suckling mice with SADS-CoV. We found that mice less than 7 days old are susceptible to the virus, which caused notable multitissue infections and damage. The mortality rate was the highest in 2-day-old mice and decreased in older mice. Moreover, a preliminary neuroinflammatory response was observed in 7-day-old SADS-CoV-infected mice. Thus, our results indicate that SADS-CoV has potential pathogenicity in young hosts. IMPORTANCE SADS-CoV, which likely has originated from bat coronaviruses, is highly pathogenic to piglets and poses a threat to the swine industry. Little is known about its potential to disseminate to other animals. No efficient treatment is available, and the quarantine strategy is the only preventive measure. In this study, we demonstrated that SADS-CoV can efficiently replicate in suckling mice younger than 7 days. In contrast to infected piglets, in which intestinal tropism is shown, SADS-CoV caused infection and damage in all murine tissues evaluated in this study. In addition, neuroinflammatory responses were detected in some of the infected mice. Our work provides a preliminary cost-effective model for the screening of antiviral drugs against SADS-CoV infection.

The origin and evolution of emerged swine acute diarrhea syndrome coronavirus with zoonotic potential.

Swine acute diarrhea syndrome coronavirus (SADS-CoV) is a newly discovered alphacoronavirus with zoonotic potential that causes diarrhea and vomiting mainly in piglets. Having emerged suddenly in 2017, the prevailing opinion is that the virus originated from HKU2, an alphacoronavirus whose primary host is bats, and at some unknown point achieved interspecies transmission via some intermediate. Here, we further explore the evolutionary history and possible cross-species transmission event for SADS-CoV. Coevolutionary analysis demonstrated that HKU2 may have achieved host switch via SADS-related (SADSr)-CoV, which was isolated from the genus Rhinolophus in 2017. SADS-CoV, HKU2, and SADSr-CoV share similar codon usage patterns and showed a lower tendency to use CpG, which may reflect a method of immune escape. The analyses of virus-host coevolution and recombination support SADSr-CoV is the direct source of SADS-CoV that may have undergone recombination events during its formation. Structure-based spike glycoprotein variance analysis revealed a more nuanced evolutionary pathway to receptor recognition for host switch. We did not find a possible positive selection site, and the dN/dS of the S gene was only 0.29, which indicates that the current SADS-CoV is slowly evolving. These results provide new insights that may help predict future cross-species transmission, and possibly surveil future zoonotic outbreaks and associated public health emergencies.

SADS-CoV nsp1 inhibits the IFN-ß production by preventing TBK1 phosphorylation and inducing CBP degradation.

Swine acute diarrhea syndrome (SADS) is first reported in January 2017 in Southern China. It subsequently causes widespread outbreaks in multiple pig farms, leading to economic losses. Therefore, it is an urgent to understand the molecular mechanisms underlying the pathogenesis and immune evasion of Swine acute diarrhea syndrome coronavirus (SADS-CoV). Our research discovered that SADS-CoV inhibited the production of interferon-ß (IFN-ß) during viral infection. The nonstructural protein 1 (nsp1) prevented the phosphorylation of TBK1 by obstructing the interaction between TBK1 and Ub protein. Moreover, nsp1 induced the degradation of CREB-binding protein (CBP) through the proteasome-dependent pathway, thereby disrupting the IFN-ß enhancer and inhibiting IFN transcription. Finally, we identified nsp1-Phe39 as the critical amino acid that downregulated IFN production. In conclusion, our findings described two mechanisms in nsp1 that inhibited IFN production and provided new insights into the evasion strategy adopted by SADS-CoV to evade host antiviral immunity.

Development of a novel double-antibody sandwich quantitative ELISA for detecting SADS-CoV infection.

Swine acute diarrhea syndrome coronavirus (SADS-CoV) is an emerging swine enteric alphacoronavirus that can cause acute diarrhea, vomiting, dehydration, and death of newborn piglets. In this study, we developed a double-antibody sandwich quantitative enzyme-linked immunosorbent assay (DAS-qELISA) for detection of SADS-CoV by using an anti-SADS-CoV N protein rabbit polyclonal antibody (PAb) and a specific monoclonal antibody (MAb) 6E8 against the SADS-CoV N protein. The PAb was used as the capture antibodies and HRP-labeled 6E8 as the detector antibody. The detection limit of the developed DAS-qELISA assay was 1 ng/mL of purified antigen and 101.08TCID50/mL of SADS-CoV, respectively. Specificity assays showed that the developed DAS-qELISA has no cross-reactivity with other swine enteric coronaviruses, such as porcine epidemic diarrhea virus (PEDV), transmissible gastroenteritis virus (TGEV), and porcine deltacoronavirus (PDCoV). Three-day-old piglets were challenged with SADS-CoV and collected anal swab samples which were screened for the presence of SADS-CoV by using DAS-qELISA and reverse transcriptase PCR (RT-PCR). The coincidence rate of the DAS-qELISA and RT-PCR was 93.93%, and the kappa value was 0.85, indicating that DAS-qELISA is a reliable method for applying antigen detection of clinical samples. KEY POINTS: • The first double-antibody sandwich quantitative enzyme-linked immunosorbent assay for detection SADS-CoV infection. • The custom ELISA is useful for controlling the SADS-CoV spread.

Navigating young minds: reliability and validity of the Greek version of kiddie - schedule for affective disorders and schizophrenia - present and lifetime DSM-5 version (K-SADS-PL-GR-5).

BACKGROUND: The Kiddie-Schedule for Affective Disorders and Schizophrenia-Present and Lifetime Version (K-SADS-PL) is one of the most popular semi-structured psychiatric interviews for children and adolescents. Its latest DSM-5 version (K-SADS-PL DSM-5) has only recently been adapted and validated in various languages. In the present study, we aimed to investigate the reliability and validity of the Greek version of the K-SADS-PL DSM-5. METHODS: A total of 137 patients consecutively referred for admission, aged 7-17, were included. The K-SADS-PL DSM-IV was translated and adapted to correspond to DSM-5 categories. Convergent and divergent validity were assessed against two self-report rating scales, Children's Depression Inventory (CDI) and Screen for Child Anxiety Related Emotional Disorders (SCARED). Inter-rater reliability was calculated exclusively for instances where a diagnosis involved three or more patients. RESULTS: Our findings revealed good to excellent inter-rater reliability and good to excellent consensual validity across most psychiatric diagnoses, except for panic disorder. Diagnostic efficiency, measured by sensitivity, specificity, positive and negative predictive values, consistently showed high specificity and negative predictive validity across all diagnostic categories. CONCLUSIONS: These findings support the applicability of the Greek version of the K-SADS-PL DSM-5 as a reliable and valid diagnostic tool in Greek-speaking populations.

Swine acute diarrhea syndrome coronavirus replication in primary human cells reveals potential susceptibility to infection.

Zoonotic coronaviruses represent an ongoing threat, yet the myriads of circulating animal viruses complicate the identification of higher-risk isolates that threaten human health. Swine acute diarrhea syndrome coronavirus (SADS-CoV) is a newly discovered, highly pathogenic virus that likely evolved from closely related HKU2 bat coronaviruses, circulating in Rhinolophus spp. bats in China and elsewhere. As coronaviruses cause severe economic losses in the pork industry and swine are key intermediate hosts of human disease outbreaks, we synthetically resurrected a recombinant virus (rSADS-CoV) as well as a derivative encoding tomato red fluorescent protein (tRFP) in place of ORF3. rSADS-CoV replicated efficiently in a variety of continuous animal and primate cell lines, including human liver and rectal carcinoma cell lines. Of concern, rSADS-CoV also replicated efficiently in several different primary human lung cell types, as well as primary human intestinal cells. rSADS-CoV did not use human coronavirus ACE-2, DPP4, or CD13 receptors for docking and entry. Contemporary human donor sera neutralized the group I human coronavirus NL63, but not rSADS-CoV, suggesting limited human group I coronavirus cross protective herd immunity. Importantly, remdesivir, a broad-spectrum nucleoside analog that is effective against other group 1 and 2 coronaviruses, efficiently blocked rSADS-CoV replication in vitro. rSADS-CoV demonstrated little, if any, replicative capacity in either immune-competent or immunodeficient mice, indicating a critical need for improved animal models. Efficient growth in primary human lung and intestinal cells implicate SADS-CoV as a potential higher-risk emerging coronavirus pathogen that could negatively impact the global economy and human health.

Evaluating the Interrater Reliability of the Icelandic Version of K-SADS-PL DSM-5.

The Schedule for Affective Disorders and Schizophrenia for School-Age Children-Present and Lifetime version (K-SADS-PL) is a valuable tool for diagnosing mental disorders in children and adolescents. Previous studies have examined its interrater reliability, but there is limited information on individual disorders, on the updated DSM-5 version. This study aims to analyse the interrater reliability of the Icelandic translation of K-SADS-PL, DSM-5 version. K-SADS-PL was administered to a clinical sample of outpatients from the Icelandic Anxiety Centre for Children, Adolescents, and Young Adults, and The Department of Child and Adolescent Psychiatry at Landspítali, the National University Hospital in Reykjavík, Iceland. In total, 135 patients aged 6-18 were included in this study. We assessed the interrater reliability using Cohen's κ, with results ranging from poor to excellent (0.3-1.0), though most disorders showed excellent reliability (κ > 0.75). The Icelandic translation of the DSM-5 K-SADS-PL is generally reliable when used by properly trained post-graduate students, which supports its use in clinical settings.

Dietary abundance distributions: Dominance and diversity in vertebrate diets.

Diet composition is among the most important yet least understood dimensions of animal ecology. Inspired by the study of species abundance distributions (SADs), we tested for generalities in the structure of vertebrate diets by characterising them as dietary abundance distributions (DADs). We compiled data on 1167 population-level diets, representing >500 species from six vertebrate classes, spanning all continents and oceans. DADs near-universally (92.5%) followed a hollow-curve shape, with scant support for other plausible rank-abundance-distribution shapes. This strong generality is inherently related to, yet incompletely explained by, the SADs of available food taxa. By quantifying dietary generalisation as the half-saturation point of the cumulative distribution of dietary abundance (sp50, minimum number of foods required to account for 50% of diet), we found that vertebrate populations are surprisingly specialised: in most populations, fewer than three foods accounted for at least half the diet. Variation in sp50 was strongly associated with consumer type, with carnivores being more specialised than herbivores or omnivores. Other methodological (sampling method and effort, taxonomic resolution), biological (body mass, frugivory) and biogeographic (latitude) factors influenced sp50 to varying degrees. Future challenges include identifying the mechanisms underpinning the hollow-curve DAD, its generality beyond vertebrates, and the biological determinants of dietary generalisation.