KYNA Ameliorates Glutamate Toxicity of HAND by Enhancing Glutamate Uptake in A2 Astrocytes.

Reactive astrocytes are key players in HIV-associated neurocognitive disorders (HAND), and different types of reactive astrocytes play opposing roles in the neuropathologic progression of HAND. A recent study by our group found that gp120 mediates A1 astrocytes (neurotoxicity), which secrete proinflammatory factors and promote HAND disease progression. Here, by comparing the expression of A2 astrocyte (neuroprotective) markers in the brains of gp120 tgm mice and gp120+/α7nAChR-/- mice, we found that inhibition of alpha 7 nicotinic acetylcholine receptor (α7nAChR) promotes A2 astrocyte generation. Notably, kynurenine acid (KYNA) is an antagonist of α7nAChR, and is able to promote the formation of A2 astrocytes, the secretion of neurotrophic factors, and the enhancement of glutamate uptake through blocking the activation of α7nAChR/NF-κB signaling. In addition, learning, memory and mood disorders were significantly improved in gp120 tgm mice by intraperitoneal injection of kynurenine (KYN) and probenecid (PROB). Meanwhile, the number of A2 astrocytes in the mouse brain was significantly increased and glutamate toxicity was reduced. Taken together, KYNA was able to promote A2 astrocyte production and neurotrophic factor secretion, reduce glutamate toxicity, and ameliorate gp120-induced neuropathological deficits. These findings contribute to our understanding of the role that reactive astrocytes play in the development of HAND pathology and provide new evidence for the treatment of HAND via the tryptophan pathway.

A Viral Protein 4-Based Trivalent Nanoparticle Vaccine Elicited High and Broad Immune Responses and Protective Immunity against the Predominant Rotaviruses.

The current live rotavirus (RV) vaccines show reduced effectiveness in developing countries, calling for vaccine strategies with improved efficacy and safety. We generated pseudovirus nanoparticles (PVNPs) that display multiple ectodomains of RV viral protein 4 (VP4), named S-VP4e, as a nonreplicating RV vaccine candidate. The RV spike protein VP4s that bind host receptors and facilitate viral entry are excellent targets for vaccination. In this study, we developed scalable methods to produce three S-VP4e PVNPs, each displaying the VP4e antigens from one of the three predominant P[8], P[4], and P[6] human RVs (HRVs). These PVNPs were recognized by selected neutralizing VP4-specific monoclonal antibodies, bound glycan receptors, attached to permissive HT-29 cells, and underwent cleavage by trypsin between VP8* and VP5*. 3D PVNP models were constructed to understand their structural features. A trivalent PVNP vaccine containing the three S-VP4e PVNPs elicited high and well-balanced VP4e-specific antibody titers in mice directed against the three predominant HRV P types. The resulting antisera neutralized the three HRV prototypes at high titers; greater than 4-fold higher than the neutralizing responses induced by a trivalent vaccine consisting of the S60-VP8* PVNPs. Finally, the trivalent S-VP4e PVNP vaccine provided 90-100% protection against diarrhea caused by HRV challenge. Our data supports the trivalent S-VP4e PVNPs as a promising nonreplicating HRV vaccine candidate for parenteral delivery to circumvent the suboptimal immunization issues of all present live HRV vaccines. The established PVNP-permissive cell and PVNP-glycan binding assays will be instrumental for further investigating HRV-host cell interactions and neutralizing effects of VP4-specific antibodies and antivirals.

The 2.6 Å Structure of a Tulane Virus Variant with Minor Mutations Leading to Receptor Change.

Human noroviruses (HuNoVs) are a major cause of acute gastroenteritis, contributing significantly to annual foodborne illness cases. However, studying these viruses has been challenging due to limitations in tissue culture techniques for over four decades. Tulane virus (TV) has emerged as a crucial surrogate for HuNoVs due to its close resemblance in amino acid composition and the availability of a robust cell culture system. Initially isolated from rhesus macaques in 2008, TV represents a novel Calicivirus belonging to the Recovirus genus. Its significance lies in sharing the same host cell receptor, histo-blood group antigen (HBGA), as HuNoVs. In this study, we introduce, through cryo-electron microscopy (cryo-EM), the structure of a specific TV variant (the 9-6-17 TV) that has notably lost its ability to bind to its receptor, B-type HBGA-a finding confirmed using an enzyme-linked immunosorbent assay (ELISA). These results offer a profound insight into the genetic modifications occurring in TV that are necessary for adaptation to cell culture environments. This research significantly contributes to advancing our understanding of the genetic changes that are pivotal to successful adaptation, shedding light on fundamental aspects of Calicivirus evolution.

Pseudovirus Nanoparticles Displaying Plasmodium Circumsporozoite Proteins Elicited High Titers of Sporozoite-Binding Antibody.

BACKGROUND: malaria caused by Plasmodium parasites remains a public health threat. The circumsporozoite proteins (CSPs) of Plasmodium sporozoite play a key role in Plasmodium infection, serving as an excellent vaccine target. METHODS: using a self-assembled S60 nanoparticle platform, we generated pseudovirus nanoparticles (PVNPs) displaying CSPs, named S-CSPs, for enhanced immunogenicity. RESULTS: purified Hisx6-tagged or tag-free S-CSPs self-assembled into PVNPs that consist of a norovirus S60 inner shell and multiple surface-displayed CSPs. The majority of the PVNPs measured ~27 nm with some size variations, and their three-dimensional structure was modeled. The PVNP-displayed CSPs retained their glycan receptor-binding function. A mouse immunization study showed that PVNPs induced a high antibody response against CSP antigens and the PVNP-immunized mouse sera stained the CSPs of Plasmodium sporozoites at high titer. CONCLUSIONS AND DISCUSSION: the PVNP-displayed CSPs retain their authentic antigenic feature and receptor-binding function. The CSP-specific antibody elicited by the S-CSP PVNPs binds original CSPs and potentially inhibits the attachment of Plasmodium sporozoites to their host cells, a key step for liver invasion by the sporozoites. Thus, S-CSP PVNPs may be an excellent vaccine candidate against malaria caused by Plasmodium parasites.

Role of the Metal Precursor in Preparing Dual-Atom Catalysts for the Oxygen Reduction Reaction.

Dual-atom catalysts (DACs) have arisen as a novel type of heterogeneous catalyst that extends from single-atom catalysts (SACs) by incorporating two kinds of metals. These materials have demonstrated enhanced performance when compared to SACs. The choice of metal precursors plays an important role in the synthesis of DACs. Here, we choose Fe and Co as DAC models and study types, contents, molar ratios of two precursors, and oxygen reduction reaction (ORR) activity. The Fe,Co DACs were synthesized by an adsorption-annealing approach, using nitrogen-doped graphitic carbon (NC) as the support. As a result, the adsorption ability of metal precursors on the support determines the metal loadings in Fe and Co DACs, leading to differences in ORR performance. The Fe precursors win the adsorption competitions in most cases, resulting in a much higher loading than that of Co precursors. Importantly, it is difficult to increase the precursor content by simply increasing the initial amount. Choosing the right combination of metal precursors, such as ferrocene and cobalt chloride, can yield Fe,Co DACs with enhanced ORR performance..

A bioengineered pseudovirus nanoparticle displaying SARS-CoV 2 RBD fully protects mice from mortality and weight loss caused by SARS-CoV 2 challenge.

The COVID-19 pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused considerable morbidity and mortality worldwide. Although authorized COVID-19 vaccines have been shown highly effective, their significantly lower efficacy against heterologous variants, and the rapid decrease of vaccine-elicited immunity raises serious concerns, calling for improved vaccine tactics. To this end, a pseudovirus nanoparticle (PVNP) displaying the receptor binding domains (RBDs) of SARS-CoV-2 spike, named S-RBD, was generated and shown it as a promising COVID-19 vaccine candidate. The S-RBD PVNP was produced using both prokaryotic and eukaryotic systems. A 3D structural model of the S-RBD PVNPs was built based on the known structures of the S60 particle and RBDs, revealing an S60 particle-based icosahedral symmetry with multiple surface-displayed RBDs that retain authentic conformations and receptor-binding functions. The PVNP is highly immunogenic, eliciting high titers of RBD-specific IgG and neutralizing antibodies in mice. The S-RBD PVNP demonstrated exceptional protective efficacy, and fully (100%) protected K18-hACE2 mice from mortality and weight loss after a lethal SARS-CoV-2 challenge, supporting the S-RBD PVNPs as a potent COVID-19 vaccine candidate. By contrast, a PVNP displaying the N-terminal domain (NTD) of SARS-CoV-2 spike exhibited only 50% protective efficacy. Since the RBD antigens of our PVNP vaccine are adjustable as needed to address the emergence of future variants, and various S-RBD PVNPs can be combined as a cocktail vaccine for broad efficacy, these non-replicating PVNPs offer a flexible platform for a safe, effective COVID-19 vaccine with minimal manufacturing cost and time.

Efficient Biosynthesis of Odd-Chain Fatty Acids via Regulating the Supply and Consumption of Propionyl-CoA in Schizochytrium sp.

Odd chain fatty acids (OCFAs) are high-value-added compounds with great application in the field of food and medicine. As an oleaginous microorganism, Schizochytrium sp. has the potential to produce OCFAs efficiently. Propionyl-CoA is used as a precursor to synthesize OCFAs through the fatty acid synthetase (FAS) pathway, so its flow direction determines the yield of OCFAs. Here, different substrates were assessed to promote propionyl-CoA supply for OCFA accumulation. Moreover, the methylmalonyl-CoA mutase (MCM) was identified as the key gene responsible for propionyl-CoA consumption, which promotes the propionyl-CoA to enter into the tricarboxylic acid cycle rather than the FAS pathway. As one of the classic B12-dependent enzymes, the activity of MCM can be inhibited in the absence of B12. As expected, the OCFA accumulation was greatly increased. However, the removal of B12 caused growth limitation. Furthermore, the MCM was knocked out to block the consumption of propionyl-CoA and to maintain cell growth; results showed that the engineered strain achieved the OCFAs titer of 2.82 g/L, which is 5.76-fold that of wild type. Last, a fed-batch co-feeding strategy was developed, resulting in the highest reported OCFAs titer of 6.82 g/L. This study provides guidance for the microbial production of OCFAs.

The αTSR Domain of Plasmodium Circumsporozoite Protein Bound Heparan Sulfates and Elicited High Titers of Sporozoite Binding Antibody After Displayed by Nanoparticles.

Introduction: Malaria is a devastating infectious illness caused by protozoan Plasmodium parasites. The circumsporozoite protein (CSP) on Plasmodium sporozoites binds heparan sulfate proteoglycan (HSPG) receptors for liver invasion, a critical step for prophylactic and therapeutic interventions. Methods: In this study, we characterized the αTSR domain that covers region III and the thrombospondin type-I repeat (TSR) of the CSP using various biochemical, glycobiological, bioengineering, and immunological approaches. Results: We found for the first time that the αTSR bound heparan sulfate (HS) glycans through support by a fused protein, indicating that the αTSR is a key functional domain and thus a vaccine target. When the αTSR was fused to the S domain of norovirus VP1, the fusion protein self-assembled into uniform S60-αTSR nanoparticles. Three-dimensional structure reconstruction revealed that each nanoparticle consists of an S60 nanoparticle core and 60 surface displayed αTSR antigens. The nanoparticle displayed αTSRs retained the binding function to HS glycans, indicating that they maintained authentic conformations. Both tagged and tag-free S60-αTSR nanoparticles were produced via the Escherichia coli system at high yield by scalable approaches. They are highly immunogenic in mice, eliciting high titers of αTSR-specific antibody that bound specifically to the CSPs of Plasmodium falciparum sporozoites at high titer. Discussion and Conclusion: Our data demonstrated that the αTSR is an important functional domain of the CSP. The S60-αTSR nanoparticle displaying multiple αTSR antigens is a promising vaccine candidate potentially against attachment and infection of Plasmodium parasites.

Combined Live Oral Priming and Intramuscular Boosting Regimen with Rotarix® and a Nanoparticle-Based Trivalent Rotavirus Vaccine Evaluated in Gnotobiotic Pig Models of G4P[6] and G1P[8] Human Rotavirus Infection.

Human rotavirus (HRV) is the causative agent of severe dehydrating diarrhea in children under the age of five, resulting in up to 215,000 deaths each year. These deaths almost exclusively occur in low- and middle-income countries where vaccine efficacy is the lowest due to chronic malnutrition, gut dysbiosis, and concurrent enteric viral infection. Parenteral vaccines for HRV are particularly attractive as they avoid many of the concerns associated with currently used live oral vaccines. In this study, a two-dose intramuscular (IM) regimen of the trivalent, nanoparticle-based, nonreplicating HRV vaccine (trivalent S60-VP8*), utilizing the shell (S) domain of the capsid of norovirus as an HRV VP8* antigen display platform, was evaluated for immunogenicity and protective efficacy against P[6] and P[8] HRV using gnotobiotic pig models. A prime-boost strategy using one dose of the oral Rotarix® vaccine, followed by one dose of the IM trivalent nanoparticle vaccine was also evaluated. Both regimens were highly immunogenic in inducing serum virus neutralizing, IgG, and IgA antibodies. The two vaccine regimens failed to confer significant protection against diarrhea; however, the prime-boost regimen significantly shortened the duration of virus shedding in pigs challenged orally with the virulent Wa (G1P[8]) HRV and significantly shortened the mean duration of virus shedding, mean peak titer, and area under the curve of virus shedding after challenge with Arg (G4P[6]) HRV. Prime-boost-vaccinated pigs challenged with P[8] HRV had significantly higher P[8]-specific IgG antibody-secreting cells (ASCs) in the spleen post-challenge. Prime-boost-vaccinated pigs challenged with P[6] HRV had significantly higher numbers of P[6]- and P[8]-specific IgG ASCs in the ileum, as well as significantly higher numbers of P[8]-specific IgA ASCs in the spleen post-challenge. These results suggest the promise of and warrant further investigation into the oral priming and parenteral boosting strategy for future HRV vaccines.

Enhancing the biomass and docosahexaenoic acid-rich lipid accumulation of Schizochytrium sp. in propionate wastewater.

In order to find a more effective way to obtain docosahexaenoic acid (DHA) rich lipid from Schizochytrium sp., a widespread propionate wastewater (PW) is used. PW is a common industrial and domestic wastewater, and transforming it into valuable products is a potential treatment method. Schizochytrium sp. is a rapidly growing oleaginous organism, which has been used commercially for DHA production. Herein, PW is completely used for DHA production by Schizochytrium sp. by genetic engineering and fermentation optimization, which can alleviate the increasingly tense demand for water resources and environmental pollution caused by industrial wastewater. Firstly, the methylmalonyl-CoA mutase (MCM) was overexpressed in Schizochytrium sp. to enhance the metabolism of propionate, then the engineered strain of overexpressed MCM (OMCM) can effectively use propionate. Then, the effects of PW with different concentration of propionate were investigated, and results showed that OMCM can completely replace clean water with PW containing 5 g L-1 propionate. Furthermore, in the fed-batch fermentation, the OMCM obtained the highest biomass of 113.4 g L-1 and lipid yield of 64.4 g L-1 in PW condition, which is 26.8% and 51.7% higher than that of wild type (WT) in PW condition. Moreover, to verify why overexpression of MCM can promote DHA and lipid accumulation, the comparative metabolomics, ATP production level, the antioxidant system, and the transcription of key genes were investigated. Results showed that ATP induced by PW condition could drive the synthesis of DHA, and remarkably improve the antioxidant capacity of cells by enhancing the carotenoids production. Therefore, PW can be used as an effective and economical substrate and water source for Schizochytrium sp. to accumulate biomass and DHA.

Enhanced docosahexaenoic acid production from cane molasses by engineered and adaptively evolved Schizochytrium sp.

Cane molasses (CM) is a sugar-rich agro-industrial byproduct. The purpose of this study is to synthesize docosahexaenoic acid (DHA) in Schizochytrium sp. by using CM. The single factor analysis showed that sucrose utilization was the main factor limiting the utilization of CM. Therefore, the endogenous sucrose hydrolase (SH) was overexpressed in Schizochytrium sp., which enhanced the sucrose utilization rate 2.57-fold compared to the wild type. Furthermore, adaptive laboratory evolution was used to further improve sucrose utilization from CM. Comparative proteomics and RT-qPCR were used out to analyze the metabolic differences of evolved strain grown on CM and glucose, respectively. Finally, a constant flow rate CM feeding strategy was implemented, whereby the DHA titer and lipid yield of the final strain OSH-end reached 25.26 g/L and 0.229 g/g sugar, respectively. This study demonstrated the CM is a cost-effective carbon source for industrial DHA fermentation.

Kynurenic acid blunts A1 astrocyte activation against neurodegeneration in HIV-associated neurocognitive disorders.

Despite extensive astrocyte activation in patients suffering from HIV-associated neurocognitive disorders (HAND), little is known about the contribution of astrocytes to HAND neuropathology. Here, we report that the robust activation of neurotoxic astrocytes (A1 astrocytes) in the CNS promotes neuron damage and cognitive deficits in HIV-1 gp120 transgenic mice. Notably, knockout of α7 nicotinic acetylcholine receptors (α7nAChR) blunted A1 astrocyte responses, ultimately facilitating neuronal and cognitive improvement in the gp120tg mice. Furthermore, we provide evidence that Kynurenic acid (KYNA), a tryptophan metabolite with α7nAChR inhibitory properties, attenuates gp120-induced A1 astrocyte formation through the blockade of α7nAChR/JAK2/STAT3 signaling activation. Meanwhile, compared with gp120tg mice, mice fed with tryptophan showed dramatic improvement in cognitive performance, which was related to the inhibition of A1 astrocyte responses. These initial and determinant findings mark a turning point in our understanding of the role of α7nAChR in gp120-mediated A1 astrocyte activation, opening up new opportunities to control neurotoxic astrocyte generation through KYNA and tryptophan administration.

Economical downstream processing of microbial polyunsaturated fatty acids.

Polyunsaturated fatty acids (PUFAs) are important nutrients for humans and animals. Microorganisms, such as yeast, filamentous fungi, and microalgae, have successfully been modified to produce PUFAs. Apart from strain improvement and fermentation optimization, efficient and cost-effective downstream processing will determine whether production can advance from the laboratory to the factory.

Function of the Polyketide Synthase Domains of Schizochytrium sp. on Fatty Acid Synthesis in Yarrowia lipolytica.

It is well known that polyunsaturated fatty acids (PUFAs) in Schizochytrium sp. are mainly synthesized via the polyketide synthase (PKS) pathway. However, the specific mechanism of PKS in fatty acid synthesis is still unclear. In this work, the functions of ORFA, ORFB, ORFC, and their individual functional domain genes on fatty acid synthesis were investigated through heterologous expression in Yarrowia lipolytica. The results showed that the expression of ORFA, ORFB, ORFC, and their individual functional domains all led to the increase of the very long-chain PUFA content (mainly eicosapentaenoic acid). Furthermore, the transcriptomic analysis showed that except for the 3-ketoacyl-ACP synthase (KS) domain of ORFB, the expression of an individual functional domain, including malonyl-CoA: ACP acyltransferase, 3-hydroxyacyl-ACP dehydratase (DH), 3-ketoacyl-ACP reductase, and KS domains of ORFA, acyltransferase domains of ORFB, and two DH domains of ORFC resulted in upregulation of the tricarboxylic acid cycle and pentose phosphate pathway, downregulation of the triacylglycerol biosynthesis, fatty acid synthesis pathway, and ß-oxidation in Yarrowia lipolytica. These results provide a theoretical basis for revealing the function of PKS in fatty acid synthesis in Y. lipolytica and elucidate the possible mechanism for PUFA biosynthesis.

Metal cofactor regulation combined with rational genetic engineering of Schizochytrium sp. for high-yield production of squalene.

The increasing market demand for squalene requires novel biotechnological production platforms. Schizochytrium sp. is an industrial oleaginous host with a high potential for squalene production due to its abundant native acetyl-CoA pool. We first found that iron starvation led to the accumulation of 1.5 g/L of squalene by Schizochytrium sp., which was 40-fold higher than in the control. Subsequent transcriptomic and lipidomic analyses showed that the high squalene titer is due to the diversion of precursors from lipid biosynthesis and increased triglycerides (TAG) content for squalene storage. Furthermore, we constructed the engineered acetyl-CoA C-acetyltransferase (ACAT)-overexpressing strain 18S::ACAT, which produced 2.79 g/L of squalene, representing an 86% increase over the original strain. Finally, a nitrogen-rich feeding strategy was developed to further increase the squalene titer of the engineered strain, which reached 10.78 g/L in fed-batch fermentation, a remarkable 161-fold increase over the control. To our best knowledge, this is the highest squalene yield in thraustochytrids reported to date.

The Association between Symptomatic Rotavirus Infection and Histo-Blood Group Antigens in Young Children with Diarrhea in Pretoria, South Africa.

OBJECTIVES: Recently, histo-blood group antigens (HBGAs) have been identified as receptors or attachment factors of several viral pathogens. Among rotaviruses, HBGAs interact with the outer viral protein, VP4, which has been identified as a potential susceptibility factor, although the findings are inconsistent throughout populations due to HBGA polymorphisms. We investigated the association between HBGA phenotypes and rotavirus infection in children with acute gastroenteritis in northern Pretoria, South Africa. METHODS: Paired diarrheal stool and saliva samples were collected from children aged ≤ 59 months (n = 342) with acute moderate to severe diarrhea, attending two health care facilities. Rotaviruses in the stool samples were detected by commercial EIA and the rotavirus strains were characterized by RT-PCR targeting the outer capsid VP7 (G-type) and VP4 (P-type) antigens for genotyping. Saliva-based ELISAs were performed to determine A, B, H, and Lewis antigens for blood group typing. RESULTS: Blood type O was the most common blood group (62.5%) in this population, followed by groups A (26.0%), B (9.3%), and AB (2.2%). The H1-based secretors were common (82.7%) compared to the non-secretors (17.3%), and the Lewis antigen positive phenotypes (Le(a+b+)) were predominant (54.5%). Blood type A children were more likely to be infected by rotavirus (38.8%) than any other blood types. P[4] rotaviruses (21/49; 42.9%) infected only secretor individuals, whereas P[6] rotaviruses (3/49; 6.1%) only infected Le(a-b-), although the numbers were very low. On the contrary, P[8] rotaviruses infected children with a wide range of blood group phenotypes, including Le(a-b-) and non-secretors. CONCLUSIONS: Our findings demonstrated that Lewis antigens, or the lack thereof, may serve as susceptibility factors to rotaviral infection by specific VP4 genotypes as observed elsewhere. Potentially, the P[8] strains remain the predominant human VP4 genotype due to their ability to bind to a variety of HBGA phenotypes.

Efficient co-production of EPA and DHA by Schizochytrium sp. via regulation of the polyketide synthase pathway.

Presently, the supply of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) traditionally produced by marine fisheries will be insufficient to meet their market demand in food industry. Thus a sustainable alternative source is urgently required. Schizochytrium sp. is an ideal producer of DHA; however, its ability to co-produce DHA and EPA has not yet been proved. Herein, we first described a cobalamin-independent methionine synthase-like (MetE-like) complex, which contains independent acyltransferase and 3-ketoacyl synthase domains, independent of the traditional polyketide synthase (PKS) system. When the MetE-like complex was activated, the EPA content was increased from 1.26% to 7.63%, which is 6.06-folds higher than that in the inactivated condition. Through lipidomics, we find that EPA is more inclined to be stored as triglyceride. Finally, the EPA production was enhanced from 4.19 to 29.83 (mg/g cell dry weight) using mixed carbon sources, and the final yield reached 2.25 g/L EPA and 9.59 g/L DHA, which means that Schizochytrium sp. has great market potential for co-production of EPA and DHA.

Engineering the xylose metabolism in Schizochytrium sp. to improve the utilization of lignocellulose.

BACKGROUND: Schizochytrium sp. is a heterotrophic, oil-producing microorganism that can efficiently produce lipids. However, the industrial production of bulk chemicals using Schizochytrium sp. is still not economically viable due to high-cost culture medium. Replacing glucose with cheap and renewable lignocellulose is a highly promising approach to reduce production costs, but Schizochytrium sp. cannot efficiently metabolize xylose, a major pentose in lignocellulosic biomass. RESULTS: In order to improve the utilization of lignocellulose by Schizochytrium sp., we cloned and functionally characterized the genes encoding enzymes involved in the xylose metabolism. The results showed that the endogenous xylose reductase and xylulose kinase genes possess corresponding functional activities. Additionally, attempts were made to construct a strain of Schizochytrium sp. that can effectively use xylose by using genetic engineering techniques to introduce exogenous xylitol dehydrogenase/xylose isomerase; however, the introduction of heterologous xylitol dehydrogenase did not produce a xylose-utilizing engineered strain, whereas the introduction of xylose isomerase did. The results showed that the engineered strain 308-XI with an exogenous xylose isomerase could consume 8.2 g/L xylose over 60 h of cultivation. Xylose consumption was further elevated to 11.1 g/L when heterologous xylose isomerase and xylulose kinase were overexpressed simultaneously. Furthermore, cultivation of 308-XI-XK(S) using lignocellulosic hydrolysates, which contained glucose and xylose, yielded a 22.4 g/L of dry cell weight and 5.3 g/L of total lipid titer, respectively, representing 42.7 and 30.4% increases compared to the wild type. CONCLUSION: This study shows that engineering of Schizochytrium sp. to efficiently utilize xylose is conducive to improve its utilization of lignocellulose, which can reduce the costs of industrial lipid production.

A Pseudovirus Nanoparticle-Based Trivalent Rotavirus Vaccine Candidate Elicits High and Cross P Type Immune Response.

Rotavirus infection continues to cause significant morbidity and mortality globally. In this study, we further developed the S60-VP8* pseudovirus nanoparticles (PVNPs) displaying the glycan receptor binding VP8* domains of rotavirus spike proteins as a parenteral vaccine candidate. First, we established a scalable method for the large production of tag-free S60-VP8* PVNPs representing four rotavirus P types, P[8], P[4], P[6], and P[11]. The approach consists of two major steps: selective precipitation of the S-VP8* proteins from bacterial lysates using ammonium sulfate, followed by anion exchange chromatography to further purify the target proteins to a high purity. The purified soluble proteins self-assembled into S60-VP8* PVNPs. Importantly, after intramuscular injections, the trivalent vaccine consisting of three PVNPs covering VP8* antigens of P[8], P[4], and P[6] rotaviruses elicited high and broad immunogenicity in mice toward the three predominant P-type rotaviruses. Specifically, the trivalent vaccine-immunized mouse sera showed (1) high and balanced IgG and IgA antibody titers toward all three VP8* types, (2) high blocking titer against the VP8*-glycan receptor interaction, and (3) high and broad neutralizing titers against replications of all P[8], P[4], and P[6] rotaviruses. Therefore, trivalent S60-VP8* PVNPs are a promising non-replicating, parenteral vaccine candidate against the most prevalent rotaviruses worldwide.

Generalist endophyte Phomopsis liquidambaris colonization of Oryza sativa L. promotes plant growth under nitrogen starvation.

Fungal endophytes establish symbiotic relationships with host plants, which results in a mutual growth benefit. However, little is known about the plant genetic response underpinning endophyte colonization. Phomopsis liquidambaris usually lives as an endophyte in a wide range of asymptomatic hosts and promotes biotic and abiotic stress resistance. In this study, we show that under low nitrogen conditions P. liquidambaris promotes rice growth in a hydroponic system, which is free of other microorganisms. In order to gain insights into the mechanisms of plant colonization by P. liquidambaris under low nitrogen conditions, we compared root and shoot transcriptome profiles of root-inoculated rice at different colonization stages. We determined that genes related to plant growth promotion, such as gibberellin and auxin related genes, were up-regulated at all developmental stages both locally and systemically. The largest group of up-regulated genes (in both roots and shoots) were related to flavonoid biosynthesis, which is involved in plant growth as well as antimicrobial compounds. Furthermore, genes encoding plant defense-related endopeptidase inhibitors were strongly up-regulated at the early stage of colonization. Together, these results provide new insights into the molecular mechanisms of plant-microbe mutualism and the promotion of plant growth by a fungal endophyte under nitrogen-deficient conditions.