Bat coronavirus was detected positive from insectivorous bats in Krau Wildlife Reserve Forest.

Bats are flying mammals with unique immune systems that allow them to hold many pathogens. Hence, they are recognised as the reservoir of many zoonotic pathogens. In this study, we performed molecular detection to detect coronaviruses, paramyxoviruses, pteropine orthoreoviruses and dengue viruses from samples collected from insectivorous bats in Krau Reserve Forest. One faecal sample from Rhinolophus spp. was detected positive for coronavirus. Based on BLASTN, phylogenetic analysis and pairwise alignment-based sequence identity calculation, the detected bat coronavirus is most likely to be a bat betacoronavirus lineage slightly different from coronavirus from China, Philippines, Thailand and Luxembourg. In summary, continuous surveillance of bat virome should be encouraged, as Krau Reserve Forest reported a wide spectrum of biodiversity of insectivorous and fruit bats. Moreover, the usage of primers for the broad detection of viruses should be reconsidered because geographical variations might possibly affect the sensitivity of primers in a molecular approach.

Bat coronavirus was detected positive from insectivorous bats in Krau Wildlife Reserve Forest

@#Bats are flying mammals with unique immune systems that allow them to hold many pathogens. Hence, they are recognised as the reservoir of many zoonotic pathogens. In this study, we performed molecular detection to detect coronaviruses, paramyxoviruses, pteropine orthoreoviruses and dengue viruses from samples collected from insectivorous bats in Krau Reserve Forest. One faecal sample from Rhinolophus spp. was detected positive for coronavirus. Based on BLASTN, phylogenetic analysis and pairwise alignment-based sequence identity calculation, the detected bat coronavirus is most likely to be a bat betacoronavirus lineage slightly different from coronavirus from China, Philippines, Thailand and Luxembourg. In summary, continuous surveillance of bat virome should be encouraged, as Krau Reserve Forest reported a wide spectrum of biodiversity of insectivorous and fruit bats. Moreover, the usage of primers for the broad detection of viruses should be reconsidered because geographical variations might possibly affect the sensitivity of primers in a molecular approach.

CD40L-stimulated B cells for ex-vivo expansion of polyspecific non-human primate regulatory T cells for translational studies.

The therapeutic applications of regulatory T cells (Tregs ) include treating autoimmune diseases, graft-versus-host disease and induction of transplantation tolerance. For ex-vivo expanded Tregs to be used in deceased donor transplantation, they must be able to suppress T cell responses to a broad range of human leukocyte antigen (HLA). Here, we present a novel approach for the expansion of polyspecific Tregs in cynomolgus macaques that was adapted from a good manufacturing practice-compliant protocol. Tregs were isolated by fluorescence-activated cell sorting (FACS) and expanded in the presence of a panel of CD40L-stimulated B cells (CD40L-sBc). Prior to Treg culture, CD40L-sBc were expanded in vitro from multiple major histocompatibility complex (MHC)-disparate macaques. Expanded Tregs expressed high levels of forkhead box protein 3 (FoxP3) and Helios, a high percentage of Treg -specific demethylated region (TSDR) demethylation and strong suppression of naïve T cell responses in vitro. In addition, these Tregs produced low levels of inflammatory cytokines and were able to expand post-cryopreservation. Specificity assays confirmed that these Tregs were suppressive upon activation by any antigen-presenting cells (APCs) whose MHC was shared by CD40L-sBc used during expansion, proving that they are polyspecific. We developed an approach for the expansion of highly suppressive cynomolgus macaque polyspecific Tregs through the use of a combination of CD40L-engineered B cells with the potential to be translated to clinical studies. To our knowledge, this is the first report that uses a pool of MHC-mismatched CD40L-sBc to create polyspecific Tregs suitable for use in deceased-donor transplants.

Ambulance use affects timely emergency treatment of acute ischaemic stroke.

INTRODUCTION: For acute ischaemic stroke patients, treatment with intravenous tissue plasminogen activator within a 4.5-hour therapeutic window is essential. We aimed to assess the time delays experienced by stroke patients arriving at the emergency department and to compare ambulance users and non-ambulance users. METHODS: We performed a prospective cohort study in a tertiary hospital in Hong Kong. All acute stroke patients attending the emergency department from January to June 2017 were recruited. Patients who were in hospital at the time of stroke onset and those who transferred from other hospitals were excluded. Three phases were compared between ambulance users and non-ambulance users: phase I, between stroke onset and calling for help; phase II, between calling for help and arriving at the emergency department; and phase III, between arriving and receiving medical assessment. RESULTS: Of 102 consecutive patients recruited, 48 (47%) patients arrived at the emergency department by ambulance. The percentage of stroke patients attending emergency department within the therapeutic window was significantly higher for ambulance users than for non-ambulance users (64.6% vs 29.6%; P<0.001). For phases I, II and III, the median times were significantly shorter for ambulance users (77.5, 32 and 8 min, respectively) than for non-ambulance users (720, 44.5 and 15 min, respectively; all P<0.001). CONCLUSION: Transport of patients to the emergency department by ambulance is important for timely and effective stroke treatment.

Predicting arboviral disease emergence using Bayesian networks: a case study of dengue virus in Western Australia.

A Bayesian Belief Network (BBN) for assessing the potential risk of dengue virus emergence and distribution in Western Australia (WA) is presented and used to identify possible hotspots of dengue outbreaks in summer and winter. The model assesses the probabilities of two kinds of events which must take place before an outbreak can occur: (1) introduction of the virus and mosquito vectors to places where human population densities are high; and (2) vector population growth rates as influenced by climatic factors. The results showed that if either Aedes aegypti or Ae. albopictus were to become established in WA, three centres in the northern part of the State (Kununurra, Fitzroy Crossing, Broome) would be at particular risk of experiencing an outbreak. The model can also be readily extended to predict the risk of introduction of other viruses carried by Aedes mosquitoes, such as yellow fever, chikungunya and Zika viruses.

Initial results of selective renal parenchymal clamping with an adjustable kidney clamp in nephron-sparing surgery: an easy way to minimise renal ischaemia.

INTRODUCTION: A renal parenchymal clamp has been used at our centre since March 2012. It is used in position over the kidney to achieve optimal vascular control of a tumour while minimising parenchymal ischaemia. This study aimed to report the feasibility, surgical outcome, and oncological control of a kidney clamp in partial nephrectomy. METHODS: This study was conducted at a teaching hospital in Hong Kong. Partial nephrectomies performed from January 2009 to March 2015 were reviewed. The tumour characteristics and surgical outcomes of kidney clamp were studied and compared with traditional hilar clamping. RESULTS: A total of 92 patients were identified during the study period. Kidney clamps were used in 20 patients and hilar clamping in 72, with a mean follow-up of 27 and 37 months, respectively. For patients in whom a kidney clamp was applied, all tumours were exophytic to a different extent and the majority (90%) were located at the polar region. The PADUA (preoperative aspects and dimensions used for an anatomical) classification nephrometry score was also lower than those in whom hilar clamping was used (7.07 vs 8.34; P=0.002). The clamp was used in open, laparoscopic, and robot-assisted surgery. Operating time was shorter (207 ± 72 mins vs 306 ± 80 mins; P<0.001) and estimated blood loss was lower (205 ± 191 mL vs 331 ± 275 mL; P=0.045) with kidney clamp. No acute kidney injury occurred. Postoperative renal function was comparable between the two groups. CONCLUSIONS: Partial nephrectomy using parenchymal clamping is safe and feasible in selected cases. The postoperative renal function and oncological control were satisfactory.

Development of an E-learning System for the Endoscopic Diagnosis of Early Gastric Cancer: An International Multicenter Randomized Controlled Trial.

BACKGROUND: In many countries, gastric cancer is not diagnosed until an advanced stage. An Internet-based e-learning system to improve the ability of endoscopists to diagnose gastric cancer at an early stage was developed and was evaluated for its effectiveness. METHODS: The study was designed as a randomized controlled trial. After receiving a pre-test, participants were randomly allocated to either an e-learning or non-e-learning group. Only those in the e-learning group gained access to the e-learning system. Two months after the pre-test, both groups received a post-test. The primary endpoint was the difference between the two groups regarding the rate of improvement of their test results. FINDINGS: 515 endoscopists from 35 countries were assessed for eligibility, and 332 were enrolled in the study, with 166 allocated to each group. Of these, 151 participants in the e-learning group and 144 in the non-e-learning group were included in the analysis. The mean improvement rate (standard deviation) in the e-learning and non-e-learning groups was 1·24 (0·26) and 1·00 (0·16), respectively (P<0·001). INTERPRETATION: This global study clearly demonstrated the efficacy of an e-learning system to expand knowledge and provide invaluable experience regarding the endoscopic detection of early gastric cancer (R000012039).

Toxicity of repeated intravenous injection of gene therapeutics for X-CGD in mice.

We made gene therapeutics for X-chronic granulomatous disease (CGD) by transducing murine bone marrow-derived stem cells with MT-gp91 retrovirus and evaluated possible toxicity in mice as a prerequisite for human clinical trials. Male C57BL/6 mice were injected intravenously with gene therapeutics for X-CGD twice at an interval of two weeks at 5 x 10(7) cells/kg and sacrificed 2 weeks after the last administration. Significant changes noted in gene therapeutics for X-CGD-treated animals were an increase in white blood cell counts and a slight decrease in albumin/globulin ratio. The red pulp hyperplasia in the spleen accompanied with an increase in organ weight was considered to result from the accumulation of gene therapeutics for X-CGD, bone marrow-derived stem cells, in the spleen. No anti-gp91 antibody was detected in the sera collected from the animals treated with gene therapeutics for X-CGD. No integration of gp91 DNA from retroviral vector was detected in chromosomal DNA of gonads in animals dosed with the test substance, indicating no potential of genomic integration. In conclusion, the repeated dose of gene therapeutics for X-CGD exerted no toxicity. The splenic red pulp hyperplasia and the increase observed in white blood cell counts and in spleen weights were considered as pharmacological changes induced by the treatment.

Immunogenicity and safety profiles of genetic vaccines against human Her-2/neu in cynomolgus monkeys.

Her-2/neu is a well-characterized tumor-associated antigen, the overexpression of which in human carcinomas correlates with a poor prognosis. Here, we evaluated Her-2/neu-specific humoral and cellular immune responses in immunized monkeys after immunization with nonreplicating adenovirus (AdHM) expressing the extracellular and transmembrane domain of human Her-2/neu (HM) and/or naked DNA vaccine (pHM-hGM-CSF) expressing human granulocyte-macrophage colony-stimulating factor together with HM. Priming of monkeys with AdHM generated Her-2/neu-specific long-lasting antibody production. Furthermore, these Her-2/neu-specific antibodies produced by AdHM immunization, some of which shared epitope specificity with Herceptin, were able to induce antibody-dependent cellular cytotoxicity against Her-2-expressing target cells. Cellular immune responses were elicited in all monkeys immunized with Her-2/neu-expressing vaccine; interferon-gamma was secreted when these splenocytes were restimulated with Her-2/neu-expressing autologous cells, and immunization with AdHM induced Her-2/neu-specific lymphoproliferative responses. Further, immunization with pHM-hGM-CSF before AdHM immunization noticeably enhanced cytotoxic T-lymphocyte activity. In addition, we observed no abnormalities that would indicate that the genetic vaccines had toxic effects in the immunized monkeys. Thus, we can conclude that our genetic vaccines efficiently elicited Her-2/neu-specific humoral and cellular immune responses without causing severe adverse effects in nonhuman primates and that as such they warrant further clinical investigation.

A combination of rough-based feature selection and RBF neural network for classification using gene expression data.

This paper presents a novel rough-based feature selection method for gene expression data analysis. It can find the relevant features without requiring the number of clusters to be known a priori and identify the centers that approximate to the correct ones. In this paper, we attempt to introduce a prediction scheme that combines the rough-based feature selection method with radial basis function neural network. For further consider the effect of different feature selection methods and classifiers on this prediction process, we use the NaIve Bayes and linear support vector machine as classifiers, and compare the performance with other feature selection methods, including information gain and principle component analysis. We demonstrate the performance by several published datasets and the results show that our proposed method can achieve high classification accuracy rate.

C. elegans SIN-3 and its associated HDAC corepressor complex act as mediators of male sensory ray development.

Mab21 gene family members are required for embryonic development and sensory organ formation in both invertebrates and vertebrates. However, their mechanistic role on differentiation is largely unexplored. We report here the isolation of SIN-3 as a MAB-21 interacting molecule. sin-3 is co-expressed with mab-21 in the ray structural cells and genetically interacts with mab-21 to control sensory organ development. Using pharmacological and RNAi approaches, we demonstrated that histone deacetylase and conserved SIN-3-associated components are required for ray patterning. Conserved physical interactions between these components were also observed, implicating the recruitment of HDAC complex by MAB-21/SIN-3 may occur to determine ray identity in males.

Recurrent prostatic stromal tumour of uncertain malignant potential (STUMP) presenting with urinary retention 6 Years after transurethral resection of prostate (TURP).

CLINICAL PRESENTATION: A 56-year-old Chinese male with previously diagnosed prostatic stromal tumour of uncertain malignant potential (STUMP) presented with urinary retention 6 years after transurethral resection of prostate (TURP). TREATMENT AND OUTCOME: Cystoscopy showed a papillary tumour of the prostatic urethra causing near-complete obstruction. Repeat TURP was performed. He has been asymptomatic since. CONCLUSION: There has been fewer than 100 cases of this lesion reported worldwide. Definitive treatment is not well established. Longterm follow-up to monitor progression and possible recurrence is required, and repeat TURP or radical surgery may be necessary.

Viral vector-mediated transduction of a modified thrombospondin-2 cDNA inhibits tumor growth and angiogenesis.

Gene therapy represents a possible alternative to the chronic delivery of recombinant antiangiogenic proteins to cancer patients. We have constructed retroviral and adenoviral vectors that express murine N-terminal fragments of thrombospondin-2 (NfTSP2), a potent endogenous inhibitor of tumor growth and angiogenesis. To test the possibility of anticancer gene therapy using NfTSP2, we tested whether an ex vivo retrovirus-mediated procedure could be used for the treatment of tumors. The treatment of tumor-bearing mice with syngenic immortalized cell lines expressing NfTSP2 led to a tumor volume reduction up to 70% as compared with the controls (P<0.005). In addition, the established tumors were eradicated in 40% of the mice treated with NfTSP2-expressing cells. Furthermore, the intratumoral injection of the NfTSP2-expressing adenoviral vector to the human squamous cell carcinoma in nude mice resulted in a significant reduction of the growth rates and the volumes of the carcinoma (P<0.05). Immunohistochemical staining of the tumors indicated that the total area and the average size of tumor vessels were significantly reduced in the treatment group versus the controls (P<0.05). In conclusion, the present study clearly demonstrates that the viral vector-mediated transfer of the NfTSP2 gene could inhibit the growth of tumors by perturbing tumor-associated angiogenesis.

Extracorporeal shockwave lithotripsy and intracorporeal lithotripsy for proximal ureteric calculi--a comparative assessment of efficacy and safety.

INTRODUCTION: Extracorporeal shockwave lithotripsy (ESWL) is the treatment modality of choice of many urologists for proximal ureteric calculi. In this study, we compared the efficacy and safety of ESWL versus ureteroscopy with holmium laser lithotripsy for the treatment of this group of stones. MATERIALS AND METHODS: Between May 1999 and October 2000, 50 patients had ESWL and another 51 patients underwent ureteroscopy with holmium laser lithotripsy for proximal ureteric calculi. The two groups were similar in age, sex ratio and stone size. ESWL was performed with the Dornier Compact lithotriptor whereas holmium laser lithotripsy was performed via retrograde ureteric access with a Wolf 7.5 Fr semirigid ureteroscope. RESULTS: Ureteroscopy with holmium laser lithotripsy was significantly better in terms of the mean procedure time (56 min in ESWL; 25 min in ureteroscopy; P < 0.001) and the 1-month stone free rate (50% in ESWL; 80% in ureteroscopy; P = 0.001). The 3-month stone free rate was also higher for ureteroscopy (78% in ESWL; 90% in ureteroscopy) but this difference was not statistically significant (P = 0.09). Minor complications of steinstrasse (6%) occurred in ESWL and proximal stone migration (8%) occurred during ureteroscopy. CONCLUSION: Ureteroscopy with holmium laser lithotripsy is a viable and safe alternative to ESWL for the management of proximal ureteric calculi.

Protection against collagen-induced arthritis by intramuscular gene therapy with an expression plasmid for the interleukin-1 receptor antagonist.

The interleukin-1 receptor antagonist (IL-1Ra) is an endogenous protein that can prevent the binding of IL-1 to its cell-surface receptors. Among a number of techniques for gene transfer in vivo, the direct injection of naked DNA into muscle is simple, inexpensive and safe. In this study, we evaluated the potential of intramuscular gene therapy with plasmid DNA containing the cDNA for IL-1Ra in the prevention of murine collagen-induced arthritis (CIA). DBA/1 mice were immunized with bovine type II collagen. At 4 weeks after the initial immunization, expression plasmid for IL-1Ra was injected into four selected sites in the thigh and calf muscles of DBA/1 mice. Control mice received the same plasmid, but lacking the IL-1Ra coding sequence. Macroscopic analysis of paws for redness, swelling and deformities showed that the onset of moderate to severe CIA in the paws of mice injected with IL-1Ra DNA was significantly prevented (P<0.05). In addition, both the synovitis and the cartilage erosion in knee joints were dramatically reduced in mice treated with IL-1Ra DNA (P<0.05). The expression of IL-1beta was significantly decreased in the ankle joints of mice treated with IL-1Ra (P<0.01). Interestingly, the levels of IL-1Ra in sera and joints after intramuscular injection of IL-1Ra DNA were significantly lower than when protein had been used in previous reports, suggesting that the therapeutic effect may be achieved by an alternative mechanism(s) rather than by systemic elevation of IL-1Ra. These observations provide the first evidence that direct intramuscular injection of expression plasmid for IL-1Ra may effectively suppress the inflammatory pathology in arthritis.

Electro-gene therapy of collagen-induced arthritis by using an expression plasmid for the soluble p75 tumor necrosis factor receptor-Fc fusion protein.

Tumor necrosis factor (TNF) is a proinflammatory cytokine involved in the pathogenesis of rheumatoid arthritis, and antagonism of TNF may reduce the activity of the disease. Among a number of techniques for gene transfer in vivo, the direct injection of plasmid DNA into muscle is simple, inexpensive, and safe. In this study, we attempted to treat collagen-induced arthritis (CIA) with anti-TNF gene therapy by transferring the plasmid encoding soluble p75 TNF receptor linked to the Fc portion of human IgG1 (sTNFR:Fc) using in vivo electroporation. DBA/1 mice were immunized with bovine type II collagen and boosted with the same antigen. At 2 days after boosting, the plasmid vector containing cDNA for the sTNFR:Fc was injected into one selected site in the gastrocnemius muscle followed by electroporation. Serum levels of sTNFR:Fc reached 2.3 ng/ml on day 5 when gene expression reached its peak. Macroscopic analysis of paws for redness, swelling and deformities showed that the onset of moderate-to-severe CIA in mice treated with sTNFR:Fc was prevented on a significant level compared with the control mice (P<0.05). The beneficial effect of sTNFR:Fc DNA transfer lasted for at least 18 days following treatment. In addition, both the synovitis and the erosion of cartilage in the knee joints were dramatically reduced in mice treated with sTNFR:Fc (P<0.05). The expression of IL-1beta and IL-12 in the paw was also decreased by sTNFR:Fc treatment (P<0.01) while there was little change in the levels of IL-17 and vWF. These data showed that sTNFR:Fc expression plasmid was effective in the prevention of CIA, and in vivo electroporation-mediated gene transfer may provide a new approach to cytokine therapy in autoimmune arthritis.

STR data for the PowerPlex 16 loci for the Chinese population in Hong Kong.

Allele frequencies for the 15 STR loci included in the PowerPlex 16 kit were obtained from a sample of 247 unrelated Chinese in Hong Kong.