Ribosomal protein L32 contributes to the growth, antibiotic resistance and virulence of Glaesserella parasuis.

Glaesserella parasuis is the pathogen that causes Glässer's disease in pigs, which is characterized by fibrinous polyserositis, arthritis and meningitis. Research on ribosomal protein L32 in microorganisms has mainly focused on regulating gene transcription and translation, but its effect on bacterial virulence is unclear. The role of L32 gene in G. parasuis is not clear, and in order to study the function of L32 gene, a suicide plasmid-mediated natural transformation method was used to construct a L32 gene deletion mutant. We found that although L32 was shown to be non-essential for cell proliferation, the growth curve of ΔL32 is clearly different compared with that of ZJ1208. ΔL32 produced more outer membrane vesicles (OMVs) with a variety of irregular shapes, but produced similar biofilm to the parental strain. ΔL32 is more sensitive to osmotic pressure, oxidation pressure and heat shock stress. Meanwhile, ΔL32 is significantly more susceptible to antimicrobials such as spectinomycin, apramycin, sulfafurazole, but not to other antibiotics used in this study. In the mouse challenge experiment, the mortality of mice infected with the mutant strain decreased by 40% compared to those infected with the wild-type strain, indicating that L32 is a virulence-associated factor which contributes to bacterial fitness in host environments. The above results show that L32 is important for the growth, stress resistance and virulence of G. parasuis, and this study also confirms for the first time that L32 plays an important role in antibiotic resistance against aminoglycosides and sulfonamides.

Association between strenuous sports or other exercises and lung cancer risk: a mendelian randomization study.

Background: Studying the relationship between strenuous sports or other exercises (SSOE) and lung cancer risk remains underexplored. Traditional observational studies face challenges like confounders and inverse causation. However, Mendelian randomization (MR) provides a promising approach in epidemiology and genetics, using genetic variants as instrumental variables to investigate causal relationships. By leveraging MR, we have scrutinized the causal link between SSOE and lung cancer development. Methods: Twelve single-nucleotide polymorphisms (SNPs) associated with SSOE, as identified in previously published genome-wide association studies, were utilized as instrumental variables in our investigation. Summary genetic data at the individual level were obtained from relevant studies and cancer consortia. The study encompassed a total of 11,348 cases and 15,861 controls. The statistical technique of inverse variance-weighting (IVW), commonly employed in meta-analyses and MR studies, was employed to assess the causal relationship between SSOE and lung cancer risk. Results: The MR risk analysis indicated a causal relationship between SSOE and the incidence of lung cancer, with evidence of a reduced risk for overall lung cancer [odds ratio (OR) =0.129; 95% confidence interval (CI): 0.021-0.779; P=0.03], lung adenocarcinoma (OR =0.161; 95% CI: 0.012-2.102; P=0.16) and squamous cell lung cancer (OR =0.045; 95% CI: 0.003-0.677; P=0.03). The combined OR for lung cancer from SSOE (controlling for waist circumference and smoking status) was 0.054 (95% CI: 0.010-0.302, P<0.001). Conclusions: Our MR analysis findings indicate a potential correlation between SSOE and a protective effect against lung cancer development. Further investigation is imperative to uncover the precise mechanistic link between them.

Occurrence Mechanism of the Abnormal Gelation Phenomenon of High Temperature Cementing Slurry Induced by a Polycarboxylic Retarder.

The class G oil well cement is a type of special cement that can be subjected to a high temperature formation environment. It was found that the class G cement tail slurry with a low polycarboxylic retarder dosage (usually ≤1% by weight of cement) was more prone to cause the abnormal gelation phenomenon (AGP) than the lead slurry with a high retarder dosage at a high temperature (usually when T ≥ 120 °C). This study aimed at the occurrence mechanism of this unfavorable phenomenon that seriously endangers the cementing security. Results showed that the abnormal gelatinous region underwent premature hydration; namely, the calcium hydroxide and calcium silicate hydrate (C-S-H) content were all higher than the nongelatinous region, while the copolymer content was the opposite. Correspondingly, the theory of "premature hydration and crystal nucleation" was proposed to explain the abnormal gelation mechanism of a cementing tail slurry with an insufficient retarder dosage. Furthermore, a novel functionalized copolymer retarder "PAIANS" was synthesized to alleviate the AGP.

Assisted reproductive technology induces different secondary sex ratio: parental and embryonic impacts.

BACKGROUND: Assisted reproduction technology (ART) has advanced significantly, raising concerns regarding its impact on the secondary sex ratio (SSR), which is the sex ratio at birth in offspring. This study aimed to explore factors affecting SSR in singletons, singletons from twin gestation, and twins from twin gestation within the context of ART. METHODS: A retrospective analysis was conducted on data from 8335 births involving 6,223 couples undergoing ART. Binary logistic regression assessed relationships between parental and embryonic factors and SSR in singletons and singletons from twin gestation. Multinomial logistic regression models were utilized to identify factors influencing SSR in twins from twin gestation. RESULTS: Secondary infertility (OR = 1.164, 95% CI: 1.009-1.342), advanced paternal age (OR = 1.261, 95% CI: 1.038-1.534), and blastocyst embryo transfer (OR = 1.339, 95% CI: 1.030-1.742) were associated with an increased SSR, while frozen embryo transfer (FET) showed a negative association with SSR (OR = 0.738, 95% CI: 0.597-0.912) in singletons. A longer duration of gonadotropin (Gn) usage reduced SSR in singletons (OR = 0.961, 95% CI: 0.932-0.990) and singletons from twin gestation (OR = 0.906, 95% CI: 0.838-0.980). In singletons from twin gestation, male-induced infertility (OR = 2.208, 95% CI: 1.120-4.348) and higher Gn dosage (OR = 1.250, 95% CI: 1.010-1.548) were significantly associated with an increased SSR. Women aged > 35 years and intracytoplasmic sperm injection (ICSI) were associated with lower SSR (OR = 0.539, 95% CI: 0.293-0.990 and OR = 0.331, 95% CI: 0.158-0.690, respectively). In twins from twin gestation, paternal age exceeded maternal age (OR = 0.682, 95% CI: 0.492-0.945) and higher Gn dosage (OR = 0.837, 95% CI: 0.715-0.980) were associated with a higher proportion of male twins. Cleavage stage transfer (OR = 1.754, 95% CI: 1.133-2.716) resulted in a higher percentage of boy-girl twins compared to blastocyst transfer. CONCLUSION: This study demonstrates the complex interplay of various factors in determining the SSR in ART, highlighting the importance of considering infertility type, paternal age, fertilization method, embryo transfer stage, and Gn use duration when assessing SSR. Nevertheless, further research with a large sample size is necessary to confirm and expand upon the findings of this study.

Enhanced in vivo antiviral activity against pseudorabies virus through transforming gallic acid into graphene quantum dots with stimulation of interferon-related immune responses.

With the urgent need for antiviral agents, antiviral materials with high biocompatibility and antiviral effects have attracted a lot of attention. In this study, gallic acid, a natural polyphenolic compound, was transformed into biocompatible graphene quantum dots (GAGQDs) which exhibit enhanced antiviral activity against pseudorabies virus (PRV). The as-prepared GAGQDs inhibit PRV proliferation with a 104-fold reduction in viral titers. Investigation of the antiviral mechanism revealed that GAGQDs inhibit the adsorption, invasion and replication of PRV infection. Treatment with GAGQDs regulates the expression levels of interferon-related antiviral proteins, including mitochondrial antiviral-signaling protein (MAVS), signal transducer and activator of transcription 1 (STAT1) and 2',5'-oligoadenylate synthetase 1 (OAS1), suggesting that GAGQDs can stimulate innate antiviral immune responses, resulting in enhanced antiviral effects. More importantly, GAGQD treatments alleviate clinical symptoms and reduce mortality in PRV-infected mice. Our results reveal the enhanced therapeutic effects of GAGQDs against PRV infection in vitro and in vivo, suggesting the potential of GAGQDs as a promising novel antiviral agent.

Integrative transcriptomic and metabolomic analysis in mice reveals the mechanism by which ginseng stem-leaf saponins enhance mucosal immunity induced by a porcine epidemic diarrhea virus vaccination.

Porcine epidemic diarrhea virus (PEDV) is a main cause of severe enteric disease in piglets, leading to millions of dollars lost annually in the global pig industry. Parenteral vaccination is limited in generating sufficient mucosal immunity, which is crucial for early defense against PEDV. Here, we orally administered ginseng stem-leaf saponins (GSLS) to mice before parenteral vaccination and found that GSLS significantly enhanced the phagocytosis of dendritic cells, promoted the activities of CD4+ T cells and increased PEDV-specific IgA antibodies in the intestinal mucosa. Transcriptomic results showed that the altered genes following GSLS treatment were mostly related to the immune response and metabolism. In addition, integrated analysis of the transcriptome and metabolome revealed that the mechanism by which GSLS enhances mucosal immunity may be associated with progesterone-related pathways. Further studies are needed to explore the detailed molecular mechanisms.

Isolation, Identification and Drug Resistance Rates of Bacteria from Pigs in Zhejiang and Surrounding Areas during 2019-2021.

This study aimed to determine the prevalence of bacterial diseases in pig farms in various regions of Zhejiang Province and surrounding areas. A total of 526 samples were collected from 85 pig farms in Zhejiang Province and surrounding areas. In this study, samples were analyzed using bacterial isolation and purification, Gram staining, PCR amplification, and antimicrobial susceptibility testing. A total of 36 Pasteurella multocida (Pm) isolates were detected, with an isolation rate of 6.84%; 37 Bordetella bronchiseptica (Bb) isolates were detected, with an isolation rate of 7.03%; 60 Glasserella parasuis (G. parasuis) isolates were detected, with an isolation rate of 11.41%; 170 Escherichia coli (E. coli) isolates were detected, with an isolation rate of 32.32%; 67 Streptococcus suis (SS) isolates were detected, with an isolation rate of 12.74%; 44 Actinobacillus pleuropneumoniae (APP) isolates were detected, with an isolation rate of 8.37%; and 7 Salmonella enteritis (SE) isolates were detected, with an isolation rate of 1.33%. Antimicrobial drug susceptibility testing against 21 types of antibiotics was carried out on the isolated strains, and the results showed that 228 strains had varying degrees of resistance to 21 antibiotics, including Pm, Bb, E. coli, and APP, with the highest resistance to lincomycin, at 100%. Pm and APP were the most sensitive to cephalothin, with resistance rates of 0. In terms of strains, Pm had the highest overall sensitivity to 21 antibiotics, and E. coli had the highest resistance. In short, bacterial diseases in Zhejiang and the surrounding areas were harmful, and the drug resistance situation was severe. This study provides scientific guidance for the clinical treatment of bacterial diseases.

Sequential application of copy number variation sequencing and quantitative fluorescence polymerase chain reaction in genetic analysis of miscarriage and stillbirth.

BACKGROUND: Copy number variation sequencing (CNV-seq) could detect most chromosomal abnormalities except polyploidy, and quantitative fluorescence polymerase chain reaction (QF-PCR) is a supplementary method to CNV-seq in triploid detection. This study aimed to evaluate the feasibility of sequential application of CNV-seq and QF-PCR in genetic analysis of miscarriage and stillbirth. METHODS: A total of 261 fetal specimens were analyzed by CNV-seq, and QF-PCR was only further performed for samples with normal female karyotype identified by CNV-seq. Cost and turnaround time (TAT) was analyzed for sequential detection strategy. Subgroup analysis and logistic regression were carried out to evaluate the relationship between clinical characteristics (maternal age, gestational age, and number of pregnancy losses) and the occurrence of chromosomal abnormalities. RESULTS: Abnormal results were obtained in 120 of 261 (45.98%) cases. Aneuploidy was the most common abnormality (37.55%), followed by triploidy (4.98%) and pathogenic copy number variations (pCNVs) (3.45%). CNV-seq could detect the triploidy with male karyotype, and QF-PCR could further identify the remaining triploidy with female karyotype. In this study, we found more male triploidies than female triploidies. With the same ability in chromosomal abnormalities detection, the cost of sequential strategy decreased by 17.35% compared with combined strategy. In subgroup analysis, significant difference was found in the frequency of total chromosomal abnormalities between early abortion group and late abortion group. Results of logistic regression showed a trend that pregnant women with advanced age, first-time abortion, and abortion earlier than 12 weeks were more likely to detect chromosomal aberrations in their products of conception. CONCLUSION: Sequential application of CNV-seq and QF-PCR is an economic and practical strategy to identify chromosomal abnormalities in fetal tissue.

Early Oral Administration of Ginseng Stem-Leaf Saponins Enhances the Peyer's Patch-Dependent Maternal IgA Antibody Response to a PEDV Inactivated Vaccine in Mice, with Gut Microbiota Involvement.

Neonatal piglets during the first week of life are highly susceptible to porcine epidemic diarrhoea virus (PEDV) infection, with mortality rates reaching 80-100%. Passive lactogenic immunity remains the most effective way to protect neonates from infection. Although safe, inactivated vaccines provide little or no passive protection. Here, we administered ginseng stem-leaf saponins (GSLS) to mice before parenteral immunization with an inactivated PEDV vaccine to investigate the effect of GSLS on the gut-mammary gland (MG)-secretory IgA axis. Early oral GSLS administration potently increased PEDV-specific IgA plasma cell generation in the intestine, facilitated intestinal IgA plasma cell migration to the MG by enhancing the chemokine receptor (CCR)10-chemokine ligand (CCL)28 interaction, and ultimately promoted specific IgA secretion into milk, which was dependent on Peyer's patches (PPs). Additionally, GSLS improved the gut microbiota composition, especially increasing probiotic abundance, and these microflora members promoted the GSLS-enhanced gut-MG-secretory IgA axis response and were regulated by PPs. In summary, our findings highlight the potential of GSLS as an oral adjuvant for PEDV-inactivated vaccines and provide an attractive vaccination strategy for lactogenic immunity induction in sows. Further studies are required to evaluate the mucosal immune enhancement efficacy of GSLS in pigs.

The role of DNA methylation in the maintenance of phenotypic variation induced by grafting chimerism in Brassica.

Grafting facilitates the interaction between heterologous cells with different genomes, resulting in abundant phenotypic variation, which provides opportunities for crop improvement. However, how grafting-induced variation occurs and is transmitted to progeny remains elusive. A graft chimera, especially a periclinal chimera, which has genetically distinct cell layers throughout the plant, is an excellent model to probe the molecular mechanisms of grafting-induced variation maintenance. Here we regenerated a plant from the T-cell layer of a periclinal chimera, TCC (where the apical meristem was artificially divided into three cell layers - from outside to inside, L1, L2, and L3; T = Tuber mustard, C = red Cabbage), named rTTT0 (r = regenerated). Compared with the control (rsTTT, s = self-grafted), rTTT0 had multiple phenotypic variations, especially leaf shape variation, which could be maintained in sexual progeny. Transcriptomes were analyzed and 58 phenotypic variation-associated genes were identified. Whole-genome bisulfite sequencing analyses revealed that the methylome of rTTT0 was changed, and the CG methylation level was significantly increased by 8.74%. In rTTT0, the coding gene bodies are hypermethylated in the CG context, while their promoter regions are hypomethylated in the non-CG context. DNA methylation changes in the leaf shape variation-associated coding genes, ARF10, IAA20, ROF1, and TPR2, were maintained for five generations of rTTT0. Interestingly, grafting chimerism also affected transcription of the microRNA gene (MIR), among which the DNA methylation levels of the promoters of three MIRs associated with leaf shape variation were changed in rTTT0, and the DNA methylation modification of MIR319 was maintained to the fifth generation of selfed progeny of rTTT0 (rTTT5). These findings demonstrate that DNA methylation of coding and non-coding genes plays an important role in heterologous cell interaction-induced variation formation and its transgenerational inheritance.

Metabolic and molecular mechanisms underlying the foliar Zn application induced increase of 2-acetyl-1-pyrroline conferring the 'taro-like' aroma in pumpkin leaves.

Introduction: Fresh pumpkin leaf is popular vegetable for its rich nutrition. The pleasant taro-like odour is important aroma quality of crops, and mostly contributed by 2-acetyl-1-pyrroline in pumpkin. Element Zn can impact metabolite biosynthesis in plants, including aroma formation. However, Zn-induced biochemical responses, especially 2-acetyl-1-pyrroline formation in pumpkin, haven't been elucidated. Methods: This study integrated metabolome and transcriptome to explore molecular fluctuations in pumpkin leaves at different time intervals after foliar Zn treatment. Result and Discussion: We first identified more than one thousand metabolites from pumpkin leaves by integrating different mass spectrometry methods according to the form in which a metabolite exists. Comparative metabolomic analysis revealed there were separately 25 out of 50 and 286 out of 963 metabolites that were respectively identified by gas chromatography-mass spectrometry and liquid chromatography-tandem mass spectrometry, differentially regulated by Zn treatment. Our findings revealed that 50mg/L of Zn significantly enhanced 2-acetyl-1-pyrroline production by more than 38%, which was contributed by increased biosynthesis of its precursors, including ornithine and proline. The following transcriptome analysis discovered 30,574 genes, including 953 novel genes. Zn treatment induced the differential expression of 41.6% of identified genes which were supposed to regulate the downstream metabolite changes in a time-dependent manner. Pathway analysis indicated that alternations in primary metabolism, including carbon metabolism and biosynthesis of amino acids, were vital to the fluctuated aromatic compound generation. Phytohormones and transcription factors may regulate the expression of gene P5CS and proline biosynthesis, which, therefore, affect 2-acetyl-1-pyrroline production. This research reveals molecular mechanisms of 2-acetyl-1-pyrroline formation in pumpkin, which will provide the molecular basis for desired aroma compound production through metabolite engineering.

QTL mapping reveals candidate genes for main agronomic traits in Luffa based on a high-resolution genetic map.

Luffa is an important medicinal and edible vegetable crop of Cucurbitaceae. Strong heterosis effects and strikingly complementary characteristics were found between the two domesticated Luffa cultivars, Luffa acutangula and Luffa cylindrica. To explore the genetic basis underlying their important agronomic traits, we constructed the first interspecific high-density genetic linkage map using a BC1 population of 110 lines derived from a cross between S1174 (Luffa acutangula) and P93075 (Luffa cylindrica). The map spanned a total of 2246.74 cM with an average distance of 0.48 cM between adjacent markers. Thereafter, a large-scale field-based quantitative trait loci (QTLs) mapping was conducted for 25 important agronomic traits and 40 significant genetic loci distributed across 11 chromosomes were detected. Notably, a vital QTL (qID2) located on chromosome 9 with a minimum distance of 23 kb was identified to be responsible for the internode diameter and explained 11% of the phenotypic variation. Lac09g006860 (LacCRWN3), encoding a nuclear lamina protein involved in the control of nuclear morphology, was the only gene harbored in qID2. Sequence alignment showed completely different promoter sequences between the two parental alleles of LacCRWN3 except for some nonsynonymous single nucleotide polymorphisms (SNPs) in exons, and the expression level in thick-stem P93075 was distinctively higher than that in thin-stem S1174. According to the natural variation analysis of a population of 183 inbred lines, two main haplotypes were found for LacCRWN3: the P93075-like and S1174-like, with the former haplotype lines exhibiting significantly thicker internode diameters than those of the latter haplotype lines. It showed that LacCRWN3, as the only CRWN3 gene in Cucurbitaceae, was the most likely candidate gene regulating the internode diameter of Luffa. Our findings will be beneficial for deciphering the molecular mechanism of key phenotypic traits and promoting maker-assisted breeding in Luffa.

Immune Enhancement of Nanoparticle-Encapsulated Ginseng Stem-Leaf Saponins on Porcine Epidemic Diarrhea Virus Vaccine in Mice.

Porcine epidemic diarrhea virus (PEDV) causes severe enteric disease in pigs, particularly neonatal piglets. Current vaccines do not provide complete protection against PEDV. Ginseng stem-leaf saponins (GSLS), a promising oral adjuvant candidate, can improve intestinal immune responses in poultry and mice. However, its low stability limits further use. Poly lactic-co-glycolic acid (PLGA), a biocompatible and biodegradable nanoparticle, has been widely used in biomedicine for stable and targeted drug delivery. In this study, we developed GSLS-PLGA nanoparticles (GSLS-NPs) and evaluated the mucosal adjuvant efficacy in vitro and in vivo. GSLS-NPs significantly enhanced antigen internalization and pro-inflammatory cytokine secretion by DC2.4 cells. Mice orally administered GSLS-NPs before intramuscular inoculation generated CD11b+CD8α- and CD11b-CD103+ dendritic cells in the spleen and draining mesenteric lymph nodes, respectively, which are the types mainly responsible for antigen presentation. Additionally, enhanced neutralizing and non-neutralizing antibody responses and expanded activities of specific effector and memory CD4+ and CD8+ T cells were also observed in mice immunized with PEDV vaccines plus GSLS-NPs compared to mice receiving the vaccines alone. Furthermore, GSLS-NPs showed a good safety profile and presented great advantages over GSLS aqueous solution. Collectively, our results highlight the potential of GSLS-NPs as a mucosal adjuvant and provide an attractive vaccination strategy for combatting PEDV. Further study is required to evaluate the efficacy of this mucosal adjuvant in swine.

Molecular Detection and Genotyping of Toxoplasma gondii from Pigs for Human Consumption in Zhejiang and Jiangsu Provinces, Eastern China.

Toxoplasma gondii infections are common in humans and animals worldwide. Ingestion of raw or undercooked meat containing tissue cysts of T. gondii is one major source of transmission of this parasite. It is important to guarantee the meat quality of China since our pork industry produces about half of the world's pork. In this study, a total of 746 pig samples were collected from Zhejiang and Jiangsu provinces in eastern China, and examined for T. gondii infection by PCR amplification targeting B1 gene. In this study, we found that 57 of 746 (7.6%) pigs were positive for B1 gene, with 8.5% (48/562) in Zhejiang province and 4.9% (9/184) in Jiangsu province, respectively. The positive DNA samples were further genotyped at 11 genetic markers, including SAG1, 5'-and 3'-SAG2, alternative SAG2, SAG3, BTUB, GRA6, L358, PK1, c22-8, c29-2, and an apicoplast locus Apico through PCR-restriction fragment length polymorphism (PCR-RFLP) technology. Two genotypes (ToxoDB 9 and ToxoDB 10) of T. gondii were identified by PCR-RFLP in Zhejiang province. However, both genotypes were not determined from Jiangsu province, which is speculated on the low DNA concentration and the small number of samples. These results indicate that T. gondii infection is endemic in pigs in eastern China and may raise public food safety concerns, suggesting more interventions for T. gondii-related risks are needed in the future.

Characterization of a Hyaluronidase-Producing Bacillus sp. CQMU-D Isolated from Soil.

Hyaluronidase (HAase) can depolymerize mucopolysaccharide hyaluronic acid (HA) to increase the efficacy of drug diffusion in the case of pathogenic bacteria. Due to its widespread medical applications, HAase originating from microorganisms has attracted significant attention. In this study, the HAase-producing bacterium Bacillus sp. CQMU-D was isolated from soil and identified based on its morphology and 16S rRNA gene sequencing. Enzyme activity was detected by measuring the content of reducing sugar in HA degradation products with 3,5-dinitrosalicylic acid (DNSA) or p-dimethylaminobenzaldehyde (DMAB) as the detection reagent. The results revealed that HAase reached maximum activity after 48 h of cultivation. Gene function annotation after full-length sequencing showed increased transport and metabolic activities associated with HAase. Additionally, the HAase gene of Bacillus sp. CQMU-D was different from the existing microbial HAase, and its protein was predicted to be a stable secretory protein with a conserved GAG_Lyase domain. These results characterized a new HAase-producing Bacillus from the soil via enzyme activity and bioinformatic analysis, expanding the knowledge on Bacillus HAase for potential industrial applications.

[Pregnancy Outcomes and Prognosis of Fetal Ventriculomegaly: A Retrospective Cohort Study].

Objective: To evaluate the pregnancy outcomes and neurodevelopment prognosis of subjects prenatally diagnosed with fetal ventriculomegaly (VM). Methods: All the subjects with VM diagnosed by ultrasound and were admitted and treated at West China Second Hospital, Sichuan University between March 2011 and September 2020 were retrospectively enrolled for a chohort study, while non-VM subjects of the same period were selected with a random number table to form the control group. Pregnancy outcomes of the two groups were compared, and the fetuses of both groups were followed up after birth for further assessment and comparison of their neurodevelopmental prognosis. Results: The live birth rate of the VM group was lower than that of the control group (77.63% [229/295] vs. 94.31% [265/281], P<0.001). Furthermore, the proportion of subjects that were transferred to NICU for monitoring and observation after birth was higher in the VM group than that of the control group (20.96% [48/229] vs. 4.53% [12/265], P<0.001). During the follow-up, it was found that the rate of neurodevelopmental abnormalities of the VM group was significantly higher than that of the control group (11.79% [27/229] vs. 1.90% [5/265], P<0.001). Moreover, neurodevelopmental abnormalities of VM fetuses were correlated to the following factors, the degree of VM ( P=0.010), intrauterine progression of VM ( P=0.024), and whether the postnatal cranial ultrasound result was suggestive of VM ( P=0.001). In addition, postnatal cranial ultrasound suggestive of VM was found to be an independent risk factor for neurodevelopmental abnormalities ( OR=9.434, 95% CI: 1.791-49.688, P=0.008). Conclusion: VM reduces the fetal live birth rate and may increase the risks of neurodevelopmental abnormalities after birth. All VM fetuses should be closely followed up for neurodevelopment status after birth, especially those with severe VM, intrauterine progression, and postnatal cranial ultrasound indicative of VM.

Metabolome and Transcriptome Analyses of Cucurbitacin Biosynthesis in Luffa (Luffa acutangula).

Cucurbitacins are extremely bitter compounds mainly present in Cucurbitaceae, where Luffa belongs. However, there is no comprehensive analysis of cucurbitacin biosynthesis in Luffa fruit. Therefore, this study analyzed bitter (WM709) and non-bitter (S1174) genotypes of Luffa to reveal the underlying mechanism of cucurbitacin biosynthesis by integrating metabolome and transcriptome analyses. A total of 422 metabolites were detected, including vitamins, essential amino acids, antioxidants, and antitumor substances. Of these, 131 metabolites showed significant differences between bitter (WM709) and non-bitter (S1174) Luffa fruits. The levels of isocucurbitacin B, cucurbitacin D, 23,24-dihydro cucurbitacin E, cucurbitacin F were significantly higher in bitter than in non-bitter Luffa. Transcriptome analysis showed that Bi, cytochromes P450s (CYP450s), and acyltransferase (ACT) of the cucurbitacin biosynthesis pathway, were significantly up-regulated. Moreover, drought stress and abscisic acid (ABA) activated genes of the cucurbitacin biosynthesis pathway. Furthermore, dual-luciferase reporter and yeast one-hybrid assays demonstrated that ABA-response element binding factor 1 (AREB1) binds to the Bi promoter to activate Bi expression. Comparative analysis of the Luffa and cucumber genomes showed that Bi, CYP450s, and ACT are located in the conserved syntenic loci, and formed a cucurbitacin biosynthesis cluster. This study provides important insights into major genes and metabolites of the cucurbitacin biosynthetic pathway, deepening the understanding of regulatory mechanisms of cucurbitacin biosynthesis in Luffa.

Identification of suitable reference genes for quantitative reverse transcription PCR in Luffa (Luffa cylindrica).

Reverse transcription real-time quantitative PCR is widely used to quantify gene expression. Reference genes are usually used as internal controls to measure the target gene expression level. To date, there is no consensus on the use of systematically validated reference genes in different tissues of Luffa. This study evaluated the expression stability of 11 candidate reference genes in different tissues using five algorithms (BestKeeper, comparative delta-Ct method, GeNorm, NormFinder, and RefFinder). Protein phosphatase 2A was the most stable gene, while alpha Tubulin was the least stable. The relative expression of ethylene-related genes in different tissues was also analyzed to reveal their role in sex determination. This study provides the basis for using suitable reference genes to evaluate targeted gene expression. Supplementary Information: The online version contains supplementary material available at 10.1007/s12298-022-01182-8.

A mutation in LacDWARF1 results in a GA-deficient dwarf phenotype in sponge gourd (Luffa acutangula).

KEY MESSAGE: A dwarfism gene LacDWARF1 was mapped by combined BSA-Seq and comparative genomics analyses to a 65.4 kb physical genomic region on chromosome 05. Dwarf architecture is one of the most important traits utilized in Cucurbitaceae breeding because it saves labor and increases the harvest index. To our knowledge, there has been no prior research about dwarfism in the sponge gourd. This study reports the first dwarf mutant WJ209 with a decrease in cell size and internodes. A genetic analysis revealed that the mutant phenotype was controlled by a single recessive gene, which is designated Lacdwarf1 (Lacd1). Combined with bulked segregate analysis and next-generation sequencing, we quickly mapped a 65.4 kb region on chromosome 5 using F2 segregation population with InDel and SNP polymorphism markers. Gene annotation revealed that Lac05g019500 encodes a gibberellin 3ß-hydroxylase (GA3ox) that functions as the most likely candidate gene for Lacd1. DNA sequence analysis showed that there is an approximately 4 kb insertion in the first intron of Lac05g019500 in WJ209. Lac05g019500 is transcribed incorrectly in the dwarf mutant owing to the presence of the insertion. Moreover, the bioactive GAs decreased significantly in WJ209, and the dwarf phenotype could be restored by exogenous GA3 treatment, indicating that WJ209 is a GA-deficient mutant. All these results support the conclusion that Lac05g019500 is the Lacd1 gene. In addition, RNA-Seq revealed that many genes, including those related to plant hormones, cellular process, cell wall, membrane and response to stress, were significantly altered in WJ209 compared with the wild type. This study will aid in the use of molecular marker-assisted breeding in the dwarf sponge gourd.

Escherichia coli Heat-Labile Enterotoxin B Subunit Combined with Ginsenoside Rg1 as an Intranasal Adjuvant Triggers Type I Interferon Signaling Pathway and Enhances Adaptive Immune Responses to an Inactivated PRRSV Vaccine in ICR Mice.

Porcine reproductive and respiratory syndrome virus (PRRSV) is a major pathogen that has threatened the global swine industry for almost 30 years. Because current vaccines do not provide complete protection, exploration of new preventive strategies is urgently needed. Here, we combined a heat-labile enterotoxin B subunit of Escherichia coli (LTB) and ginsenoside Rg1 to form an intranasal adjuvant and evaluated its enhancement of immune responses in mice when added to an inactivated-PRRSV vaccine. The combination adjuvant synergistically elicited higher neutralizing and non-neutralizing (immunoglobulin G and A) antibody responses in the circulatory system and respiratory tract, and enhanced T and B lymphocyte proliferation, CD4+ T-cell priming, and cytotoxic CD4+ T cell activities in mononuclear cells from spleen and lung tissues when compared to the PRRSV vaccine alone, and it resulted in balanced Th1/Th2/Th17 responses. More importantly, we observed that the combination adjuvant also up-regulated type I interferon signaling, which may contribute to improvement in adaptive immune responses. These results highlight the potential value of a combined adjuvant approach for improving the efficacy of vaccination against PRRSV. Further study is required to evaluate the efficacy of this combined adjuvant in swine.