Enhanced Cellular Delivery of Tildipirosin by Xanthan Gum-Gelatin Composite Nanogels.

Tildipirosin has no significant inhibitory effect on intracellular bacteria because of its poor membrane permeability. To this end, tildipirosin-loaded xanthan gum-gelatin composite nanogels were innovatively prepared to improve the cellular uptake efficiency. The formation of the nanogels via interactions between the positively charged gelatin and the negatively charged xanthan gum was confirmed by powder X-ray diffraction and Fourier transform infrared. The results indicate that the optimal tildipirosin composite nanogels possessed a 3D network structure and were shaped like a uniformly dispersed ellipse, and the particle size, PDI, and ζ potential were 229.4 ± 1.5 nm, 0.26 ± 0.04, and -33.2 ± 2.2 mV, respectively. Interestingly, the nanogels exhibited gelatinase-responsive characteristics, robust cellular uptake via clathrin-mediated endocytosis, and excellent sustained release. With those pharmaceutical properties provided by xanthan gum-gelatin composite nanogels, the anti-Staphylococcus aureus activity of tildipirosin was remarkably amplified. Further, tildipirosin composite nanogels demonstrated good biocompatibility and low in vivo and in vitro toxicities. Therefore, we concluded that tildipirosin-loaded xanthan gum-gelatin composite nanogels might be employed as a potentially effective gelatinase-responsive drug delivery for intracellular bacterial infection.

Differential analysis of ovarian tissue between high and low-yielded laying hens in the late laying stage and the effect of LECT2 gene on follicular granulosa cells proliferation.

The ovaries of high-yield laying hens exhibited signs of aging beyond 400 days of age, subsequently resulting in a decline in both egg production and egg quality. Oxidative stress, characterized by an increase in the production of reactive oxygen species (ROS), stands as one of the principal processes contributing to ovarian aging. Elevated ROS levels are implicated in the induction of apoptosis in granulosa cells (GCs), provoking mitochondrial impairment, and diminishing the capacity of the antioxidant defense system. This investigation stratified laying hens into two distinct groups, predicated upon their egg production levels: high-yield hens (HH) and low-yield hens (LL). The study focused on evaluating oxidative stress markers and identifying differentially expressed genes between these two groups. The findings revealed that the LL group exhibited follicular atresia, mitochondrial disruptions, and apoptotic occurrences in ovarian GCs. Notably, ROS levels, Malondialdehyde (MDA) concentrations, and 8-hydroxy-2'-deoxyguanosine (8-OHdG) concentrations in ovarian tissue and follicular GCs were substantially higher in the HH group. Furthermore, the RNA-sequencing results unveiled differential expression of the LECT2 gene between the HH and LL groups. Consequently, an overexpression vector for the LECT2 gene was successfully constructed and introduced into GCs. The quantitative polymerase chain reaction (QPCR) analysis exhibited significant downregulation (p < 0.01) of key apoptotic genes such as Caspase-3 and C-myc and significant upregulation (p < 0.01) of BCL2 following the overexpression of the LECT2 gene in GCs. In conclusion, oxidative stress emerges as a pivotal factor influencing the laying traits of both high and low-yield laying hens. The accumulation of reactive oxygen species (ROS) within the ovaries precipitates apoptosis in GCs, subsequently leading to follicular atresia and a reduction in egg production. Furthermore, we employed RNA sequencing technology to examine the ovarian matrix tissue in high and low laying hens during the late phase of egg laying. Our analysis revealed a substantial upregulation of the LECT2 gene in the ovarian matrix tissue of high laying hens. This observation implies that the LECT2 gene exerts a pivotal influence on driving the proliferation and differentiation of follicular GCs, thereby exerting a crucial regulatory role in follicular development.

Chrysanthemum sporopollenin: A novel vaccine delivery system for nasal mucosal immunity.

Objective: Mucosal immunization was an effective defender against pathogens. Nasal vaccines could activate both systemic and mucosal immunity to trigger protective immune responses. However, due to the weak immunogenicity of nasal vaccines and the lack of appropriate antigen carriers, very few nasal vaccines have been clinically approved for human use, which was a major barrier to the development of nasal vaccines. Plant-derived adjuvants are promising candidates for vaccine delivery systems due to their relatively safe immunogenic properties. In particular, the distinctive structure of pollen was beneficial to the stability and retention of antigen in the nasal mucosa. Methods: Herein, a novel wild-type chrysanthemum sporopollenin vaccine delivery system loaded with a w/o/w emulsion containing squalane and protein antigen was fabricated. The unique internal cavities and the rigid external walls within the sporopollenin skeleton construction could preserve and stabilize the inner proteins. The external morphological characteristics were suitable for nasal mucosal administration with high adhesion and retention. Results: Secretory IgA antibodies in the nasal mucosa can be induced by the w/o/w emulsion with the chrysanthemum sporopollenin vaccine delivery system. Moreover, the nasal adjuvants produce a stronger humoral response (IgA and IgG) compared to squalene emulsion adjuvant. Mucosal adjuvant benefited primarily from prolongation of antigens in the nasal cavity, improvement of antigen penetration in the submucosa and promotion of CD8+ T cells in spleen. Disccusion: Based on effective delivering both the adjuvant and the antigen, the increase of protein antigen stability and the realization of mucosal retention, the chrysanthemum sporopollenin vaccine delivery system has the potential to be a promising adjuvant platform. This work provide a novel idea for the fabrication of protein-mucosal delivery vaccine.

Borneol-driven meningeal lymphatic drainage clears amyloid-ß peptide to attenuate Alzheimer-like phenotype in mice.

Rationale: The accumulation and clearance of amyloid-ß (Aß) peptides play a crucial role in the pathogenesis of Alzheimer's disease (AD). The (re)discovery of meningeal lymphatic vessels in recent years has focused attention on the lymphatic clearance of Aß and has become a promising therapeutic target for such diseases. However, there is a lack of small molecular compounds that could clearly regulate meningeal lymphatic drainage to remove Aß from the brain. Methods: We investigated the effect of borneol on meningeal lymphatic clearance of macromolecules with different molecular weights (including Aß) in the brain. To further investigate the mechanism of borneol regulating meningeal lymphatic drainage, immunofluorescence staining, western blotting, ELISA, RT-qPCR, and Nitric Oxide assay kits were used. The cognitive function of AD mice after borneol treatment was evaluated using two behavioral tests: open field (OF) and Morris water maze (MWM). Results: This study discovered that borneol could accelerate the lymphatic clearance of Aß from the brain by enhancing meningeal lymphatic drainage. Preliminary mechanism analysis revealed that borneol could improve the permeability and inner diameter of lymphatic vessels, allowing macromolecules to drain into the cervical lymph nodes (CLNS) and then be transported to the lymphatic circulation. To speed up the clearance of macromolecules, borneol also stimulated lymphatic constriction by lowering the level of nitric oxide in the meninges. In addition, borneol stimulated lymphangiogenesis by increasing the levels of FOXC2, VEGFC, and LYVE-1 in the meninges, which promoted the clearance rates of macromolecules. Borneol improved meningeal lymphatic clearance not only for Aß but also for other macromolecular polymers (molecular weight in the range of 2 KD - 45 KD. Borneol ameliorated cognitive deficits and alleviated brain Aß burden in Aß-injected mice. Conclusions: Our findings not only provide a strategy to regulate lymphatic clearance pathways of macromolecules in the brain, but also new targets and ideas for treating neurodegenerative diseases like AD. Furthermore, our findings indicate that borneol is a promising therapeutic drug for AD.

Menthol nanoliposomes enhanced anti-tumor immunotherapy by increasing lymph node homing of dendritic cell vaccines.

Menthol, a cyclic terpene alcohol, plays a critical role in overcoming the blood-brain barrier and stratum corneum barrier. Herein, we innovatively propose a menthol nanoliposome (Men-nanoLips) that can dramatically increase lymph node accumulation of the dendritic cell (DC)-based anti-tumor vaccines. Specifically, Men-nanoLips efficiently enhanced lymphatic endothelial cell (EC) barrier permeability by reducing the expression of tight junction proteins. And interestingly, Men-nanoLips not only up-regulated the expression of CCR7 in DCs but also increased the secretion of CCL21 in lymphatic ECs. Moreover, Men-nanoLips promoted DC vaccine maturation as evidenced by increasing the expression of costimulatory molecules and up-regulating the pseudopodia-like protein. With those complementary mechanisms provided by Men-nanoLips, the number of the B16 whole-tumor cell lysate-loaded DCs that target the draining LN enhanced remarkably and significantly boosted the treatment efficacy of DC anti-tumor vaccines. Therefore, we concluded that Men-nanoLips could be instructive for increasing LN homing of DC vaccines.

Ultra-high photoactive thiadiazolo[3,4-g]quinoxaline nanoparticles with active-targeting capability for deep photodynamic therapy.

Improving the effective treatment depth of photodynamic therapy (PDT) is an important issue to resolve for its clinical application. In this study, a new biocompatible photosensitizer (PS), namely TQs-PEG4, based on thiadiazolo[3,4-g]quinoxaline (TQ) with ultra-high photoactive property is designed and synthesized. TQs-PEG4 possesses an ultra-high singlet oxygen quantum yield (ΦΔ = 1.04). After encapsulating it with a biodegradable copolymer (DSPE-mPEG2000-cRGD), well distributed organic TQs-PEG4 nanoparticles (NPs) are formed with good water dispersity and excellent active tumor-targeting property. In vitro PDT experiments reveal that TQs-PEG4 NPs present excellent phototoxicities towards different cancer cell lines with an ultra-low dosage (<0.3 µg mL-1). TQs-PEG4 NP mediated PDT significantly inhibited tumor growth even when the tumor was covered with a 6 mm thick piece of pork tissue under 660 nm laser irradiation. Both the histological analysis and biochemical testing demonstrated the good biosafety of TQs-PEG4 NPs towards mice. This study not only develops an ultra-high photoactive organic PS, TQs-PEG4, but also proves the great potential of TQs-PEG4 NPs for application in deep PDT.

Oxygen-Based Nanocarriers to Modulate Tumor Hypoxia for Ameliorated Anti-Tumor Therapy: Fabrications, Properties, and Future Directions.

Over the past five years, oxygen-based nanocarriers (NCs) to boost anti-tumor therapy attracted tremendous attention from basic research and clinical practice. Indeed, tumor hypoxia, caused by elevated proliferative activity and dysfunctional vasculature, is directly responsible for the less effectiveness or ineffective of many conventional therapeutic modalities. Undeniably, oxygen-generating NCs and oxygen-carrying NCs can increase oxygen concentration in the hypoxic area of tumors and have also been shown to have the ability to decrease the expression of drug efflux pumps (e.g., P-gp); to increase uptake by tumor cells; to facilitate the generation of cytotoxic reactive oxide species (ROS); and to evoke systematic anti-tumor immune responses. However, there are still many challenges and limitations that need to be further improved. In this review, we first discussed the mechanisms of tumor hypoxia and how it severely restricts the therapeutic efficacy of clinical treatments. Then an up-to-date account of recent progress in the fabrications of oxygen-generating NCs and oxygen-carrying NCs are systematically introduced. The improved physicochemical and surface properties of hypoxia alleviating NCs for increasing the targeting ability to hypoxic cells are also elaborated with special attention to the latest nano-technologies. Finally, the future directions of these NCs, especially towards clinical translation, are proposed. Therefore, we expect to provide some valued enlightenments and proposals in engineering more effective oxygen-based NCs in this promising field in this comprehensive overview.

Identification of stress-responsive transcription factors with protein-bound Escherichia coli genomic DNA libraries.

Bacteria promoters along with operators are crucial elements in the control of gene expression in microbes in response to environmental stress changes. A genome-wide promoter DNA regulatory library is in demand to be developed for a microbe reporter method to monitor the existence of any given environmental stress substance. In this study, we utilized Escherichia coli (E. coli) as a model system for the preparation of both cell lysates and genomic DNA fragments. Through enriching protein-bound DNA fragments to construct luciferase reporter libraries, we found that, of 280 clones collected and sequenced, 131 clones contained either the promoter-35 and -10 conservative sequences and/or an operator transcription factor binding sites (TFBS) region. To demonstrate the functionality of the identified clones, five of 131 clones containing LexA binding sequence have been demonstrated to be induced in response to mitomycin C treatment. To evaluate our libraries as a functional screening library, 80 randomly picked up clones were cultured and treated with and without MMC, where two clones were shown to have greater than twofold induction. In addition, two arsenite-responsive clones were identified from 90 clones, one having the well-known ArsR and another having the osmotically inducible lipoprotein (OsmE1). The newly discovered osmE1 has been quantitatively validated to be induced by arsenite treatment with real-time PCR in a dose response and time course manner. This enriching protein-bound DNA luciferase reporter libraries and functional screening facilitate the identification of stress-responsive transcriptional factors in microbes. We developed functional libraries containing E. coli genomic-wide protein-bound DNA as enhancers/operators to regulate downstream luciferase in response to stress.

A water-soluble pyrazino[2,3-g]quinoxaline photosensitizer for high-efficiency one- and two-photon excited bioimaging and photodynamic therapy.

A water-soluble photosensitizer, PQs-PEG5, based on a novel pyrazino[2,3-g]quinoxaline (PQ) prototype compound (PQs-5), was designed and synthesized. It has strong linear absorption within 400-600 nm and a desirable two-photon absorption cross section (100-1290 GM) within 740-1000 nm, and it exhibits both a high fluorescence quantum yield (∼0.55) and singlet oxygen quantum yield (∼0.47). By carrying out in vitro cell imaging and photodynamic therapy (PDT) experiments on PQs-PEG5 with HeLa and 4T1 cells, it was demonstrated that PQs-PEG5 can present outstanding bioimaging performance and PDT efficacies simultaneously. Under one-photon excitation (1PE) of a 635 nm diode laser (60 mW cm-2, 5 min) or two-photon excitation (2PE) of an 820 nm fs laser (100 mW, 80 MHz, 140 fs, 5 min), PQs-PEG5 (10 µM) could cause HeLa and 4T1 cell death effectively, which indicated its great potential as a novel photosensitizer for both 1PE- and 2PE-PDT applications.

Theileria ovis (Piroplasmida: Theileriidae) Detected in Melophagus ovinus (Diptera: Hippoboscoidea) and Ornithodoros lahorensis (Ixodida: Argasidae) Removed From Sheep in Xinjiang, China.

Theileria spp. are tick-transmitted, intracellular apicomplexan protozoan parasites that infect a wide range of animals and, as such, can cause significant economic losses. The aim of the present study was to detect and analyze apicomplexan parasites from two different ectoparasites that were collected from Xinjiang Uygur Autonomous Region, China. The PCR-based detection of 18S rRNA indicated that Ornithodoros lahorensis specimens from Kashgar, Xinjiang, and Aksu were positive for Theileria spp., as were Melophagus ovinus specimens from Aksu. Meanwhile, phylogenetic analysis, based on the 18S rRNA gene sequences, revealed that the four amplified Theileria sequences could be attributed to T. ovis. To the best of our knowledge, this is the first study to report the detection of T. ovis DNA in M. ovinus and the first molecular identification study to confirm the detection of T. ovis in O. lahorensis in China. Accordingly, the present study extends the known distribution of T. ovis.

Novel mitochondrial-targeted thiadiazolo[3,4-g]quinoxaline dyes as efficient photosensitizers for ultra-low dose operable photodynamic therapy.

Two novel thiadiazolo[3,4-g]quinoxaline (TQ) photosensitizers (PSs), TQs-3 and TQs-4, were designed and synthesized. Both of them presented ultra-high singlet oxygen quantum yields under red light irradiation. By carrying out in vitro photodynamic therapy (PDT) experiments using TQs-4 loaded nanoparticles (TQs-4 NPs) to treat three kinds of tumor cell lines: 4T1, HeLa and MCF-7 cells, it was demonstrated that TQs-4 NPs had outstanding PDT efficacies. An ultra-low dose of TQs-4 (0.14 µg mL-1) can realize the death of more than 90% HeLa cells (635 nm, 60 mW cm-2, 10 min), which indicated that TQs-4 show promising potential as a novel PS for PDT applications.

PEGylated gold nanorods are not cytotoxic to human endothelial cells but affect kruppel-like factor signaling pathway.

Gold (Au) nanomaterials (NMs), particularly those with PEG surface functionalization, are generally considered to be biocompatible for biomedical applications due to relatively low cytotoxicity. Herein, we investigated the toxicity of PEGylated Au nanorods (NRs) to human umbilical vein endothelial cells (HUVECs), a commonly used in vitro model for human endothelium. We found a previously unknown effect that up to 10 µg/mL Au NRs, albeit not cytotoxic, decreased the mRNA and protein levels of kruppel-like factor 2 (KLF2), a transcription factor with well-documented vasoprotective effects. The results from PCR array showed that a number of genes associated with risk of cardiovascular diseases were altered by Au NRs, and several genes are downstream genes of KLF2 according to ingenuity pathway analysis (IPA). These effects could be observed with or without the presence of inflammatory stimuli lipopolysaccharide (LPS), which suggests a pre-existing inflammatory state is not required for Au NRs to alter KLF2 signaling pathway. We further identified that Au NRs significantly decreased eNOS mRNA/p-eNOS proteins as well as increased MCP-1 mRNA/sMCP-1 release, which are targets of KLF2. Combined, our data revealed a novel pathway that PEGylated Au NPs at non-cytotoxic concentrations might alter KLF leading to the increase of risk of cardiovascular diseases in human endothelial cells. Given the importance of KLF in vascular homeostasis, our data indicate that it is necessary to evaluate the influence of engineered NPs to KLF signaling pathways, especially for NPs with biomedical uses.

First report of border disease virus in Melophagus ovinus (sheep ked) collected in Xinjiang, China.

Melophagus ovinus (sheep ked) is a blood-sucking ectoparasite that is parasitic primarily on sheep. It is widely distributed in different geographical regions worldwide. In China, it has been mainly found in Xinjiang, Gansu, and Tibet in recent years. In addition to causing direct damage to the animal hosts, M. ovinus also carries pathogens and serves as a vector for disease transmission. Border disease virus (BDV) is a positive-sense, single-stranded RNA pestivirus that mainly infects and causes border disease (BD) in sheep and goats worldwide. Since 2012, this disease has been reported in 4 provinces in China. In the present study, we investigated the presence of BDV in M. ovinus from Xinjiang and Gansu. Frozen M. ovinus collected during 2017 and 2018 from Xinjiang and Gansu and preserved in our laboratory were studied. First, total RNA of M. ovinus was extracted, followed by reverse transcription, PCR (RT-PCR) amplification of the 5'-UTR of BDV, and sequencing of the amplified products. Finally, the sequencing results were analyzed using DNAStar, MEGA 5.0 molecular biology software, and the BLAST online platform. The results from RT-PCR and sequencing analyses showed that among the samples included in the study, only the M. ovinus collected from Qinghe County in Alta, Xinjiang in 2018 tested positive for BDV. BLAST analysis showed that the viral strain with the most similar nucleotide identity to the sequence of the China/BDV/2018 fragment was the goat-derived BDV strain AH12-02 collected in Anhui, China, in 2012. A phylogenetic-tree analysis showed the strain to exhibit a BDV-3 genotype. This is the first report globally on BDV detected in M. ovinus and is also the first report of BDV discovered in Xinjiang, China. This study reconfirms the presence of BDV in China.

Contribution of nonconsensus base pairs within ArsR binding sequences toward ArsR-DNA binding and arsenic-mediated transcriptional induction.

BACKGROUND: A transcriptional reporter is the key component in bacterial biosensors which are employed to monitor the induction or repression of a reporter gene corresponding to environmental change. Interaction of a transcription factor with its consensus sequence generated by using a position weight matrix (PWM) model is crucial for its sensitivity of the reporter. However, recent studies suggest that PWM model based on independent contribution of individual consensus base pairs to protein interaction is often insufficient to explain complex regulation, such as the effect of nonconsensus sequences on the protein-DNA binding affinity. In the present study, we employed a simpler prokaryotic arsenic repressor (ArsR) regulation system to access the protein-DNA recognition. Contribution of nonconsensus base pairs within ArsR binding sequences toward ArsR-DNA binding and arsenic-mediated transcriptional induction was studied. RESULTS: We constructed a series of arsenic responsive reporters, each comprising two copies of the ArsR binding sequences from different resources. We found that high arsenic-mediated induction specifically requires the binding sequence from Escherichia coli to be placed at the first binding sequence; however, no such preference was observed for the second binding sequence, which could be from Acidithiobacillus ferrooxidans, plasmid R773, Synechococcus, or a core binding sequence of arsR. By creating a series of reporters differed at the nonconsensus base pairs of the second binding sequence, we observed that some constructs bound weakly while others strongly to ArsR. Most interestingly, although a number of these reporters showed similar binding affinity to ArsR, their arsenic-dependent induction differed significantly. CONCLUSIONS: The results indicated that nonconsensus base pairs could have profound influence on protein binding and may also modulate post-binding function. These findings provide new insights into the complex regulation of gene expression and facilitate the development of transcriptional reporter-based biosensors.

The toxicity of multi-walled carbon nanotubes (MWCNTs) to human endothelial cells: The influence of diameters of MWCNTs.

The biological applications of multi-walled carbon nanotubes (MWCNTs) may lead to their exposure to human blood vessels, but the influence of their physicochemical properties on toxicity to endothelial cells is incompletely known. Here, human umbilical vein endothelial cells (HUVECs) were exposed to three commercially available MWCNTs, namely XFM4, XFM22, and XFM34 (diameters XFM4 < XFM22 < XFM34), to understand the possible role of their diameter on toxicity. Based on the same mass concentration, XFM4 induced significantly higher level of cytotoxicity than the other two MWCNTs, and HUVECs internalized more XFM4. Cytokine release, monocyte adhesion, and intracellular reactive oxygen species levels were significantly induced only after XFM4 treatment. The exposure to XFM4 significantly reduced the expression of autophagic genes autophagy-related 7 (ATG7), autophagy-related 12 (ATG12), and beclin 1 (BECN1) and increased the expression of endoplasmic reticulum (ER) stress genes DNA damage inducible transcript 3 (DDIT3) and X-box binding protein 1 spliced (XBP-1s). Moreover, the modulation of autophagy-ER stress by chemicals resulted in a significant increase in the cytotoxicity of XFM4 but had minimal impact on the cytotoxicity of XFM34. These data indicate that the diameter of MWCNTs may influence their toxicity to HUVECs, probably through autophagy dysfunction and ER stress.

Palmitate enhanced the cytotoxicity of ZnO nanomaterials possibly by promoting endoplasmic reticulum stress.

We recently synthesized ZnO nanomaterials (denoted as ZnO nanorods [NRs] and Mini-NRs) and suggested that their cytotoxicity could be related with the activation of endoplasmic reticulum (ER) stress apoptosis. However, in a complex biological microenvironment, the ER stress-apoptosis pathway could also be modulated by biological molecules, such as free fatty acids, leading to unpredicted biological effects. In this study, we investigated the combined toxicity of ZnO NRs/Mini-NRs and palmitate (PA) to THP-1 macrophages. PA influenced the zeta potential and solubility of ZnO NRs and ZnO Mini-NRs in water, which indicated a change of colloidal stability. Exposure to ZnO NRs and Mini-NRs dose-dependent decreased cellular viability and release of soluble monocyte chemotactic protein 1 (sMCP-1), and these effects were significantly promoted with the presence of PA. However, ZnO NR- and Mini-NR-induced intracellular Zn ions or reactive oxygen species were not significantly affected by PA. ZnO NRs and ZnO Mini-NRs significantly promoted the expression of ER stress genes HSPA5, DDIT3, XBP-1s and apoptotic gene CASP3, whereas PA also modestly promoted the expression of HSPA5, DDIT3 and CASP3. Interestingly, the ER stress inducer thapsigargin showed a similar effect as PA to promote the cytotoxicity of ZnO NRs and ZnO Mini-NRs. It is suggested that PA might promote the cytotoxicity of ZnO NRs and ZnO Mini-NRs possibly by promoting ER stress.

Molecular Identification of Bartonella melophagi and Wolbachia Supergroup F from Sheep Keds in Xinjiang, China.

To confirm that Bartonella and Wolbachia were carried by sheep keds (Melophagus ovinus) in southern Xinjiang of China, 17 M. ovinus samples, which were collected in Aksu Prefecture, Xinjiang, were randomly selected. In this study, the Bartonella gltA and Wolbachia 16S rRNA gene were amplified through conventional PCR and the sequence of those amplified products, were analyzed. The results demonstrated that Bartonella was carried by all of the 17 sheep keds and Wolbachia was carried by 15 out of them. Bartonella was identified as B. melophagi. Three strains of Wolbachia were supergroup F and 1 strain has not been confirmed yet. It is the first report about Wolbachia supergroup F was found in sheep keds and provided the molecular evidence that B. melophagi and Wolbachia supergroup F were carried by sheep keds in Aksu Prefecture of southern Xinjiang, China. The 2 pathogens were found in sheep keds around Taklimakan Desert for the first time.

Copy number of ArsR reporter plasmid determines its arsenite response and metal specificity.

The key component in bacteria-based biosensors is a transcriptional reporter employed to monitor induction or repression of a reporter gene corresponding to environmental change. In this study, we made a series of reporters in order to achieve highly sensitive detection of arsenite. From these reporters, two biosensors were developed by transformation of Escherichia coli DH5α with pLHPars9 and pLLPars9, consisting of either a high or low copy number plasmid, along with common elements of ArsR-luciferase fusion and addition of two binding sequences, one each from E. coli and Acidithiobacillus ferrooxidans chromosome, in front of the R773 ArsR operon. Both of them were highly sensitive to arsenite, with a low detection limit of 0.04 µM arsenite (~ 5 µg/L). They showed a wide dynamic range of detection up to 50 µM using high copy number pLHPars9 and 100 µM using low copy number pLLPars9. Significantly, they differ in metal specificity, pLLPars9 more specific to arsenite, while pLHPars9 to both arsenite and antimonite. The only difference between pLHPars9 and pLLPars9 is their copy numbers of plasmid and corresponding ratios of ArsR to its binding promoter/operator sequence. Therefore, we propose a working model in which DNA bound-ArsR is different from its free form in metal specificity.

Toxicity of ZnO nanoparticles (NPs) with or without hydrophobic surface coating to THP-1 macrophages: interactions with BSA or oleate-BSA.

It is recently shown that biological macromolecules in food could interact with nanoparticles (NPs) and consequently change the biological effects of NPs. In this study, the interactions between ZnO NPs with or without hydrophobic surface coating and bovine serum albumin (BSA) or oleate (OA) complexed to BSA (OA-BSA) were assessed. Atomic force microscope (AFM) showed topographic changes of both types of NPs by BSA or OA-BSA, which could indicate the formation of protein corona. ZnO NPs showed negative Zeta potential, which was slightly decreased by BSA or OA-BSA, with OA-BSA being more effective. The UV-Vis was increased, whereas the fluorescence and synchronous fluorescence was decreased by the presence of ZnO NPs. Exposure to both types of ZnO NPs was associated with cytotoxicity to THP-1 macrophages, which was equally mitigated by BSA or OA-BSA associated with decreased cellular Zn elements. Exposure to ZnO NPs was associated with decreased release of cytokines, which was not affected by BSA or OA-BSA. In combination, the results from this study suggested that both BSA and OA-BSA could be adsorbed to ZnO NPs regardless of hydrophobic surface coating, which reduced the cytotoxicity of NPs to macrophages probably due to reduced association between NPs and cells. BSA and OA-BSA equally protected THP-1 macrophages from ZnO NP exposure, which might indicate that complexation to OA did not compromise the cytoprotective effects of BSA. These data might also indicate the complex interaction between NPs and biological macromolecules as food components, which should be considered for future nanotoxicological studies.

First report of Anaplasma ovis in pupal and adult Melophagus ovinus (sheep ked) collected in South Xinjiang, China.

BACKGROUND: Melophagus ovinus (sheep ked) is a blood-feeding ectoparasite that belongs to the family Hippoboscidae (Diptera: Hippoboscoidea) and mainly parasitizes sheep. The life-cycle of M. ovinus consists of three stages: larva, pupa and adult. It has a worldwide distribution and has been found in four provinces of China, especially South Xinjiang. In addition to causing direct damage to animal hosts, M. ovinus serves as a vector for disease transmission. In this study, our aim was to investigate the presence of Anaplasma spp. in pupal and adult M. ovinus. METHODS: A total of 93 specimens (including eight pupal specimens) of M. ovinus collected in South Xinjiang were selected for isolation of genomic DNA, followed by PCR amplification and sequencing of the msp4 gene of Anaplasma spp. The sequences were analyzed in MEGA 7.0 software and via online BLAST. RESULTS: PCR and sequencing results showed that all the specimens collected in 2013 were free of Anaplasma spp., whereas three and 25 specimens (including five pupal specimens) collected in 2016 and 2017, respectively, tested positive for Anaplasma spp. The analysis of 24 msp4 gene sequences (from four pupal specimens) confirmed the presence of A. ovis in M. ovinus specimens collected in South Xinjiang, China. The detected A. ovis isolates belong to Genotypes II and III. CONCLUSIONS: To the best of our knowledge, this is the first report of the detection of A. ovis DNA in pupal M. ovinus, confirming the vertical transmission of A. ovis in M. ovinus and the potential of M. ovinus to serve as a vector for A. ovis.