MicroRNA-3061 downregulates the expression of PAX7/Wnt/Ca2+ signalling axis genes to induce premature ovarian failure in mice.

The in-depth mechanisms of microRNA regulation of premature ovarian failure (POF) remain unclear. Crispr-cas9 technology was used to construct transgenic mice. The qPCR and Western blot was used to detect the expression level of genes. H&E staining were used to detect ovarian pathological phenotypes. We found that the expression levels of microRNA-3061 were significantly higher in ovarian granulosa cells (OGCs) of POF mouse models than in controls. The miR-3061+/-/AMH-Cre+/- transgenic mice manifested symptoms of POF. RNA-Seq and luciferase reporter assay confirmed that the PAX7 was one of the target genes negatively regulated by microRNA-3061 (miR-3061-5p). Moreover, PAX7 mediated the expression of non-canonical Wnt/Ca2+ signalling pathway by binding to the motifs of promoters to stimulate the transcriptional activation of Wnt5a and CamK2a. In contrast, specific knock-in of microRNA-3061 in OGCs significantly downregulated the expression levels of PAX7 and inhibited the expression of downstream Wnt/Ca2+ signalling pathway. We also discerned a correlation between the expression levels of mRNAs of the Wnt/Ca2+ signalling pathway and the levels of E2 and FSH in POF patients by examining gene expression in the follicular fluid-derived exosomes of women. We confirmed that overexpression of microRNA-3061 induced proliferative inhibition of OGCs and ultimately induced POF in mice by suppressing the transcription factor PAX7 and downregulating expression levels of its downstream Wnt/Ca2+ signalling pathway genes.

Detection of SARS-CoV-2 virus in middle ear effusions and its association with otitis media with effusion.

A large-scale outbreak of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) occurred in Shanghai, China, in early December 2022. To study the incidence and characteristics of otitis media with effusion (OME) complicating SARS-CoV-2, we collected 267 middle ear effusion (MEE) samples and 172 nasopharyngeal (NP) swabs from patients. The SARS-CoV-2 virus was detected by RT-PCR targeting. The SARS-CoV-2 virus, angiotensin-converting enzyme 2 (ACE2), and transmembrane serine protease 2 (TMPRSS2) expression in human samples was examined via immunofluorescence. During the COVID-19 epidemic in 2022, the incidence of OME (3%) significantly increased compared to the same period from 2020 to 2022. Ear symptoms in patients with SARS-CoV-2 complicated by OME generally appeared late, even after a negative NP swab, an average of 9.33 ± 6.272 days after COVID-19 infection. The SARS-CoV-2 virus was detected in MEE, which had a higher viral load than NP swabs. The insertion rate of tympanostomy tubes was not significantly higher than in OME patients in 2019-2022. Virus migration led to high viral loads in MEE despite negative NP swabs, indicating that OME lagged behind respiratory infections but had a favorable prognosis. Furthermore, middle ear tissue from adult humans coexpressed the ACE2 receptor for the SARS-CoV-2 virus and the TMPRSS2 cofactors required for virus entry.

Comparison of Endoscopic Cartilage Myringoplasty in Dry and Wet Ears With Chronic Suppurative Otitis Media.

OBJECTIVES: This study compared the rate of graft success, as well as hearing improvement and dry ear time between dry ears and wet ears with otomycosis or without otomycosis in patients with chronic suppurative otitis media (CSOM) after endoscopic cartilage myringoplasty. METHODS: This retrospective study was conducted in a tertiary hospital in Shanghai. In total, 83 patients with CSOM (43 with dry ears and 40 with wet ears) were included. Among the 40 patients with CSOM and wet ears, 25 exhibited otomycosis. All patients underwent endoscopic myringoplasty, and perforations were repaired using tragal cartilage with a single-sided perichondrium. Patients were followed up for at least 6 months. Pure-tone hearing was examined preoperatively and at 3 months postoperatively. The graft uptake rate, hearing improvement, and dry ear time were compared between the groups. RESULTS: The graft success rate did not differ significantly between the dry-ear and wet-ear groups (95.35% and 90.00%, respectively). Furthermore, the graft success rate also did not differ significantly between patients with wet ears and otomycosis and those with wet ears without otomycosis (92.00% and 86.67%, respectively). Hearing gain did not differ significantly between the dry-ear and wet-ear groups. No significant difference in hearing gain was also found in patients with wet ears with or without otomycosis. However, the time to dry ear was significantly longer in the wet-ear group than in the dry-ear group. CONCLUSION: Patients with CSOM and wet ears required more time to achieve a completely healthy status. However, the graft success rate and hearing improvement were not affected by a wet middle ear and otomycosis. Thus, endoscopic myringoplasty using tragus cartilage is an effective treatment for refractory CSOM in patients with wet ears and otomycosis.

Glycated Hemoglobin Electrochemical Immunosensor Based on Screen-Printed Electrode.

An electrochemical HbA1c sensor with high sensitivity and good specificity is proposed based on the electrochemical immune principle. The reproducibility and conductivity of the electrode are improved by depositing gold nanoparticles (AuNPs) on the surface of the screen-printed electrode (SPE). The HbA1c antibodies are immobilized on the surface of the modified electrode by adsorption to capture the HbA1c in the sample. The hindering effect of HbA1c on the electrode transfer reaction was exploited as the HbA1c detection mechanism. The electrode's properties were characterized by electrochemical impedance spectroscopy (EIS), and the measurement properties of the electrode were analyzed using differential pulse voltammetry (DPV) and cyclic voltammetry (CV). The experimental results show that the peak current signal of the electrochemical immunosensor produced a linear response to HbA1c in the concentration range of 20-200 µg/mL, a linear relationship coefficient of 0.9812, a detection limit of 15.5 µg/mL, and a sensitivity of 0.0938 µA/µg·mL-1. The sensor delivered satisfactory repeatability, stability, and anti-interference performance. Due to its small size, high sensitivity, and wide linear detection range, it is expected to play a significant role in managing diabetes at home.

Thymopentin alleviates premature ovarian failure in mice by activating YY2/Lin28A and inhibiting the expression of let-7 family microRNAs.

OBJECTIVE: Thymopentin (5TP) significantly improved typical murine premature ovarian failure (POF) symptoms induced by a high-fat and high-sugar (HFHS) diet. However, its effect and mechanism remain unclear. MATERIALS AND METHODS: RNA-Seq was used to detect the differentially expressed genes among each group. HFHS-induced POF mouse model was generated and injected with siRNA using Poly (lactic-co-glycolic acid) (PLGA) as a carrier. RESULTS: RNA-Seq suggested that 5TP promoted the expression of Yin Yang 2 (YY2) in mouse ovarian granulosa cell (mOGC) of HFHS-POF mice. Luciferase reporter assay indicated that 5TP promoted the binding of YY2 to the specific sequence C(C/T)AT(G/C)(G/T) on the Lin28A promoter and promoted Lin28A transcription and expression. We continuously injected PLGA-cross-linked siRNA nanoparticles targeting YY2 into HFHS-POF mice (siYY2@PLGA), which significantly reduced the therapeutic effect of 5TP. siYY2@PLGA injection also significantly attenuated the upregulation of Lin28a expression in mOGCs induced by 5TP and enhanced the expression of let-7 family microRNAs, thereby inhibiting the proliferation and division of mOGCs. qPCR results showed that there was a significant difference in the expression levels of exosome-derived Yy2 mRNAs between POF patients and normal women, and that there was a specific correlation between the expression level of exosome-derived Yy2 and the peripheral concentrations of the blood hormones pregnenolone, progesterone and oestradiol. CONCLUSIONS: Thymopentin promotes the transcriptional activation of Lin28A via stimulating transcription factor YY2 expression, inhibits the activity of let-7 family microRNAs and alleviates the ageing of ovarian granulosa cells, ultimately achieving a therapeutic effect on POF in mice.

AI powered electrochemical multi-component detection of insulin and glucose in serum.

Multi-component detection of insulin and glucose in serum is of great importance and urgently needed in clinical diagnosis and treatment due to its economy and practicability. However, insulin and glucose can hardly be determined by traditional electrochemical detection methods. Their mixed oxidation currents and rare involvement in the reaction process make it difficult to decouple them. In this study, AI algorithms are introduced to power the electrochemical method to conquer this problem. First, the current curves of insulin, glucose, and their mixed solution are obtained using cyclic voltammetry. Then, seven features of the cyclic voltammetry curve are extracted as characteristic values for detecting the concentrations of insulin and glucose. Finally, after training using machine learning algorithms, insulin and glucose concentrations are decoupled and regressed accurately. The entire detection process only takes three minutes. It can detect insulin at the pmol level and glucose at the mmol level, which meets the basic clinical requirements. The average relative error in predicting insulin concentrations is around 6.515%, and that in predicting glucose concentrations is around 4.36%. To verify the performance and effectiveness of the proposed method, it is used to determine the concentrations of insulin and glucose in fetal bovine serum and real clinical serum samples. The results are satisfactory, demonstrating that the method can meet basic clinical needs. This multi-component testing system delivers acceptable detect limit and accuracy and has the merits of low cost and high efficiency, holding great potential for use in clinical diagnosis.

Clinical Characteristics of Patients With Papilloma in the External Auditory Canal.

OBJECTIVES: To determine the clinical characteristics of papilloma involving the external auditory canal (PEAC) in a region of China. STUDY DESIGN: A retrospective study. METHODS: Demographics, manifestations, imaging results, histopathology, and treatment of 67 patients diagnosed with PEAC in a period of 6 years were analyzed at Shanghai Eye, Ear, Nose and Throat Hospital in China. RESULTS: PEAC were encountered in patients between the ages of 12 and 82 years (mean 53.8 years). It was more prevalent in men (82%) than in women (18%) (P < .05). The clinical presentation was usually a mass in EAC, aural fullness, and hearing loss. Otoscopic and radiological examination were used together for initial diagnosis and pretreatment planning. Unilateral involvement was more common than bilateral involvement (P < .05). The average time between onset of first symptom and surgical resection and/or biopsy was 6.5 months (range, 0.25-60 months). All patients underwent gross total resection. In 5 patients, (7.5%) carcinoma was detected in the specimen. Fifteen patients (22%) had recurrence; recurrent tumors were detected after an average period of 10 months after surgery (range, 4-24 months). CONCLUSION: PEAC is largely a benign lesion with a low risk of malignancy. Optimal management is via gross total resection. However, the risk of recurrence is high, which motivates a need for long-term monitoring. LEVEL OF EVIDENCE: 4 Laryngoscope, 131:1132-1137, 2021.

Bacteriological Profile of Chronic Suppurative Otitis Media and Antibiotic Susceptibility in a Tertiary Care Hospital in Shanghai, China.

OBJECTIVE: Chronic suppurative otitis media (CSOM) is mostly caused by bacterial infection of the middle ear and antibiotics are generally used empirically, which may lead to the emergence of resistant bacterial strains. The objective of the study is to assess the bacteriological profile of and evaluate the antibiotic susceptibility of strains isolated in a tertiary care hospital in Shanghai, China. METHODS: This study included 289 individuals with clinical diagnosis of CSOM. Middle ear purulent discharge was obtained with sterile swabs and cultured for bacterial pathogens. The susceptibility of the isolated microorganisms to antibiotics was assessed by a microdilution method. RESULTS: Bacterial pathogens were found in 223 (77.2%) of the 289 cases. A total of 236 strains were isolated. Staphylococcus aureus was the commonest bacteria (44.9%) followed by Pseudomonas aeruginosa (16.9%) and coagulase-negative Staphylococcus (8.5%). There were 18.9% methicillin-resistant S aureus (MRSA) among the obtained S aureus organisms. Multidrug-resistant P aeruginosa was found in 4 patients, making up 10% of all detected P aeruginosa. Staphylococcus aureus showed highest susceptibility to vancomycin (100%), then gentamicin (98.1%) and rifampicin (97.2%) and was most resistant to penicillin (61.3%) and erythromycin (50%). All isolated P aeruginosa showed susceptibility to piperacillin, piperacillin/tazobactam, and meropenem. High degree of resistance in P aeruginosa was observed toward levofloxacin (42.5%), ciprofloxacin (40%), and ceftriaxone (30%). CONCLUSION: The high prevalence of MRSA and fluoroquinolone-resistant P aeruginosa indicated cephalosporins and fluoroquinolone as primary empirical antibiotic drugs in CSOM to be cautiously used. In order to reduce the incidence of resistant strains and promote effective usage of antibiotics, all aural discharges should be cultured to determine antibacterial susceptibility patterns before treatment.

The microbiomes of adenoid and middle ear in children with otitis media with effusion and hypertrophy from a tertiary hospital in China.

INTRODUCTION: Otitis media with effusion (OME) is one of the most common pediatric diseases worldwide. Several studies have analyzed the diversity of the microbiomes found in the middle ear effusions (MEEs) of populations from developed countries. However, no microbiological studies of MEEs from Chinese children with OME have been reported. This study investigated the middle ear and adenoid microbiological profiles of children with OME, and compared the microbial flora of the adenoid between children with and without otitis media. METHODS: MEEs and adenoid swabs were acquired from 15 children undergoing ventilation tube insertion and adenoidectomy. Adenoid swabs from 15 patients with no ear disease were used as controls. Samples were analyzed by 16S rRNA sequencing. Operational taxonomic units (OTUs) abundance information were normalized. Alpha diversity analyses were used to assess the richness and diversity of the microbial community for each sample. Beta diversity analyses were used to determine the inter-group variability between microbiome structure. RESULTS: Based on the mean relative abundance, the MEEs were dominated by Haemophilus (14.75%), Staphylococcus (9.37%) and Halomonas (7.85%), and the bacterial compositions of the adenoids in the OME groups were dominated by Haemophilus (21.87%), Streptococcus (19.65%), and Neisseria (5.8%). The bacterial compositions in the adenoids of the controls were dominated by Haemophilus (15.96%), Streptococcus (13.33%), and Moraxella (12.28%). Alpha diversity analyses showed that there were no significant differences in microbiome richness or diversity between the middle ear effusions (TM) and adenoids (TA) of OME subjects. Adenoid samples from OME patients (TA) and control patients (CA) were also similar. Beta diversity analyses showed that the microbiomes of the adenoids in OME patients were also similar to that of controls. However, the microbiome structure of middle ear effusions was dissimilar to those of the adenoids in OME patients according to beta diversity analyses. CONCLUSIONS: Our results confirmed the microbial diversity of MEEs among Chinese children. However, the dissimilar microbiome composition between samples taken from the surface of the adenoids and from the middle ear effusions challenges the conventional theory that the adenoid serves as a microbial reservoir in children with otitis media with effusion.

Deep Learning in Automated Region Proposal and Diagnosis of Chronic Otitis Media Based on Computed Tomography.

OBJECTIVES: The purpose of this study was to develop a deep-learning framework for the diagnosis of chronic otitis media (COM) based on temporal bone computed tomography (CT) scans. DESIGN: A total of 562 COM patients with 672 temporal bone CT scans of both ears were included. The final dataset consisted of 1147 ears, and each of them was assigned with a ground truth label from one of the 3 conditions: normal, chronic suppurative otitis media, and cholesteatoma. A random selection of 85% dataset (n = 975) was used for training and validation. The framework contained two deep-learning networks with distinct functions: a region proposal network for extracting regions of interest from 2-dimensional CT slices; and a classification network for diagnosis of COM based on the extracted regions. The performance of this framework was evaluated on the remaining 15% dataset (n = 172) and compared with that of 6 clinical experts who read the same CT images only. The panel included 2 otologists, 3 otolaryngologists, and 1 radiologist. RESULTS: The area under the receiver operating characteristic curve of the artificial intelligence model in classifying COM versus normal was 0.92, with sensitivity (83.3%) and specificity (91.4%) exceeding the averages of clinical experts (81.1% and 88.8%, respectively). In a 3-class classification task, this network had higher overall accuracy (76.7% versus 73.8%), higher recall rates in identifying chronic suppurative otitis media (75% versus 70%) and cholesteatoma (76% versus 53%) cases, and superior consistency in duplicated cases (100% versus 81%) compared with clinical experts. CONCLUSIONS: This article presented a deep-learning framework that automatically extracted the region of interest from two-dimensional temporal bone CT slices and made diagnosis of COM. The performance of this model was comparable and, in some cases, superior to that of clinical experts. These results implied a promising prospect for clinical application of artificial intelligence in the diagnosis of COM based on CT images.

Spider Toxin Peptide Lycosin-I Functionalized Gold Nanoparticles for in vivo Tumor Targeting and Therapy.

Cell penetrating peptides (CPPs) are commonly utilized for intracellular delivery of functional materials to circumvent biomembrane barrier. However, further application of CPPs is hindered by lacking selectivity toward targeted cells. The spider venom peptide, lycosin-I, is a CPP with potent cytotoxicity to cancer cells, which might enable lycosin-I to deliver functional materials into cancer cells selectively. In this study, we demonstrated that the lycosin-I-conjugated spherical gold nanoparticles (LGNPs) not only exhibited efficient cellular internalization efficiency toward cancer cells but also displayed unprecedented selectivity over noncancerous cells. Although LGNPs were removed from the living circulatory system via reticuloendothelial system-dominant clearance modes without noticeable adverse effects to animals, they actually displayed active tumor-targeting effects and efficient accumulation in tumors in vivo. Furthermore, the potential application of this platform for cancer therapy was explored by lycosin-I-conjugated gold nanorods (LGNRs). LGNRs exhibited selective intracellular translocation towards cancer cells and efficient photothermal effect under near infrared (NIR, 808 nm) irradiation, which consequently killed cancer cells in vitro and in vivo effectively. Therefore, the established LGNPs and LGNRs possessed great potential in cancer-targeting delivery and photothermal therapy.

Sub-diffraction-limit localization imaging of a plasmonic nanoparticle pair with wavelength-resolved dark-field microscopy.

In this work, with wavelength-resolved dark-field microscopy, the center-of-mass localization information from nanoparticle pairs (i.e., spherical (45 nm in diameter) and rod (45 × 70 nm) shaped gold nanoparticle pairs with different gap distances and orientations) was explored and compared with the results determined by scanning electron microscopy (SEM) measurements. When the gap distance was less than 20 nm, the scattering spectrum of the nanoparticle pair was seriously modulated by the plasmonic coupling effect. The measured coordinate information determined by the optical method (Gaussian fitting) was not consistent with the true results determined by SEM measurement. A good correlation between the optical and SEM measurements was achieved when the gap distance was further increased (e.g., 20, 40 and 60 nm). Under these conditions, well-defined scattering peaks assigned to the corresponding individual nanoparticles could be distinguished from the obtained scattering spectrum. These results would afford valuable information for the studies on single plasmonic nanoparticle imaging applications with the optical microscopy method such as super-localization imaging, high precision single particle tracking in a crowding environment and so on.

Immunogenicity and protective immunity against otitis media caused by pneumococcus in mice of Hib conjugate vaccine with PsaA protein carrier.

This study evaluated the immunogenicity and protective immunity of a Hemophilus influenzae b (Hib) polysaccharide conjugate vaccine with the pneumococcal surface adhesin A (PsaA) protein carrier in young mice. The Hib polysaccharide was conjugated with the rPsaA protein carrier, which was produced using recombinant DNA technology. A total of 15 young mice aged 3 weeks to 5 weeks were immunized with the conjugate vaccine, and another 15 young mice of the same age were immunized with the licensed Hib-tetanus toxoid (TT) vaccine. Furthermore, the third group of 15 young mice was inoculated with phosphate buffer saline as control. The immunized mice were inoculated with pneumococcus in the middle ear. Results showed that IgG antibody responses against both the PsaA protein and Hib polysaccharide were observed in the Hib-PsaA group. However, no statistical difference was observed in the titer of IgG against the Hib polysaccharide between Hib-PsaA and Hib-TT groups. The elimination rate of pneumococcus and the inflammation of the middle ear showed the effectiveness of protective immunity against otitis media caused by pneumococcus. Our results suggest that the Hib polysaccharide can be successfully conjugated with rPsaA via amide condensation. This new Hib-PsaA conjugate vaccine can induce both anti-PsaA and anti-Hib immune responses in young mice and elicit effective protection against acute otitis media caused by pneumococcus.

Orientational Imaging of a Single Gold Nanorod at the Liquid/Solid Interface with Polarized Evanescent Field Illumination.

Understanding the mechanistic information on many kinetic processes requires the exploration of dynamic rotational information on the target object at the single particle (or molecule) level. In this work, we developed a new strategy, total internal reflection scattering (TIRS) microscopy, to determine the full three-dimensional (3D) angular information on a single gold nanorod (GNR) close to the liquid/solid interface. It was found that the 3D orientational information on individual GNR could be readily elucidated by using p-polarized TIRS illumination through deciphering the orientation-coded intensity distribution pattern in a single TIRS image. In comparison with the previously reported strategies, this method does not require complicated focal plane correction, affording a versatile pathway to track the rotational dynamics close to the interface in a high throughput manner. The methodology presented here, therefore, demonstrates a promising approach that can be applied to fluidic membranes, including membranes with polymers, bound proteins, and so on.

Combination of submicroemulsion and phospholipid complex for novel delivery of ursodeoxycholic acid.

The objective of this study was to prepare and characterize ursodeoxycholic acid submicron emulsion (UA-SME) loaded with ursodeoxycholic acid phytosomes (UA-PS) and optimize the process variables. A screening experiment with response surface methodology with Box-Behnken design (BBD) was used to optimize the process parameters of UA-SME. The blood concentrations of UA after oral administration of UA-SME and UA coarse drug were assayed. The optimum process conditions were finally obtained by using a desirability function. It was found that stirring velocity, homogenization pressure and homogenization cycles were the most important variables that affected the particles size, polydispersity index and entrapment efficiency of UA-SME. Results showed that the optimum stirring velocity, homogenization pressure and cycles were 16 000 rpm, 60 MPa and 10 cycles, respectively. The mean diameter, polydispersity index and entrapment efficiency of UA-SME were 251.9 nm, 0.241 and 74.36%, respectively. Pharmacokinetic parameters of UA and UA-SME in rats were Tmax 2.215 and 1.489 h, Cmax 0.0364 and 0.1562 µg/mL, AUC0-∞ 3.682 and 13.756 µg h/mL, respectively. The bioavailability of UA in rats was significantly different (p < 0.05) after oral administration of UA-SME compared to those of UA coarse drug. This was due to improvement of the hydrophilicity and lipophilic property of UA-SME.

Mandarin Chinese speech recognition by pediatric cochlear implant users.

OBJECTIVES: Because of difficulties associated with pediatric speech testing, most pediatric cochlear implant (CI) speech studies necessarily involve basic and simple perceptual tasks. There are relatively few studies regarding Mandarin-speaking pediatric CI users' perception of more difficult speech materials (e.g., words and sentences produced by multiple talkers). Difficult speech materials and tests necessarily require older pediatric CI users, who may have different etiologies of hearing loss, duration of deafness, CI experience. The present study investigated how pediatric CI patient demographics influence speech recognition performance with relatively difficult test materials and methods. METHODS: In this study, open-set recognition of multi-talker (two males and two females) Mandarin Chinese disyllables and sentences were measured in 37 Mandarin-speaking pediatric CI users. Subjects were grouped according to etiology of deafness and previous acoustic hearing experience. Group 1 subjects were all congenitally deafened with little-to-no acoustic hearing experience. Group 2 subjects were not congenitally deafened and had substantial acoustic hearing experience prior to implantation. Multiple linear regression analyses were performed within each group using subject demographics such as age at implantation and age at testing. RESULTS: Pediatric CI performance was generally quite good. For Group 1, mean performance was 82.3% correct for disyllables and 82.8% correct for sentences. For Group 2, mean performance was 76.6% correct for disyllables and 84.4% correct for sentences. For Group 1, multiple linear regression analyses showed that age at implantation predicted disyllable recognition, and that age at implantation and age at testing predicted sentence recognition. For Group 2, neither age at implantation nor age at testing predicted disyllable or sentence recognition. Performance was significantly better with the female than with the male talkers. CONCLUSIONS: Consistent with previous studies' findings, early implantation provided a significant advantage for profoundly deaf children. Performance for both groups was generally quite good for the relatively difficult materials and tasks, suggesting that open-set word and sentence recognition may be useful in evaluating speech performance with older pediatric CI users. Differences in disyllable recognition between Groups 1 and 2 may reflect differences in adaptation to electric stimulation. The Group 1 subjects developed speech patterns exclusively via electric stimulation, while the Group 2 subjects adapted to electric stimulation relative to previous acoustic patterns.

Intranasal vaccination with chitosan-DNA nanoparticles expressing pneumococcal surface antigen a protects mice against nasopharyngeal colonization by Streptococcus pneumoniae.

Streptococcus pneumoniae is a respiratory pathogen, and mucosal immune response plays a significant role in the defense against pneumococcal infections. Thus, intranasal vaccination may be an alternative approach to current immunization strategies, and effective delivery systems to mucosal organism are necessary. In this study, BALB/c mice were immunized intranasally with chitosan-DNA nanoparticles expressing pneumococcal surface antigen A (PsaA). Compared to levels in mice immunized with naked DNA or chitosan-pVAX1, anti-PsaA IgG antibody in serum and anti-IgA antibody in mucosal lavages were elevated significantly in mice immunized with chitosan-psaA. The balanced IgG1/IgG2a antibody ratio in serum, enhanced gamma interferon (IFN-γ) and IL-17A levels in spleen lymphocytes, and mucosal washes of mice immunized with chitosan-psaA suggested that cellular immune responses were induced. Furthermore, significantly fewer pneumococci were recovered from the nasopharynx of mice immunized with chitosan-psaA than for the control group following intranasal challenge with ATCC 6303 (serotype 3). These results demonstrated that mucosal immunization with chitosan-psaA may successfully generate mucosal and systemic immune responses and prevent pneumococcal nasopharyngeal colonization. Hence, a chitosan-DNA nanoparticle vaccine expressing pneumococcal major immunodominant antigens after intranasal administration could be developed to prevent pneumococcal infections.

Relationship between high-resolution computed tomography densitometry and audiometry in otosclerosis.

OBJECTIVE: The aim of this study is to evaluate the usefulness of high-resolution computed tomography (HRCT) densitometry in the diagnosis of otosclerosis and to investigate the relationship between CT densitometry and audiometry. METHODS: HRCT findings and audiometry were compared among 34 patients (34 ears, the otosclerosis group) with surgically confirmed otosclerosis between January 2007 and December 2007 and 33 patients (33 opposite normal ears, the control group) with facial paralysis diagnosed at the same period of time. Seven regions of interest (ROI) were set manually around the otic capsule on the axial slice of 0.75-mm-thick CT image. The mean CT values of these seven regions were measured. In each ROI, the mean CT value of the otosclerosis group and that of the control group were compared. Based on the CT findings, the ears with otosclerosis were classified into two groups: Group A showed no pathological CT findings; Group B showed low density around the cochlea. In the otosclerosis group, the relationship between the findings of CT and the results of audiometry was analyzed. RESULTS: The mean CT values in the area posterior to the oval window and anterior to the oval window were significantly lower for the otosclerosis group compared with the control group (the former t=-2.030, p=0.046; the latter Z=-4.979, p<0.01). Group A consisted of 30 patients, 7 of which (23.33%) exhibited conductive hearing loss, and 23 of which (76.67%) exhibited mixed hearing loss; Group B had 4 patients, all with mixed hearing loss. For the otosclerosis group, the mean CT value in the area posterior to the oval window was positively correlated with the mean air conduction threshold (r=0.4273, p=0.0117) and with the mean air-bone gap (r=0.3995, p=0.0192). CONCLUSION: Quantitative evaluation of CT with slices less than 1mm in thickness may provide important information for the diagnosis and assessment of otosclerosis which are unattainable through other methods.

[Improvement of immunogenicity of Streptococcus pneumoniae psaA DNA vaccine by prime and boost strategy].

OBJECTIVE: To study on the preparation of Streptococcus pneumoniae psaA DNA vaccine and to analyse the immunogenicity by the prime-boost strategy. METHODS: The psaA gene was amplified from the genome of Streptococcus pneumoniae by PCR, and then was inserted into plasmid pVAX1 and pET28a to construct recombinant expression vectors respectively. 293T cells were transiently transfected with pVAX1-psaA, and RT-PCR analysis of total cell RNA extracts showed successful expression of psaA. BALB/c mices (n = 5) were intramuscularly injected with 100 microg psaA DNA vaccine for three times, and then boosted with 50 microg recombinant PsaA protein. The antibody response against PsaA was measured by ELISA. RESULTS: The psaA gene was amplified and subcloned successfully. The constructed psaA DNA vaccine was confirmed by DNA sequencing, and the recombinant PsaA protein was purified by the one-step Ni(2+) affinity chromatography. Expression of the PsaA was observed in cells transfected with pVAX1-psaA. The animal experiment results showed that the anti-PsaA level of the DNA prime-protein boosting mice was higher significantly than the other groups (t = 87.518, P < 0.05). CONCLUSION: The psaA DNA vaccine was prepared successfully, and the immunogenicity of Streptococcus pneumoniae psaA DNA vaccine could be improved significantly by the DNA prime and protein boost strategy.

Isolation of fenitrothion-degrading strain Burkholderia sp. FDS-1 and cloning of mpd gene.

A short rod shaped, gram-negative bacterium strain Burkholderia sp. FDS-1 was isolated from the sludge of the wastewater treating system of an organophosphorus pesticides manufacturer. The isolate was capable of using fenitrothion as the sole carbon source for its growth. FDS-1 first hydrolyzed fenitrothion to 3-methyl-4-nitrophenol, which was further metabolized to nitrite and methylhydroquinone. The addition of other carbon source and omitting phosphorus source had little effect on the hydrolysis of fenitrothion. The gene encoding the organophosphorus hydrolytic enzyme was cloned and sequenced. The sequence was similar to mpd, a gene previously shown to encode a parathion-methyl-hydrolyzing enzyme in Plesiomonas sp. M6. The inoculation of strain FDS-1 (10(6) cells g(-1)) to soil treated with 100 mg fenitrothion emulsion kg(-1) resulted in a higher degradation rate than in noninoculated soils regardless of the soil sterilized or nonsterilized. These results highlight the potential of this bacterium to be used in the cleanup of contaminated pesticide waste in the environment.