Polychromatic and collimated lights generated by nondegenerate four-wave mixing in cesium vapor.

We demonstrate the generation of polychromatic and collimated lights at 456 nm, 459 nm, and 761 nm based on cesium (133Cs) 6S1/2 - 6P3/2 - 8S1/2 - 7P3/2, 7P1/2, 6P1/2 - 6S1/2 multi-diamond-type atomic system via two-photon excitation with two IR pump lasers at 852 nm and 795 nm. The 456 nm, 459 nm (7P3/2, 7P1/2 → 6S1/2) collimated blue lights result from the self-seeded four-wave mixing process (FWM), and the 761 nm coherent light (8S1/2 → 6P1/2) is from a seeded FWM process with the injection of a third laser at 895 nm. We measure the dependency of generated polychromatic fields on the temperature of 133Cs vapor cell and the powers of input lasers, clearly demonstrating the competition between the self-seeded FWM and seeded FWM, as they share the same excitation path. This work is helpful to further produce entangled multi-color photons for quantum communication.

Molecular detection and genetic characteristics of porcine circovirus 3 and porcine circovirus 4 in central China.

Porcine circoviruses (PCVs) are a significant cause of concern for swine health, with four genotypes currently recognized. Two of these, PCV3 and PCV4, have been detected in pigs across all age groups, in both healthy and diseased animals. These viruses have been associated with various clinical manifestations, including porcine dermatitis and nephropathy syndrome (PDNS) and respiratory and enteric signs. In this study, we detected PCV3 and PCV4 in central China between January 2022 and February 2023. We tested fecal swabs and tissue samples from growing-finishing and suckling pigs with or without respiratory and systemic manifestations and found the prevalence of PCV3 to be 15.15% (15/99) and that of PCV3/PCV4 coinfection to be 4.04% (4/99). This relatively low prevalence might be attributed to the fact that most of the clinical samples were collected from pigs exhibiting respiratory signs, with only a few samples having been obtained from pigs with diarrhea. In some cases, PCV2 was also detected, and the coinfection rates of PCV2/3, PCV2/4, and PCV2/3/4 were 6.06% (6/99), 5.05% (5/99), and 3.03% (3/99), respectively. The complete genomic sequences of four PCV3 and two PCV4 isolates were determined. All four of the PCV3 isolates were of subtype PCV3b, and the two PCV4 isolates were of subtype PCV4b. Two mutations (A24V and R27K) were found in antibody recognition domains of PCV3, suggesting that they might be associated with immune escape. This study provides valuable insights into the molecular epidemiology and evolution of PCV3 and PCV4 that will be useful in future investigations of genotyping, immunogenicity, and immune evasion strategies.

Detection and genetic analysis of porcine circovirus-like virus in pigs with diarrhea between 2016 and 2021 in Henan and Shanxi provinces of China.

Porcine circovirus-like virus (PCLV) is a recently discovered virus that may be associated with diarrhea in pigs. To investigate the epidemic profile and genetic characteristics of this virus, 175 clinical samples (141 intestinal samples, 17 blood samples, and 17 fecal samples) were collected from diseased piglets during outbreaks of diarrhea from 33 pig farms in 19 cities of Henan and Shanxi provinces of China between 2016 and 2021 and were screened by PCR for the presence of PCLV. The results showed that the positive rate for PCLV was 32% (56/175) at the sample level, 60.6% (20/33) at the farm level, and 57.9% (11/19) at the city level, which varied from 5.88% to 44.12% between 2016 and 2021. It was also found that PCLV occurred in coinfections with porcine circovirus type 2 (PCV2), PCV3, PCV4, porcine epidemic diarrhea virus, and porcine reproductive and respiratory syndrome virus, but no nucleic acids were detected for transmissible gastroenteritis virus, porcine deltacoronavirus, or porcine rotavirus in piglets with diarrhea. Notably, PCLV was detected in 13 diarrheal piglets from four different farms that were negative for the other porcine viruses. These findings suggest that PCLV may be associated with porcine diarrhea and that it has been circulating in piglets in Henan and Shanxi provinces of China. In addition, the complete genomes of 13 PCLV strains were sequenced and found to share 35.4%-91.0% nucleotide sequence identity with sequences available in the GenBank database. Phylogenetic analysis based on Rep amino acid sequences revealed that the 13 PCLV strains from this study clustered in group 1 and were closely related to eight Chinese PCLV strains, Bo-Circo-like virus CH, American strains 21 and 22, and Hungarian strains 288_4 and 302_4, but they differed genetically from seven other foreign PCLV strains. The whole genome and rep gene of 13 PCLV strains in this study were 72.2%-82% and 83.8%-89.7% identical, respectively, to those of Bo-Circo-like virus strain CH, indicating that PCLV is a novel virus in pigs that may be involved in cross-species transmission. Evidence of a recombination event was found in the rep region of the 13 PCLV strains sequenced. This study enriches the epidemiological data on PCLV infection in pigs in China and lays a foundation for further study on the pathogenesis of PCLV.

Simultaneous detection and phylogenetic analysis of porcine epidemic diarrhea virus and porcine circovirus 4 in Henan province, China.

Porcine circovirus 4 (PCV4) is a recently discovered circovirus that was first reported in 2019 in several pigs in Hunan province of China and has also been identified in pigs infected with porcine epidemic diarrhea virus (PEDV). To further investigate the coinfection and genetic diversity of these two viruses, 65 clinical samples (including feces and intestinal tissues) were collected from diseased piglets on 19 large-scale pig farms in Henan province of China, and a duplex SYBR Green I-based quantitative real-time polymerase chain reaction (qPCR) assay was developed for detecting PEDV and PCV4 simultaneously. The results showed that the limit of detection was 55.2 copies/µL and 44.1 copies/µL for PEDV and PCV4, respectively. The detection rate for PEDV and PCV4 was 40% (26/65) and 38% (25/65), respectively, and the coinfection rate for the two viruses was 34% (22/65). Subsequently, the full-length spike (S) gene of eight PEDV strains and a portion of the genome containing the capsid (Cap) gene of three PCV4 strains were sequenced and analyzed. Phylogenetic analysis showed that all of the PEDV strains from the present study clustered in the G2a subgroup and were closely related to most of the PEDV reference strains from China from 2011 to 2021, but they differed genetically from a vaccine strain (CV777), a Korean strain (virulent DR1), and two Chinese strains (SD-M and LZC). It is noteworthy that two PEDV strains (HEXX-24 and HNXX-24XIA) were identified in one sample, and the HNXX-24XIA strain had a large deletion at amino acids 31-229 of the S protein. Moreover, a recombination event was observed in strain HEXX-24. Phylogenetic analysis based on the amino acid sequence of the PCV4 Cap protein revealed that PCV4 strains were divided into three genotypes: PCV4a1, PCV4a2, and PCV4b. Three strains in the present study belonged to PCV4a1, and they had a high degree of sequence similarity (>98% identity) to other PCV4 reference strains. This study not only provides technical support for field investigation of PEDV and PCV4 coinfection but also provides data for their prevention and control.

Functional interplay of histone lysine 2-hydroxyisobutyrylation and acetylation in Arabidopsis under dark-induced starvation.

Lysine 2-hydroxyisobutyrylation (Khib) is a novel type of histone acylation whose prevalence and function in plants remain unclear. Here, we identified 41 Khib sites on histones in Arabidopsis thaliana, which did not overlap with frequently modified N-tail lysines (e.g. H3K4, H3K9 and H4K8). Chromatin immunoprecipitation-sequencing (ChIP-seq) assays revealed histone Khib in 35% of protein-coding genes. Most Khib peaks were located in genic regions, and they were highly enriched at the transcription start sites. Histone Khib is highly correlated with acetylation (ac), particularly H3K23ac, which it largely resembles in its genomic and genic distribution. Notably, co-enrichment of histone Khib and H3K23ac correlates with high gene expression levels. Metabolic profiling, transcriptome analyses, and ChIP-qPCR revealed that histone Khib and H3K23ac are co-enriched on genes involved in starch and sucrose metabolism, pentose and glucuronate interconversions, and phenylpropanoid biosynthesis, and help fine-tune plant response to dark-induced starvation. These findings suggest that Khib and H3K23ac may act in concert to promote high levels of gene transcription and regulate cellular metabolism to facilitate plant adaption to stress. Finally, HDA6 and HDA9 are involved in removing histone Khib. Our findings reveal Khib as a conserved yet unique plant histone mark acting with lysine acetylation in transcription-associated epigenomic processes.

Development and application of a low-priced duplex quantitative PCR assay based on SYBR Green I for the simultaneous detection of porcine deltacoronavirus and porcine sapelovirus.

Porcine deltacoronavirus (PDCoV) and porcine sapelovirus (PSV) are two viruses that can cause diarrhoea in pigs and bring great economic loss to the pig industry. In this research, a duplex real-time quantitative polymerase chain reaction (qPCR) assay based on SYBR Green I was developed to simultaneously detect PDCoV and PSV. No specific melting peaks were found in other porcine diarrhoea-associated viruses, indicating that the method developed in this study had good specificity. The detection limits of PDCoV and PSV were 1.0 × 101 copies µl-1 and 1.0 × 102 copies µl-1, respectively. The duplex real-time qPCR assay tested two hundred and three (203) intestinal and faecal samples collected from diarrhoeal and asymptomatic pigs. The positive rates of PDCoV and PSV were 20.2% and 23.2%, respectively. The co-infection rate of PDCoV and PSV was 13.8%. To evaluate the accuracy of the developed method, conventional PCR and singular TaqMan real-time qPCR assays for PDCoV/PSV were also used to detect the samples. The results showed that the duplex real-time qPCR assay was consistent with the singular assays, but its sensitivity was higher than conventional PCR methods. This duplex real-time qPCR assay provides a rapid, sensitive and reliable method in a clinic to simultaneously detect PDCoV and PSV.

Combining hydrochemistry and hydrogen and oxygen stable isotopes to reveal the influence of human activities on surface water quality in Chaohu Lake Basin.

The input of pollutants caused by human activities induces the deterioration of surface water quality. To reveal the characteristics of surface water quality in Chaohu Lake Basin and the influence of human activities, the hydrochemistry and stable isotope composition of hydrogen and oxygen in lake water and inflow river water were analyzed. The results show that the hydrochemical type of lake water is the Na-Cl type,while river water is the Na-Cl, Ca-Cl and mixed types. The ion proportional coefficient method and principal component analysis show that surface water is controlled by weathering of evaporated salt rocks and silicate rocks, in which Cl- and SO42- are affected by fertilizers and sewage to some extent. There is a strong correlation between conventional ions and nutrient indexes, which indicates that dissolved ions are affected not only by rock weathering but also by human activities (such as the discharge of domestic sewage or nitrogen-containing wastewater and the use of fertilizers). The stable isotope values of hydrogen and oxygen in surface water are distributed at the lower right portion of the local precipitation line and are close to it, indicating that surface water mainly originates from precipitation. The high value of d-excess values in surface water indicates that evaporation is weak. As pollution indicators, EC, Cl- and NO3- indicates that the Nanfei River, Dianbu River, Shiwuli River and Pai River flow in northwestern of Chaohu Lake Basin through Hefei urban city are severely polluted, NO3- originates from manure and sewage. Rivers flowing through farmland areas are less polluted, and the use of agricultural fertilizer contributes greatly to NO3-.

Mutations in ASH1L confer susceptibility to Tourette syndrome.

Tourette syndrome (TS) is a childhood-onset neuropsychiatric disorder characterized by repetitive motor movements and vocal tics. The clinical manifestations of TS are complex and often overlap with other neuropsychiatric disorders. TS is highly heritable; however, the underlying genetic basis and molecular and neuronal mechanisms of TS remain largely unknown. We performed whole-exome sequencing of a hundred trios (probands and their parents) with detailed records of their clinical presentations and identified a risk gene, ASH1L, that was both de novo mutated and associated with TS based on a transmission disequilibrium test. As a replication, we performed follow-up targeted sequencing of ASH1L in additional 524 unrelated TS samples and replicated the association (P value = 0.001). The point mutations in ASH1L cause defects in its enzymatic activity. Therefore, we established a transgenic mouse line and performed an array of anatomical, behavioral, and functional assays to investigate ASH1L function. The Ash1l+/- mice manifested tic-like behaviors and compulsive behaviors that could be rescued by the tic-relieving drug haloperidol. We also found that Ash1l disruption leads to hyper-activation and elevated dopamine-releasing events in the dorsal striatum, all of which could explain the neural mechanisms for the behavioral abnormalities in mice. Taken together, our results provide compelling evidence that ASH1L is a TS risk gene.

Benefits of Iron Chelators in the Treatment of Parkinson's Disease.

As a novel discovered regulated cell death pattern, ferroptosis has been associated with the development of Parkinson's disease (PD) and has attracted widespread attention. Nevertheless, the relationship between ferroptosis and PD pathogenesis is still unclear. This study aims to investigate the effect of iron overload on dopaminergic (DA) neurons and its correlation with ferroptosis. Here we use nerve growth factor (NGF) induced PC12 cells which are derived from pheochromocytoma of the rat adrenal to establish a classical PD in vitro model. We found significantly decreased cell viability in NGF-PC12 cell under ammonium ferric citrate (FAC) administration. Moreover, excessive intracellular iron ions induced the increase of (reactive oxygen species) ROS release as well as the decrease of mitochondrial membrane potential in PC12-NGF cells. In addition, we also found that overloaded iron can activate cell apoptosis and ferroptosis pathways, which led to cell death. Furthermore, MPP-induced PD cells were characterized by mitochondrial shrinkage, decreased expression of glutathione peroxidase 4 (Gpx4) and ferritin heavy chain (FTH1), and increased divalent metal transporter (DMT1) and transferrin receptor 1 (TfR1) expression level. In contrast, Lip-1 and DFO increased the expression level of GPX4 and FTH1 compared to MPP-induced PD cell. In conclusion, we indicated that overloaded intracellular iron contributes to neurons death via apoptosis and ferroptosis pathways, while DFO, an iron chelator, can inhibit ferroptosis in order to protect the neurons in vitro.

Detection and genetic characteristics of porcine bocavirus in central China.

To investigate the epidemic profile and genetic diversity of porcine bocavirus (PBoV), 281 clinical samples, including 236 intestinal tissue samples and 45 fecal samples were collected from diarrheic piglets on 37 different pig farms in central China, and two SYBR Green I-based quantitative PCR assays were developed to detect PBoV1/2 and PBoV3/4/5, respectively. One hundred forty-eight (52.67%) of the 281 clinical samples were positive for PBoV1/2, 117 (41.63%) were positive for PBoV3/4/5, 55 (19.57%) were positive for both PBoV1/2 and PBoV3/4/5, and 86.49% (32/37) of the pig farms were positive for PBoV. Overall, the prevalence of PBoV was 74.73% (210/281) in central China. Subsequently, nearly full-length genomic sequences of two PBoV strains (designated CH/HNZM and PBoV-TY) from two different farms were determined. Phylogenetic analysis demonstrated that the two PBoV strains obtained in this study belonged to the PBoV G2 group and had a close relationship to 10 other PBoV G2 strains but differed genetically from PBoV G1, PBoV G3, and seven other bocaviruses. CH/HNZM and PBoV-TY were closely related to the PBoV strain GD18 (KJ755666), which may be derived from the PBoV strains 0912/2012 (MH558677) and 57AT-HU (KF206160) through recombination. Compared with reference strain ZJD (HM053694)-China, more amino acid variation was found in the NS1 proteins of CH/HNZM and PBoV-TY. These data extend our understanding of the molecular epidemiology and evolution of PBoV.

A novel model to predict mental distress among medical graduate students in China.

BACKGROUND: Poor mental health was reported among medical graduate students in some studies. Identification of risk factors for predicting the mental health is capable of reducing psychological distress among medical graduate students. Therefore, the aim of the study was to identify potential risk factors relating to mental health and further create a novel prediction model to calculate the risk of mental distress among medical graduate students. METHODS: This study collected and analyzed 1079 medical graduate students via an online questionnaire. Included participants were randomly classified into a training group and a validation group. A model was developed in the training group and validation of the model was performed in the validation group. The predictive performance of the model was assessed using the discrimination and calibration. RESULTS: One thousand and fifteen participants were enrolled and then randomly divided into the training group (n = 508) and the validation group (n = 507). The prevalence of severe mental distress was 14.96% in the training group, and 16.77% in the validation group. The model was developed using the six variables, including the year of study, type of student, daily research time, monthly income, scientific learning style, and feeling of time stress. The area under the receiver operating characteristic curve (AUROC) and calibration slope for the model were 0.70 and 0.90 (95% CI: 0.65 ~ 1.15) in the training group, respectively, and 0.66 and 0.80 (95% CI, 0.51 ~ 1.09) in the validation group, respectively. CONCLUSIONS: The study identified six risk factors for predicting anxiety and depression and successfully created a prediction model. The model may be a useful tool that can identify the mental status among medical graduate students. TRIAL REGISTRATION: No. ChiCTR2000039574 , prospectively registered on 1 November 2020.

Assessments of different inactivating reagents in formulating transmissible gastroenteritis virus vaccine.

BACKGROUND: Transmissible gastroenteritis virus (TGEV) causes enteric infection in piglets, characterized by vomiting, severe diarrhea and dehydration, and the mortality in suckling piglets is often high up to 100%. Vaccination is an effective measure to control the disease caused by TGEV. METHODS: In this study, cell-cultured TGEV HN-2012 strain was inactivated by formaldehyde (FA), ß-propiolactone (BPL) or binaryethylenimine (BEI), respectively. Then the inactivated TGEV vaccine was prepared with freund's adjuvant, and the immunization effects were evaluated in mice. The TGEV-specific IgG level was detected by ELISA. The positive rates of CD4+, CD8+, CD4+IFN-γ+, CD4+IL-4+ T lymphocytes were detected by flow cytometry assay. Lymphocyte proliferation assay and gross pathology and histopathology examination were also performed to assess the three different inactivating reagents in formulating TGEV vaccine. RESULTS: The results showed that the TGEV-specific IgG level in FA group (n = 17) was earlier and stronger, while the BEI group produced much longer-term IgG level. The lymphocyte proliferation test demonstrated that the BEI group had a stronger ability to induce spleen lymphocyte proliferation. The positive rates of CD4+ and CD8+ T lymphocyte subsets of peripheral blood lymphocyte in BEI group was higher than that in FA group and BPL groups by flow cytometry assay. The positive rate of CD4+IFN-γ+ T lymphocyte subset was the highest in the BPL group, and the positive rate of CD4+IL-4+ T lymphocyte subset was the highest in the FA group. There were no obvious pathological changes in the vaccinated mice and the control group after the macroscopic and histopathological examination. CONCLUSIONS: These results indicated that all the three experimental groups could induce cellular and humoral immunity, and the FA group had the best humoral immunity effect, while the BEI group showed its excellent cellular immunity effect.

Characterization of a recombinant pseudorabies virus expressing porcine parvovirus VP2 protein and porcine IL-6.

BACKGROUND: Porcine parvovirus (PPV) and pseudorabies virus (PRV) are the important etiological agents of swine infectious diseases, resulting in huge economic losses to the Chinese swine industry. Interleukin-6 (IL-6) has the roles to support host immune response to infections as a pleiotropic cytokine. It is essential to construct a live attenuated vaccine-based recombinant PRV that expresses PPV VP2 protein and porcine IL-6 for prevention and control of PRV and PPV. METHODS: The recombinant plasmid, pGVP2-IL6, was constructed by porcine IL-6 gene substituting for EGFP gene of the PRV transfer plasmid pGVP2-EGFP containing VP2 gene of PPV. Plasmid pGVP2-IL6 was transfected into swine testicle cells pre-infected with the virus rPRV-VP2-EGFP strain through homologous recombination and plaque purification to generate a recombinant virus rPRV-VP2-IL6. The recombinant PRV was further identified by PCR and DNA sequencing, and the expression of the VP2 protein and porcine IL-6 was analyzed by reverse transcription-PCR (RT-PCR) and Western blot. The virus titer was calculated according to Reed and Muench method. The immunogenicity of the recombinant virus was preliminarily evaluated in mice by intramuscular administration twice with the rPRV-VP2-IL6 at 4-week intervals. RESULTS: A recombinant virus rPRV-VP2-IL6 was successfully constructed and confirmed in this study. The properties of rPRV-VP2-IL6 were similar to the parental virus HB98 in terms of growth curve, morphogenesis and virus plaque sizes, and rPRV-VP2-IL6 was proliferated in different cell types. It induced specific antibodies against PPV as well as a strong increase of PPV-specific lymphocyte proliferation responses in mice immunized with rPRV-VP2-IL6, and provided partial protection against the virulent PPV challenge. rPRV-VP2-IL6 also induced a high level of neutralizing antibodies against PRV, and significantly reduced the mortality rate of (1 of 10) following virulent PRV challenge compared with the control (10 of 10). CONCLUSIONS: The recombinant rPRV-VP2-IL6 might be a potential candidate vaccine against PRV and PPV infections in pigs.

Development of a duplex SYBR Greenâ based real-time PCR assay for detection of porcine epidemic diarrhea virus and porcine bocavirus3/4/5.

The duplex real-time PCR assay based on SYBR Green І was developed for detection of porcine epidemic diarrhea virus (PEDV) and porcine bocavirus (PBoV) 3/4/5 genotypes simultaneously. Two pairs of specific primers were designed targeting the N gene sequence of PEDV and VP1 gene sequence of PBoV3/4/5. PEDV and PBoV3/4/5 could be distinguished by their different melting temperatures (Tm) in one sample. The Tm value of PEDV was 83.5 °C, and the Tm value of PBoV3/4/5 was 78.5 °C, while other swine pathogens showed no specific melting peaks. The detection limits of this assay were 10 copies/µL for both PEDV and PBoV3/4/5. A total of sixty-three intestinal tissue samples were collected from piglets suffering from diarrhea, and the viral nucleic acids detected and identified by the real-time PCR assay and conventional PCR assay. The duplex real-time PCR detection results showed that the prevalence of PEDV and PBoV3/4/5 was 85.7% and 46%, respectively, and the co-infection rate of the two viruses was 28.6%. These results indicated that this duplex real-time PCR assay was a sensitive, specific and reproducible method for differentiating PEDV and PBoV3/4/5 or their co-infection.

Construction and immunogenicity of a gE/gI/TK-deleted PRV based on porcine pseudorabies virus variant.

Pseudorabies (PR) caused by re-emerging pseudorabies virus (PRV) variant has outbroken among PRV vaccine-immunized swine herds on many Chinese pig farms, with severe socioeconomic consequences since late 2011. Here, a gE/gI/TK-deleted recombinant virus (rPRV NY-gE-/gI-/TK-) was constructed based on PRV NY strain from 2012 through homologous DNA recombination and gene-editing technology termed clustered regularly interspaced palindromic repeats (CRISPR)/associated (Cas9) system. The rPRV NY-gE-/gI-/TK- strain showed similar growth kinetics to the parental PRV NY strain in vitro, and was safe for mice. Sixty mice were injected subcutaneously (s.c.) twice with 106.0 TCID50 of rPRV NY-gE-/gI-/TK- and DMEM, respectively, with two-week interval. The levels of PRV gB antibodies and neutralizing antibodies against PRV NY in mice immunized with rPRV NY-gE-/gI-/TK- were higher than those in the DMEM control group. The number of T lymphocyte subclasses CD3+, CD4+ and CD8+ in rPRV NY-gE-/gI-/TK--immunized mice was higher than that in DMEM-injected mice. After challenge with 106.0 TCID50 PRV NY at 42 dpi, all rPRV NY-gE-/gI-/TK--immunized mice survived without exhibiting any pathological lesions in different tissues and intranuclear eosinophilic inclusions of the brain, and the viral genomic copy numbers in various organs of mice were obviously lower than DMEM group. These results showed the rPRV NY-gE-/gI-/TK- could be a promising next-generation vaccine to control now epidemic PR in China.

Simultaneous detection of porcine reproductive and respiratory syndrome virus and porcine circovirus 3 by SYBR Green І-based duplex real-time PCR.

The SYBR Green І-based duplex real-time PCR assay was developed for simultaneous detection of porcine reproductive and respiratory syndrome virus (PRRSV) and porcine circovirus 3 (PCV-3) genomes. PRRSV and PCV-3 were distinguished in the same sample by their distinctive melting temperature (Tm) which was 84 °C for PRRSV and 81.5 °C for PCV-3, and other non-targeted swine viruses showed no specific melting peaks. The detection limits of this assay were 46.1copies/µL for PRRSV and 49.3copies/µL for PCV-3, respectively. Thirty-three lung samples of porcine with respiratory and reproductive failure symptoms were collected and confirmed by the SYBR Green І-based real-time PCR assay and conventional PCR assay. The real-time PCR detection results showed that the PRRSV positive rate was 45.45%, the PCV-3 positive rate was 63.63%, the PRRSV and PCV-3 co-infection positive rate was 36.36%, which were more sensitive than conventional PCR detection. This duplex real-time PCR assay could be a rapid, sensitive and reliable method for the detection of PRRSV and PCV-3 co-infection.

Characteristics of the spike and ORF3 genes of porcine epidemic diarrhea virus in Henan and Shanxi provinces of China.

To investigate the epidemic characteristics of porcine epidemic diarrhea virus (PEDV), 135 clinical samples (including intestinal tissues and feces) were collected from diseased piglets during outbreaks of diarrhea from 2015 to 2019 on farms in Henan and Shanxi provinces of China where swine had been immunized with attenuated PEDV (CV777). A total of 86 clinical samples (86/135, 63.7%) were positive for PEDV by RT-PCR, and subsequently, the complete spike (S) and ORF3 genes of 32 PEDV samples were sequenced. Phylogenetic analysis showed that the 32 PEDV strains obtained in this study belonged to group 2 (pandemic variant strains) and had a close relationship to 17 Chinese strains after 2010, two South Korean strains (KNU-1305 and KNU-1807), three American strains (PC22A-P140.BI, USA/Colorado/2013, and USA/OK10240-6/2017) and a Mexican strain (PEDV/MEX/QRO/02/2017), but differed genetically from a South Korean strain (SM98), a European strain (Br1/87), a Chinese strain (LZC), and a vaccine strain (CV777). G2-a subgroup strains were the dominant pandemic variant strains circulating in Henan and Shanxi provinces of China. Furthermore, a cross-recombination event was identified in the S region of the SX/TY2/2017 strain, and the putative parental strains were the epidemic strains CH/GDGZ/2012 and CH/YZ1/2015, identified in China in 2012 and 2015, respectively. These results provide further information about PEDV evolution, which could improve our understanding of the circulation of PEDV in Henan and Shanxi provinces. This information will also be helpful for developing new strategies for prevention and control of variant strains.

Quick selenium accumulation in the selenium-rich rice and its physiological responses in changing selenium environments.

BACKGROUND: The element selenium (Se) deficiency is thought to be a global human health problem, which could disperse by daily-supplement from Se-rich food. Increasing the accumulation of Se in rice grain is an approach matched to these nutrient demands. Nonetheless, Se is shown to be essential but also toxic to plants, with a narrow margin between deficiency and toxicity. Notably, the regulatory mechanism balancing the accumulation and tolerance of Se in Se-rich rice plants remains unknown. RESULTS: In this study, we investigated the phenotypical, physiological, and biochemical alterations of Se-rich rice in the exposure to a variety of Se applications. Results showed that the Se-rich rice was able to accumulate more abundance of Se from the root under a low Se environment comparing to the Se-free rice. Besides, excessive Se led to phytotoxic effects on Se-rich rice plants by inducing chlorosis and dwarfness, decreasing the contents of antioxidant, and exacerbating oxidative stresses. Furthermore, both phosphate transporter OsPT2 and sulfate transporters OsSultr1;2 may contribute to the uptake of selenate in rice. CONCLUSIONS: Se-rich red rice is more sensitive to exogenous application of Se, while and the most effective application of Se in roots of Se-rich rice was reached in 20 µM. Our findings present a direct way to evaluate the toxic effects of Se-rich rice in the Se contaminated field. Conclusively, some long-term field trial strategies are suggested to be included in the evaluation of risks and benefits within various field managements.

Development of a TB green II-based duplex real-time fluorescence quantitative PCR assay for the simultaneous detection of porcine circovirus 2 and 3.

Porcine circovirus 3 (PCV3), as a newly emerged circovirus, is widely distributed in pig populations worldwide. Co-infection of PCV2 and PCV3 has been reported frequently in clinical samples. In the present study, a TB Green II-based duplex real-time polymerase chain reaction (qPCR) was developed to rapidly and differentially detect PCV2 and PCV3. The assay specifically detected PCV2 and PCV3, with no fluorescence signals being detected for other non-targeted pig pathogens. The duplex qPCR showed a high degree of linearity (R2 > 0.998), and its limits of detection were 10 and 78 copies/µL for PCV2 and PCV3, respectively. The duplex qPCR could detect and differentiate PCV2 (melting peaks at 85.5 °C) and PCV3 (melting peaks at 82.5 °C), and showed high repeatability and reproducibility, with intra- and inter-assay coefficients of variation of less than 2.0%. Fifty-six tissue samples from 18 pig farms were used to evaluate the duplex qPCR method. The results revealed infection rates of 66.07% (37/56) and 39.28% (22/56) for PCV2 and PCV3, respectively. The PCV2 + PCV3 co-infection rate was 39.28% (22/56). The developed method could be used as an efficient molecular biology tool for epidemiological investigations of PCV2 and PCV3.

Analysis of genetic variation of porcine circovirus type 2 within pig populations in central China.

In order to investigate the genetic diversity of porcine circovirus type 2 (PCV2), 284 clinical tissue samples were collected from different pig farms in central China from 2015 to 2017. A total of 162 tissue samples (162/284, 57.04%) were positive for PCV2 by PCR, and subsequently, the complete genome of 36 of these PCV2 samples was cloned and sequenced. The sequencing results showed that 37 complete PCV2 sequences were obtained from 36 PCV2-positive clinical samples. These PCV2 strains were relatively conserved and extremely homologous to the representative classical PCV2 strains. Of these, 20 PCV2 strains belonged to genotype PCV2d, 14 belonged to PCV2b, and three others belonged to PCV2a. Coinfection with PCV2b and PCV2d was identified in one sample (DF-2). These results show that PCV2d may be gradually replacing PCV2b as the predominant PCV2 genotype in central China, and that other genotypes also exist in individual regions. The results of this study will aid in our understanding of the molecular epidemiology of PCV2.