Live-attenuated PruΔgra72 strain of Toxoplasma gondii induces strong protective immunity against acute and chronic toxoplasmosis in mice.

BACKGROUND: Toxoplasma gondii is an intracellular opportunistic pathogenic protozoan that poses serious threats, particularly in immunocompromised individuals. In the absence of a robust prophylactic measure, the mitigation and management of toxoplasmosis present formidable challenges to public health. We recently found that GRA72 plays an important role in parasitophorous vacuole (PV) morphology, growth and virulence of T. gondii. However, whether gra72-deficient strain can be used as a vaccine remains unknown. METHODS: We first examined the attenuated virulence of gra72 gene knockout strain (PruΔgra72) and the parasite load in organs of the infected mice. Subsequently, we evaluated the immune-protective effects of the PruΔgra72 vaccination against challenge with various types of T. gondii tachyzoites and Pru cysts. Furthermore, levels of antibodies and cytokines induced by PruΔgra72 vaccination were examined. Statistical analysis was conducted by Student's t-test or Mantel-Cox log-rank test based on data obtained from three independent experiments with GraphPad Prism 8.0. RESULTS: We found that PruΔgra72 strain exhibited a significantly attenuated virulence even at the highest dose of 5 × 107 tachyzoites in Kunming mice model. The significant decrease of brain cyst burden and parasite load in the organs of the PruΔgra72-infected mice suggested its potentiality as a live-attenuated vaccine. Hence, we explored the protective immunity of PruΔgra72 vaccination against toxoplasmosis. Results showed that vaccination with 5 × 106 PruΔgra72 tachyzoites triggered a strong and sustained Th1-biased immune response, marked by significantly increased levels of anti-T. gondii IgG antibodies, and significantly higher levels of Th1 type cytokines (IL-2, IL-12 and IFN-γ) compared to that of Th2 type (IL-4 and IL-10). Vaccination with 5 × 106 PruΔgra72 tachyzoites in mice conferred long-term protection against T. gondii infection by less virulent tachyzoites (ToxoDB#9 PYS and Pru strains) and Pru cysts, provided partial protection against acute infection by high virulent Type I RH tachyzoites and significantly decreased brain cyst burden of chronically infected mice. CONCLUSIONS: The avirulent PruΔgra72 induced strong protective immunity against acute and chronic T. gondii infection and is a promising candidate for developing a safe and effective live-attenuated vaccine against T. gondii infection.

Clinical Characteristics and Blood Test Results in COVID-19 Patients.

OBJECTIVE: An outbreak of pneumonia named COVID-19 caused by a novel coronavirus in Wuhan is rapidly spreading worldwide. The objective of the present study was to clarify further the clinical characteristics and blood parameters in COVID-19 patients. MATERIALS AND METHODS: Twenty-three suspected patients and 64 patients with laboratory-confirmed SARS-Cov-2 infection were admitted to a designated hospital. Epidemiological, clinical, laboratory, and treatment data were collected and analyzed. RESULTS: Of the 64 patients studied, 47 (73.4%) had been exposed to a confirmed source of COVID-19 transmission. On admission, the most common symptoms were fever (75%) and cough (76.6%). Twenty-eight (43.8%) COVID-19 patients showed leukopenia, 10 (15.6%) showed lymphopenia, 47 (73.4%) and 41 (64.1%) had elevated high-sensitivity C-reactive protein (hsCRP) and erythrocyte sedimentation rate (ESR), respectively, and 30 (46.9%) had increased fibrinogen concentration. After the treatment, the counts of white blood cells and platelets, and the level of prealbumin increased significantly, while aspartate aminotransferase (AST), lactate dehydrogenase (LDH), and hsCRP decreased. COVID-19 patients with the hospital stay longer than 12 days had higher body mass index (BMI) and increased levels of AST, LDH, fibrinogen, hsCRP, and ESR. CONCLUSIONS: Results of blood tests have potential clinical value in COVID-19 patients.

Influence of Ceramic Debris on Osteoblast Behaviors: An In Vivo Study.

OBJECTIVE: Wear-induced aseptic loosening has been accepted as one of the main reasons for failure of total hip arthroplasty. Ceramic wear debris is generated following prosthesis implantation and plays an important part in the upregulation of inflammatory factors in total hip arthroplasty. The present study investigates the influence of ceramic debris on osteoblasts and inflammatory factors. METHODS: Ceramic debris was prepared by mechanical grinding of an aluminum femoral head and added to cultures of MC3T3-E subclone 14 cells at different concentrations (i.e. 0, 5, 10, and 15 µg/mL). Cell proliferation was evaluated using a Cell Counting Kit (CCK-8), and cell differentiation was assessed by mRNA expression of alkaline phosphatase (ALP), osteocalcin (OCN), and osteopontin (OPN). In addition, cell bio-mineralization was evaluated through alizarin red S staining, and release of tumor necrosis factor alpha (TNF-α), interleukin-1 beta (IL-1ß), and interleukin-6 (IL-6) was measured through enzyme-linked immunosorbent assays (ELISA). Furthermore, mRNA expression of Smad1, Smad4, and Smad5 and protein expression of phosphorylated Smad1, Smad4, and Smad5 were measured by reverse transcriptase polymerase chain reaction (RT-PCR) and western blotting. RESULTS: The ceramic debris had irregular shapes and sizes, and analysis of the size distribution using a particle size analyzer indicated that approximately 90% of the ceramic debris was smaller than 3.2 µm (2.0 ± 0.4 µm), which is considered clinically relevant. The results for mRNA expression of ALP, OCN, and OPN and alizarin red S staining indicated that cell differentiation and bio-mineralization were significantly inhibited by the presence of ceramic debris at all tested concentrations (P < 0.05, and the values decreased gradually with the increase of ceramic debris concentration), but the results of the CCK-8 assay showed that cell proliferation was not significantly affected (P > 0.05; there was no significant difference between the groups at 1, 3, and 5 days). In addition, the results of ELISA, RT-PCR, and western blotting demonstrated that ceramic debris significantly promoted the release of inflammatory factors, including TNF-α, IL-ß, and IL-6 (P < 0.05, and the values increased gradually with the increase of ceramic debris concentration), and also greatly reduced the mRNA expression of Smad1, Smad4, and Smad5 (the values decreased gradually with the increase of ceramic debris concentration) as well as protein expression of phosphorylated Smad1, Smad4, and Smad5. CONCLUSION: Ceramic debris may affect differentiation and bio-mineralization of MC3T3-E subclone 14 cells through the bone morphogenetic protein/Smad signaling pathway.

PCAF acetylates Runx2 and promotes osteoblast differentiation.

Osteoblasts play a crucial role in bone formation. However, the molecular mechanisms involved in osteoblast differentiation remain largely unclear. Runt-related gene 2 (Runx2) is a master transcriptional factor for osteoblast differentiation. Here we reported that p300/CBP-associated factor (PCAF) directly binds to Runx2 and acetylates Runx2, leading to an increase in its transcriptional activity. Upregulation of PCAF in MC3T3-E1 cells increases the expression of osteogenic marker genes including alkaline phosphatase (ALP), osteocalcin (Ocn), and Osteopontin (Opn), and ALP activity was stimulated as well. Consequently, the mineralization of MC3T3-E1 cells was remarkably improved by PCAF. In contrast, PCAF knockdown decreases the mRNA levels of ALP, Ocn, and Opn. ALP activity and the mineralized area were attenuated under PCAF knockdown conditions. These results indicate that PCAF is an important regulator for promoting osteoblast differentiation via acetylation modification of Runx2.

[Progress on interface membrane around prostheses].

Aseptic loosening is one of the most frequent long-term complications after joint replacement,which limits the service life of prostheses. A lot of studies have been focused on interface membranes around prostheses recently. In interface membranes, there are plenty of macrophagocytes and desmocytes,which play vital roles in osteolysis. This study gave a review of interface membrane including its formation, cellulosity, osteolysis factor, immunity reaction and turnover. The advances of interface membranes will help us to comprehend the pathogenesy and treatment of the aseptic loosening.

Polyester-degrading thermophilic actinomycetes isolated from different environment in Taiwan.

Thermophilic actinomycetes strains were isolated from various environment in Taiwan and screened for degradation of poly(ethylene succinate) (PES), poly(epsilon-caprolactone) (PCL) and/or poly(beta-hydroxybutyrate) (PHB) by the clear-zone method. Out of 341 strains of thermophilic actinomycetes, 105 isolates were PHB-degraders (30.8%), 198 isolates were PCL-decomposers (58.1%), and 99 isolates could degrade PES (29.0%). Furthermore, 77 isolates could degrade both PHB and PCL (22.6%), 35 isolates could degrade both PHB and PES (10.3%), 81 isolates could degrade both PES and PCL (23.8%) and 31 isolates could degrade the three polyesters used in this study (9.1%). Base on the morphological and chemical characteristics, these 31 isolates belonging to Actinomadura (12.9%), Microbispora (25.8%), Streptomyces (48.4%), Thermoactinomyces (9.7%) and Saccharomonospora genus (3.22%).

Production of xylooligosaccharides from xylans by extracellular xylanases from Thermobifida fusca.

Xylooligosaccharides are produced for use as a valuable food sweetener or additive. They have many beneficial biomedical and health effects. In this study, a process for producing xylooligosaccharides from lignocellulolytic agricultural waste was developed. Bagasse, corncob, wheat bran, and peanut shell were used as carbon sources for production of xylanolytic enzymes from Thermobifida fusca NTU22. When using bagasse as the carbon source, the xylanolytic enzymes that simultaneously accumulated in the broth in a 500 mL Hinton flask after 72 h of cultivation at 50 degrees C were measured as xylanase (14.0 U/mL), beta-xylosidase (74.1 mU/mL), and acetyl esterase (29.1 mU/mL). The optimum pH and temperature for xylanases were 6.0-8.0 and 70 degrees C, respectively. Six proteins with xylanase activity were identified by zymogram analysis of isoelectric focusing gel. This was followed by heat treatment at 70 degrees C for 30 min that eliminated 90% of the beta-xylosidase activity. The xylanase and acetyl esterase activities were still 100%. Two percent of xylan extracted from the bagasse was then hydrolyzed by heat-treated crude xylanase preparation at 60 degrees C, pH 7.0, for 10 h. The xylooligosaccharides that accumulated in the broth were about 23.7%. After the purification process by activated charcoal chromatography, the purity of xylooligosaccharides was 71.4%.

Amycolatopsis taiwanensis sp. nov., from soil.

An actinomycete strain (0345M-7(T)) was isolated from a soil sample from Yilan county, Taiwan. The isolate displayed substrate mycelia, upon which were borne short spore chains. The spore chains were composed of non-motile, smooth-surfaced, oval spores. Strain 0345M-7(T) had meso-diaminopimelic acid in its peptidoglycan. Whole-cell sugars were galactose, glucose, arabinose and ribose. The only phospholipid found was phosphatidylethanolamine. The predominant menaquinone was MK-9(H(4)). Mycolic acids were not detected. Major cellular fatty acids were iso-C(16 : 0) (38.1 %) and C(17 : 1) (25.4 %). The DNA G+C content of strain 0345M-7(T) was 68.9 mol%. On the basis of phenotypic and genotypic data, it is proposed that strain 0345M-7(T) (=BCRC 16802(T)=KCTC 19116(T)) should be classified as the type strain of a novel species of the genus Amycolatopsis, Amycolatopsis taiwanensis sp. nov.

Discovery of ultrahigh-T spinel-garnet granulite with pure CO2 fluid inclusions from the Altay orogenic belt, NW China.

We first report discovery of the spinel-garnet-orthopyroxene granulite with pure CO2 fluid inclusions from the Fuyun region of the late Paleozoic Altay orogenic belt in Central Asia, NW China. The rock is characterized by an assemblage of garnet, orthopyroxene, spinel, cordierite, biotite, plagioclase and quartz. Symplectites of orthopyroxene and spinel, and orthopyroxene and cordierite indicate decompression under UHT conditions. Mineral chemistry shows that the orthopyroxenes have high XMg and Al2O3 contents (up to 9.23 wt%). Biotites are enriched in TiO2 and XMg and are stable under granulite facies conditions. The garnet and quartz from the rock carry monophase fluid inclusions which show peak melting temperatures of around -56.7 degrees C, indicating a pure CO2 species being presented during the ultrahigh-T metamorphism in the Altay orogenic belt. The inclusions homogenize into a liquid phase at temperatures around 15.3-23.8 degrees C translating into CO2 densities of the order of 0.86-0.88 g/cm3. Based on preliminary mineral paragenesis, reaction textures and petrogenetic grid considerations, we infer that the rock was subjected to UHT conditions. The CO2-rich fluids were trapped during exhumation along a clockwise P-T path following isothermal decompression under UHT conditions.

Petrology, geochemistry and geodynamics of basic granulite from the Altay area, North Xinjiang, China.

The basic granulite of the Altay orogenic belt occurs as tectonic lens in the Devonian medium- to lower-grade metamorphic beds through fault contact. The Altay granulite (AG) is an amphibole plagioclase two-pyroxene granulite and is mainly composed of two pyroxenes, plagioclase, amphibole and biotite. Its melano-minerals are rich in Mg/(Mg+Fe2+), and its amphibole and biotite are rich in TiO2. The AG is rich in Mg/(Mg+Fe2+), Al2O3 and depletion of U, Th and Rb contents. The AG has moderate SigmaREE and LREE-enriched with weak positive Eu anomaly. The AG shows island-arc pattern with negative Nb, P and Ti anomalies, reflecting that formation of the AG may be associated with subduction. Geochemical and mineral composition data reflect that the protolith of the AG is calc-alkaline basalt and formed by granulite facies metamorphism having peak P-T conditions of 750 degrees C-780 degrees C and 0.6-0.7 Gpa. The AG formation underwent two stages was suggested. In the early stage of oceanic crustal subduction, calc-alkaline basalt with island-arc environment underwent granulite facies metamorphism to form the AG in deep crust, and in the late stage, the AG was thrust into the upper crust.