E. hellem Ser/Thr protein phosphatase PP1 targets the DC MAPK pathway and impairs immune functions.

Microsporidia are difficult to be completely eliminated once infected, and the persistence disrupts host cell functions. Here in this study, we aimed to elucidate the impairing effects and consequences of microsporidia on host DCs. Enterocytozoon hellem, one of the most commonly diagnosed zoonotic microsporidia species, was applied. In vivo models demonstrated that E. hellem-infected mice were more susceptible to further pathogenic challenges, and DCs were identified as the most affected groups of cells. In vitro assays revealed that E. hellem infection impaired DCs' immune functions, reflected by down-regulated cytokine expressions, lower extent of maturation, phagocytosis ability, and antigen presentations. E. hellem infection also detained DCs' potencies to prime and stimulate T cells; therefore, host immunities were disrupted. We found that E. hellem Ser/Thr protein phosphatase PP1 directly interacts with host p38α (MAPK14) to manipulate the p38α(MAPK14)/NFAT5 axis of the MAPK pathway. Our study is the first to elucidate the molecular mechanisms of the impairing effects of microsporidia on host DCs' immune functions. The emergence of microsporidiosis may be of great threat to public health.

Microsporidia persistence in host impairs epithelial barriers and increases chances of inflammatory bowel disease.

Microsporidia are intracellular fungus-like pathogens and the infection symptoms include recurrent diarrhea and systematic inflammations. The major infection route of microsporidia is the digestive tract. Since microsporidia are hard to fully eliminate, the interactions and persistence of the pathogen within epithelium may modulate host susceptibility to digestive disorders. In this study, both in vitro and in vivo infection models were applied. The alterations of epithelial barrier integrity, permeability, and tight junction proteins after microsporidia infection were assessed on MDCK/Caco-2 monolayers. The fecal intestinal microbiota and tissue alterations after microsporidia infection were assessed on C57BL/6 mice. Moreover, the susceptibility to develop dextran sulfate sodium (DSS)-induced inflammatory bowel diseases (IBDs) was also analyzed by the murine infection model. The results demonstrated that microsporidia infection increased epithelium permeability, weakened wound healing capability, and destructed tight junction protein zonula occludens-1. Microsporidia infection also dysregulates intestinal microbiota. These impairing effects of microsporidia increased host vulnerability to develop enteritis as shown by the murine model of DSS-induced IBD. Our study is the first to elucidate molecular mechanisms of the damaging effects of microsporidia on host epithelium and pointed out the cryptic threats of latent microsporidia infection to public health as reflected by the increased chances of developing more severe diseases.IMPORTANCEMicrosporidia are widely present in nature and usually cause latent and persistent infections in hosts. Given the fact that the digestive tract is the major infection route, it is of great importance to explore the consequences of microsporidia infection on the intestinal epithelial barrier and the risks to the host. In this study, we demonstrated the destructing effects of microsporidium infection on epithelial barriers manifested as increased epithelial permeability, weakened wound healing ability, and disrupted tight junctions. Moreover, microsporidia made the host more susceptible to dextran sulfate sodium-induced inflammatory bowel disease. These findings provide new evidence for us to better understand and develop novel strategies for microsporidia prevention and disease control.

Encephalitozoon hellem Infection Promotes Monocytes Extravasation.

Background: Microsporidia are a group of obligated intracellular fungus pathogens. Monocytes and the derivative macrophages are among the most important players in host immunity. The invasion of microsporidia may significantly affect the monocytes maturation and extravasation processes. Methods: We utilized a previously established microsporidia infection murine model to investigate the influences of microsporidia Encephalitozoon hellem (E. hellem) infection on monocyte maturation, releasing into the circulation and extravasation to the inflammation site. Flow cytometry and qPCR analysis were used to compare the monocytes and derivative macrophages isolated from bone marrow, peripheral blood and tissues of E. hellem-infected and control mice. Results: The results showed that the pro-inflammatory group of CD11b+Ly-6C+ monocytes are promoted in E. hellem-infected mice. Interestingly, the percentage of Ly-6C+ monocytes from E. hellem-infected mice are significantly lower in peripheral blood while significantly higher in the inflamed small intestine, together with up-regulated ratio of F4/80 macrophage in small intestine as well. Conclusions: Our findings demonstrated that E. hellem infection leads to promoted monocytes maturation in bone marrow, up-regulation of extravasation from peripheral blood to inflammation site and maturation into macrophages. Our study is the first systematic analysis of monocytes maturation and trafficking during microsporidia infection, and will provide better understanding of the pathogen-host interactions.

First Report on Megaselia scalaris Loew (Diptera: Phoridae) Infestation of the Invasive Pest Spodoptera frugiperda Smith (Lepidoptera: Noctuidae) in China.

The invasive pest Spodoptera frugiperda first emerged in China in January 2019 and has, to date, migrated to 29 provinces and municipalities in China, causing heavy crop damage in large areas. As a response to this invasive species from the environment, some indigenous natural enemies have been discovered and reported after S. frugiperda invasion. In this paper, parasitic flies were collected and identified from S. frugiperda collected in the Yunnan, Guangxi, and Henan provinces and the Chongqing municipality in China. By using both conventional and molecular approaches, we were able to show that all the parasitic flies of S. frugiperda identified in the four regions were Megaselia. scalaris, and that they attacked the pest larvae and pupae. This is the first report on an indigenous Chinese Megaselia species that has parasitic ability against the invasive pest S. frugiperda, potentially providing new ideas for pest control in China.

Characterization of a Murine Model for Encephalitozoon hellem Infection after Dexamethasone Immunosuppression.

BACKGROUND: Encephalitozoon hellem (E. hellem) belongs to a group of opportunistic pathogens called microsporidia. Microsporidia infection symptoms vary and include diarrhea, ocular disorders and systemic inflammations. Traditionally, immunodeficient animals were used to study microsporidia infection. To overcome the difficulties in maintenance and operation using immunodeficient mice, and to better mimic natural occurring microsporidia infection, this study aims to develop a pharmacologically immunosuppressed murine model of E. hellem infection. METHODS: Wild-type C57BL/6 mice were immunosuppressed with dexamethasone (Dex) and then E. hellem spores were inoculated into the mice intraperitoneally. Control groups were the Dex-immunosuppressed but noninoculated mice, and the Dex-immunosuppressed then lipopolysaccharide (LPS)-treated mice. Mice body weights were monitored and all animals were sacrificed at the 15th day after inoculation. Tissue fragments and immune cells were collected and processed. RESULTS: Histopathological analysis demonstrated that E. hellem inoculation resulted in a disseminated nonlethal infection. Interestingly, E. hellem infection desensitized the innate immunity of the host, as shown by cytokine expressions and dendritic cell maturation. We also found that E. hellem infection greatly altered the composition of host gut microbiota. (4) Conclusions: Dex-immunosuppressed mice provide a useful tool for study microsporidiosis and the interactions between microsporidia and host immunity.

Proliferation characteristics of the intracellular microsporidian pathogen Nosema bombycis in congenitally infected embryos.

Nosema bombycis is an obligate intracellular pathogen that can be transmitted vertically from infected females to eggs, resulting in congenital infections in embryos. Here we investigated the proliferation characteristics of N. bombycis in silkworm embryos using a histopathological approach and deep RNA sequencing. We found that N. bombycis proliferated mainly around yolk granules at the early stage of the embryonic development, 1-2 days post oviposition (dpo). At 4-6 dpo, a portion of N. bombycis in different stages adjacent to the embryo were packaged into the newly formed intestinal lumen, while the remaining parasites continued to proliferate around yolk granules. In the newly hatched larvae (9 dpo), the newly formed spores accumulated in the gut lumen and immediately were released into the environment via the faeces. Transcriptional profiling of N. bombycis further confirmed multiplication of N. bombycis throughout every stage of embryonic development. Additionally, the increased transcriptional level of spore wall proteins and polar tube proteins from 4 dpo indicated an active formation of mature spores. Taken together, our results have provided a characterization of the proliferation of this intracellular microsporidian pathogen in congenitally infected embryos leading to vertical transmission.

Development of a nucleic acid lateral flow strip for rapid, visual detection of Nosema bombycis in silkworm eggs.

Nosema bombycis, the pathogen of silkworm pébrine, causes enormous economic losses to sericulture. As such, quarantine of commercial silkworm eggs represents an important safeguard to the silkworm industry. Here, we established a user-friendly detection system based on a nucleic acid lateral flow strip (NAFLS) that combines polymerase chain reaction (PCR) and a colloidal gold strip. PCR primers were designed based on the sequence of LSU rDNA of N. bombycis and has favourable specificity for common microsporidian isolates in silkworms. The forward and reverse primers were labeled on the 5' end with biotin and carboxyfluorescein (FAM), respectively. Genomic DNA was extracted from egg samples and was used as a template for PCR, followed by subsequent detection by NALFS. The detection limit of purified N. bombycis genomic DNA was 1 pg, 100× more sensitive than that of agarose gel electrophoresis (AGE). Furthermore, the sensitivity of detection of simulated "infected" silkworm eggs was 10-100× higher than that of AGE. NALFS detected infection in 27 of 29 samples of silkworm eggs oviposited by female moths infected in lab; ≥2% infected eggs per batch are detected as positive, while ≥40% infected eggs per batch are required for detection by AGE. Collectively, NALFS is easy to use and has great potential for widespread use in the detection of N. bombycis in silkworm egg production.

[Value of virtual endoscopy in diagnosis and treatment for hydrocephalus after failed shunt].

OBJECTIVE: To investigate the value of virtual endoscopy in diagnosis and treatment of hydrocephalus after failed shunt. METHODS: we performed magnetic resonance imaging (MRI) 3D-SPACE sequence scanning for 76 patients suffering from hydrocephalus after failed shunt. Based on the DICOM data of 3D-SPACE sequence, we three-dimensionally reconstructed the ventricles and simulated virtual endoscopy with 3D-Slicer software. Surgical strategy was made after evaluation of virtual endoscopy images. RESULTS: Virtual endoscopy based on 3D-SPACE images was helpful to depict the aqueduct or foramen of Monro obstructions, and confirm the reason of shunt failure. Finally, there were 56 hydrocephalus patients (73.7%) were treated with endoscopic assisted third ventricle fistula (ETV), without shunt catheter insertion. CONCLUSION: For patients suffering from shunt failure of hydrocephalus, virtual endoscopy has high diagnostic yield rate for providing more detailed anatomical information than conventional MR imaging. Hence, this advanced imaging method is helpful for surgical treatment strategy decision making.

[Combined application of neuroendoscope and laparoscope in ventriculo-peritoneal shunt for treatment of communicating hydrocephalus].

OBJECTIVE: To summarize the clinical efficacy of ventriculo-peritoneal shunt (VPS) assisted by neuroendoscopy and laparoscopy for treatment of communicating hydrocephalus. METHODS: From January 2010 to January 2014, 209 cases (male 93, female 116) who suffered communicating hydrocephalus performed VPS with neuroendoscopy and laparoscopy in Department of Neurosurgery of People's Liberation Army General Hospital. The age of the patients were from 7 months to 79 years (mean 38.1 years), average duration were 20 days to 4 years (mean (2.4 ± 0.7) months). Neuroendoscopy and laparoscopy were used to help respectively to place shunt catheter to better position, both in the ventricle and peritoneal cavity. The effect of subsequent shunt system survival was analyzed with Kaplan-Meier survival analysis. RESULTS: There were 209 patients received 255 times of VPS. All operations were successfully completed. No craniotomy or open operation were needed for technical-related complications. Forty-six revisions were performed in all patients. After the operation, 203 patients with hydrocephalus improved at different level after surgery. Thirteen cases occurred intracranial hypotension syndrome and improved after the pressure adjusted. All patients were followed up for 1 month to 4 years, with a median follow-up time of 2.1 years, while the shunt system efficiencies were 91.0%, 86.7%, 83.9% and 82.0% respectively from the end of the 1st year to the end of the 4th year. CONCLUSIONS: For VPS, neuroendoscopy and laparoscopy can respectively help to place shunt catheter to better position, both in the ventricle and peritoneal cavity. Hence, the combination of these two modalities can reduce the failure rate of shunt catheter insertion and has significant impact on shunt system survival.

[Endoscopic treatment strategy of obstructive hydrocephalus induced by pineal region tumors].

OBJECTIVE: To explore the efficacies of endoscopic third ventriculostomy (ETV) and tumor biopsy in surgical treatment of obstructive hydrocephalus induced by pineal region tumors. METHODS: For 65 patients of obstructive hydrocephalus induced by pineal region tumors, endoscopic third ventriculostomy (ETV) and tumor biopsy were performed from 2012 to 2014. Tumor specimens were harvested and analyzed histopathologically. According to pathological diagnosis, appropriate treatment strategies were followed. RESULTS: Sixty-five patients underwent successfully ETV and pineal region tumor biopsy. And the histological diagnosis yield rate was 98.5% (64/65). PEB chemotherapy (cisplatin+etoposide+bleomycin) and radiotherapy were offered for germ cell tumors (n = 37) and pineal blastomas (n = 6). The therapy effective rate of germ cell tumors was 91.7% (33/36). A total of 56.9% (37/65) patients benefited without traditional craniotomy or shunt. CONCLUSION: For pineal region tumor induced non-communicating hydrocephalus, ETV and endoscopic tumor biopsy can relieve hydrocephalus while obtaining pathological diagnosis at a relative low risk. Most patients of germ cell tumors and pineoblastomas may avoid craniotomy and shunt. It will become a routine treatment for pineal region tumors with non-communicating hydrocephalus.

[Magnetic resonance three dimensional sampling perfection with application optimized contrasts using different flip angle evolution sequence for obstructive hydrocephalus: impact on diagnosis and surgical strategy modification].

OBJECTIVE: To investigate the value of magnetic resonance three dimensional sampling perfection with application optimized contrasts using different flip angle evolution (3D-SPACE) sequences in diagnosis and surgical strategy modification for obstructive hydrocephalus. METHODS: From March 2013 to July 2014, there were 152 cases admitted in People's Liberation Army General Hospital suffered from hydrocephalus, including 88 male patients and 64 female patients aging from 8 months to 79 years. All patients were performed magnetic resonance T2WI and 3D-SPACE sequence scanning before operation. Surgical strategy was made after evaluation of 3D-SPACE sequence. Non-communicating hydrocephalus was treated with endoscopic third ventriculostomy (ETV) and communicating hydrocephalus was treated with ventriculo-peritoneal shunt. According to MR images of direct observation to site of obstruction to determine the detection rate. MRI 3D-SPACE and cranial CT examination were performed in regular follow-up studies. RESULTS: The relevance ratio of 3D-SPACE for the diagnosis of non-communicating hydrocephalus was 98.3% (114/116), while the relevance ratio of conventional T2-weighted MRI was 72.4% (84/116). Among the 152 patients, there were 36 cases with cerebral aqueduct film obstruction, 22 cases with space-occupying lesions in pineal region, posterior part of the third ventricle, or space-occupying lesions in quadrigeminal bodies area, 10 cases with Dandy-Walker symptom, 18 cases with cyst of the anterior pool of the bridge, 16 cases with cysticercosis, 4 cases with cyst of lateral ventricle, 2 cases with cyst of fourth ventricle, 2 cases with space-occupying lesion in foramen ofmonro, 2 cases with foramen ofmonro atresia, 4 cases with craniopharyngioma, 36 cases with communicating hydrocephalus. There were 112 hydrocephalus cases (73.7%) were treated with ETV, without shunt catheter insertion in follow-up study from 1 to 18 months (average (14±9) months). CONCLUSIONS: For obstructive hydrocephalus, MRI 3D-SPACE sequence image has high diagnostic yield rate for providing more detailed anatomical information than conventional MRI. Hence, the advanced imaging methods are helpful for surgical treatment strategy decision making.

[Diagnostic value of magnetic resonance spectroscopy and intraoperative magnetic resonance imaging in the treatment of brain abscesses].

OBJECTIVE: To evaluate the diagnostic value of magnetic resonance spectroscopy (MRS) and the role of intraoperative magnetic resonance imaging (iMRI) in the treatment of brain abscesses by aspiration and drainage. METHODS: From November 2009 to June 2013, Forty-one brain abscess patients were evaluated. MRS was employed to acquire the metabolic information and assist in the differential diagnosis of brain abscesses showing lactate cytosolic amino acids (AAs) with/without succinate, acetate, alanine and glycine on MRS. Diffusion-weighted imaging (DWI) was also helpful. Eleven single deep-seated abscesses underwent aspiration and drainage with 1.5 T iMRI and neuronavigation system. RESULTS: Forty-one brain abscesses were all diagnosed correctly. Ten single deep-seated abscesses underwent aspiration and drainage with 1.5 T iMRI and neuronavigation system successfully with just one puncture. One deep-seated abscess with thick-walled was punctured thrice before achieving success because of aspirated needle sidesliping. All 11 deep-seated abscess cases were cured and were also confirmed by follow-up. None of them suffered from significant complications, such as intracranial bleeding or new neurological deficit. CONCLUSION: MRS may acquire the metabolic information, confirm the presence of AAs with/without succinate, acetate, alanine and glycine and assist in the differential diagnosis of brain abscesses. iMRI system can help detect aspirated needle sidesliping and correct it in time to improve the cure rate, especially for single deep-seated brain abscess with a thick wall.

[The quantitative analysis of Ub and UbE1 in focus of traumatic epilepsy].

OBJECTIVE: To study the expression of ubiquitin proteasome system (UPS) in the traumatic epilepsy pathogenesis and its value in traumatic epilepsy by quantitative analysis. METHODS: Fifteen specimens from human epileptic temporal cortex from PTE were collected as the PTE group. Fifteen specimens from non-PTE were collected as the non-PTE group. Fifteen normal cerebral cortex specimens died from acute traffic accident were collected as the control group. The expression of mRNA and protein of Ub and UbE1 were detected by RT-PCR and Western blot. Statistical analysis was used to compare the data between three groups. RESULTS: The expression of mRNA and protein of Ub and UbE1 were the following order: PTE group(high), non-PTE group(middle) and control group(low). CONCLUSION: The study confirms that UPS is up-regulated in the epilepsy's focus, especially in traumatic epilepsy. The activation of UPS may be an important pathological change in neurons in pathogenesis of traumatic epilepsy.

[Morphology changes of ubiquitin-proteasome system in traumatic epilepsy].

OBJECTIVE: To investigate the value of ubiquitin(Ub) and ubiquitin-activating enzymel(UbE1) for the appraisement of post traumatic epilepsy (PTE). METHODS: Fifteen specimens from human epileptic temporal cortex originating from PTE were collected as the PTE group. Fifteen specimens from non-PTE were collected as the non-PTE group. Meanwhile, 15 normal cerebral cortex specimens from people dead from acute traffic accident were collected as the control groups. Observe morphology changes of each group with HE, then with immunohistochemistry of Ub and UbE1. RESULTS: Compared to the control group, morphology changes of neuron quantity reduction, neuron denaturation and so on were observed both in the PTE group and the non-PTE group under HE, especially in the PTE group. Ub and UbE1 mainly expressed in the nucleus and cytoplasm of the neurons in epilepsy spot without extracellular expression. The expression of Ub and UbE1 is PTE group > non-PTE group > control group (P < 0.05). CONCLUSIONS: The neuron denaturation are one of the main pathology changes of epilepsy, and it is more obvious in the PTE group. Immunohistochemistry of Ub and UbE1 may be more helpful to distinguish PTE and non-PTE than HE staining.