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Background An outbreak of Plasmodiummalariae infection among forest goers in Sanya City of Hainan Island, China was reported in 2015. In response to this outbreak, an innovative three-layer strategy (TLS) targeted forest goers was adapted based on the 1-3-7 approach. Main text Key elements of TLS are: (i) The village with five malaria cases and adjacent villages were set as the first layer. All residents including forest goers were taken as the high-risk population (HRP). Active case detection (ACD) by blood smear microscopy and PCR was selected as the primary measure, and passive case detection (PCD) as complementary measure. One case was identified under TLS implementation. (ii) The township with cases (Gaofeng Town) and the nearby towns were chosen as the second layer. Only forest goers were screened by ACD, while PCD as a routine screening method. 7831 blood smears collected by ACD and PCD and tested with negative results. (iii) The city with cases (Sanya City) and others 12 counties/county-level cities were selected as the third layer. Malaria cases were monitored passively. A total of 77,555 blood slides were screened by PCD with zero positive sample. For each layer, the malaria vector mosquitoes were monitored using light traps, cattle-baited/human-bait traps.Anophelesminimus (dominant species), An.sinensis and An.dirus were captured. Vector control measures mainly include insecticide residual spraying and long-lasting insecticide nets. The capacity of clinicians, public health practitioners and laboratory technicians has been improved through training. During 2016‒2018, TLS and chemoprophylaxis were implemented in the same areas. In the first layer, all residents were monitored by ACD, and malaria chemoprophylaxis were distributed, 89.5% of forest goers were using chemoprophylaxis against malaria. The blood smears (3126 by ACD plus 1516 by PCD) were with zero positive results. Chemoprophylaxis and ACD were offered to forest goers once a year, and PCD in residents as a complementary measure in the second and third layer, 77.8% and 95.1% of forest goers received chemoprophylaxis. In each layer, vector surveillance and control of malaria and trainings for medical staff were still in place. Conclusions TLS was effective in blocking the outbreak by P.malariae among forest goers in Hainan in malaria elimination stage. However, whether it could prevent the malaria resurgence in the post-elimination phase needs to be further assessed. Graphical Abstract

As viral genomic imprints in host genomes, endogenous viral elements (EVEs) shed light on the deep evolutionary history of viruses, ancestral host ranges, and ancient viral-host interactions. In addition, they may provide crucial information for calibrating viral evolutionary timescales. In this study, we conducted a comprehensive in silico screening of a large data set of available mammalian genomes for EVEs deriving from members of the viral family Flaviviridae, an important group of viruses including well-known human pathogens, such as Zika, dengue, or hepatitis C viruses. We identified two novel pestivirus-like EVEs in the reference genome of the Indochinese shrew (Crocidura indochinensis). Homologs of these novel EVEs were subsequently detected in vivo by molecular detection and sequencing in 27 shrew species, including 26 species representing a wide distribution within the Crocidurinae subfamily and one in the Soricinae subfamily on different continents. Based on this wide distribution, we estimate that the integration event occurred before the last common ancestor of the subfamily, about 10.8 million years ago, attesting to an ancient origin of pestiviruses and Flaviviridae in general. Moreover, we provide the first description of Flaviviridae-derived EVEs in mammals even though the family encompasses numerous mammal-infecting members. This also suggests that shrews were past and perhaps also current natural reservoirs of pestiviruses. Taken together, our results expand the current known Pestivirus host range and provide novel insight into the ancient evolutionary history of pestiviruses and the Flaviviridae family in general. ispartof: MOLECULAR BIOLOGY AND EVOLUTION vol:39 issue:10 ispartof: location:United States status: published

There are many DNA taxonomy methods available, but single-locus data coupled with distance-based analyses still dominate species identification in practice. Genetic distance thresholds are often used for assigning genetic diversity into entities corresponding to species but are criticized for non-universality or arbitrary. This study investigated an alternative approach for determining genetic thresholds that used iteratively sister-clade clustering for sister species boundary detection within a phylogenetic framework. This method was separately applied to two close subfamilies of Callosciurinae and Sciurinae in Rodentia using the cyt b gene. Our results showed that genetic thresholds for Callosciurinae and Sciurinae were 4.0% and 6.0%, respectively, indicating that the optimized thresholds could be lineage-specific. The use of these thresholds for taxa partitioning yielded a very similar putative species number as the prevailing ABGD method, and increased species diversity by 74.2% and 20.7% in Callosciurinae and Sciurinae, respectively. This suggested that additional cryptic species were present that warrant further investigation. We further tested the performance limitation of our method by simulating the impacts of tree construction and sampling limitation. The results showed that it preformed equally well for different trees but failed to work when inter- or intraspecific sampling is insufficient. These findings support the feasibility of this approach as an alternative tool for species delineation when only single-locus information is available for large datasets.

Yulangsan polysaccharide (YLSPS) is derived from the root of Millettia pulchra (Benth.) Kurz var. Recent studies have postulated YLSPS as a regimen for cancer treatment. However, the underlying mechanism anti-breast cancer is still poorly unknown. The aim of this study was to examine the suppressive and apoptosis effect of YLSPS on the growth of breast cancer cell 4T1 and its possible underlying mechanism. In this study, breast cancer cell 4T1 viability and apoptosis were assessed by CCK-8 and flow cytometry, relative quantitative real-time PCR and western blot after treated with drug-serum of YLSPS. Furthermore, therapy experiments were conducted using a Balb/c mouse transplanted tumor model of breast cancer. The number of apoptotic cells and microvascular density (MVD) in the tumor tissues were assessed by TUNEL and CD34 immunostaining. Immunohistochemical assays and ELISA were used to detect the expression of VEGF, Bcl-2, Bax and Caspase-3 in the tissues. The in vitro studies showed that the drug-serum of YLSPS significantly inhibition of proliferation and effectively induced apoptosis of 4T1 cells. Oral administration of YLSPS in the breast cancer models significantly reduced the tumor volume and weight. The enhanced antitumor efficacy was associated with decreased angiogenesis, an enhanced antioxidant capacity, an increased induction of apoptosis and an inhibition of lung metastasis. These findings indicate that YLSPS significantly inhibited mouse breast cancer growth in vitro and in vivo. These data suggest that YLSPS may serve as a potential therapeutic agent for breast cancer.

Silicosis is characterized by pulmonary interstitial fibrosis that arises as a result of chronic exposure to silica. The few available treatments only delay its progression. As α-lipoic acid (ALA) has been shown to have various beneficial effects, including mitoprotective, antioxidant, and anti-inflammatory effects, we hypothesized that it may exhibit therapeutic effects in pulmonary fibrosis. Therefore, in the present study, we used a murine model of silicosis to investigate whether supplementation with exogenous ALA could attenuate silica-induced pulmonary fibrosis by improving mitochondrial function. ALA was administered to the model mice via continuous intragastric administration for 28 days, and then the antioxidant and mitoprotective effects of ALA were evaluated. The results showed that ALA decreased the production of reactive oxygen species, protected mitochondria from silica-induced dysfunction, and inhibited extracellular matrix deposition. ALA also decreased hyperglycemia and hyperlipidemia. Activation of the mitochondrial AMPK/PGC1α pathway might be responsible for these ALA-mediated anti-fibrotic effects. Exogenous ALA blocked oxidative stress by activating NRF2. Taken together, these findings demonstrate that exogenous ALA effectively prevents the progression of silicosis in a murine model, likely by stimulating mitochondrial biogenesis and endogenous antioxidant responses. Therefore, ALA can potentially delay the progression of silica-induced pulmonary fibrosis.

Ethnopharmacological relevance The root of Averrhoa carambola L. (Oxalidaceae), a traditional Chinese medicine, was mainly used in ancient times in the treatment of urinary calculi, recurrent headache and joint pain. Aim of the study Our aims were to explore the potential therapeutic effect of the extract of Averrhoa carambola L. (Oxalidaceae) roots (EACR) against hepatic fibrosis in CCl4-treated rats and to understand the underlying molecular mechanism. Materials and methods Six groups of male Sprague Dawley rats were treated as follows: vehicle (olive oil), CCl4 alone, CCl4+colchicine, CCl4+EACR 1.0 g/kg, CCl4+EACR 0.5 g/kg and CCl4+EACR 0.25 g/kg. At the end of the 12th week, biomarkers of liver function, liver fibrosis, hepatic oxidative stress and antioxidant status were assayed, and histopathological and immunohistochemical evaluation of liver tissue were conducted to investigate the liver damage and fibrosis degree. Furthermore, expressions of COL-1a1, α-SMA, TGF-β1, Smad2, smad3, Smad4 and TIMP2 were examined by qPCR and/or western blot. The expressions of apoptosis-related proteins were also detected using western blot analysis. Results EACR treatment markedly reduced the CCl4-induced elevation of serum aminotransferase activities, liver fibrosis indexes, and the extent of oxidative stress. EACR treatment also significantly reduced the accumulation of collagen and the immunostaining of α-SMA, TGF-β1 and Smad2, 4 and 7 in the liver of CCl4 treated rats. In addition, EACR treatment markedly reversed the CCl4-induced increase in mRNA expression of COL-1a1, α-SMA, TIMP2, TGF-β1, Smad2 and Smad4 and suppressed the expressions of α-SMA, TIMP2, TGF-β1, smad2, 3 and 4, BAX and cleaved caspase-3 proteins. Meanwhile, EACR treatment also significantly elevated the mRNA expression of Smad7 and the protein expression of Smad7 and Bcl-2. Conclusion These results suggest that EACR has protective activity against liver fibrosis. The anti-fibrotic activity of EACR in vivo is associated with enhanced antioxidant, apoptosis-inhibition and increased MMP-2/TIMP-2 expression ratio, and with modulation of TGF-β1/Smad signaling pathway.

Oxidative stress and inflammatory processes are proposed to mediate the development of silicosis. However, antioxidant therapy has not produced consistent results during the treatment of silicosis. α-Lipoic acid synthesized by lipoic acid synthase is a powerful anti-oxidant and helps protect mitochondria. Thus far, the effect of endogenous α-Lipoic acid on silicosis has not been elucidated yet. We established an experimental model of silicosis with wildtype and LiasH/H mice, a new antioxidant mouse model which has overexpressed Lias gene (∼150 %) relative to its wild type counterpart. We systemically examined main pathological changes of pulmonary fibrosis, and explored α-lipoic acid effects on oxidative stress, inflammatory and pulmonary fibrosis biomarkers in silica-instillated mice. In LiasH/H mice over-expression of lipoic acid alleviated the severity of major pathological alterations in the early stage of pulmonary fibrosis induced by silica compared with wild type mice. Silica significantly increased oxidative stress in both wild type and LiasH/H mice. The antioxidant defense was strengthen including increased NRF2 and LIAS production in LiasH/H mice. Relieved oxidative stress resulted in decreased inflammatory response and secretion of chemokines. LiasH/H mice reduced chronic inflammatory response and inhibition of NF-κB activity after silica instillation. The LiasH/H mouse model overexpression of lipoic acid synthase gene retarded the development of silica-induced pulmonary fibrosis. Strengthen antioxidant defense by increased lipoic acid synthase is a potential strategy for protection against silica-induced pulmonary fibrosis.

Many peninsulas in the temperate zone played an important role as refugia of various flora and fauna, and the southern Korean Peninsula also served as a refugium for many small mammals in East Asia during the Pleistocene. The Asian lesser white-toothed shrew, Crocidura shantungensis, is a widely distributed species in East Asia, and is an appropriate model organism for exploring the role of the Korean Peninsula as a refugium of small mammals. Here, we investigated phylogenetic relationships and genetic diversity based on the entire sequence of the mitochondrial cytochrome b gene (1140 bp). A Bayesian tree for 98 haplotypes detected in 228 C. shantungensis specimens from East Asia revealed the presence of three major groups with at least 5 subgroups. Most haplotypes were distributed according to their geographic proximity. Pairwise FST's and analysis of molecular variance (AMOVA) revealed a high degree of genetic differentiation and variance among regions as well as among populations within region, implying little gene flow among local populations. Genetic evidence from South Korean islands, Jeju-do Island of South Korea, and Taiwan leads us to reject the hypothesis of recent population expansion. We observed unique island-type genetic characteristics consistent with geographic isolation and resultant genetic drift. Phylogeographic inference, together with estimates of genetic differentiation and diversity, suggest that the southern most part the Korean Peninsula, including offshore islands, played an important role as a refugium for C. shantungensis during the Pleistocene. However, the presence of several refugia on the mainland of northeast Asia is also proposed.

Although increased survey efforts using improved capture methods (particularly harp traps) have greatly expanded the quantity of Kerivoula specimens available in China, the understanding of the genus has been long constrained. After the recently published revision of the hardwickii-complex with the description of K. furva and re-evaluation of occurrence of K. titania in Taiwan, the critical overview of the previous data of Chinese Kerivoula (with the exception of K. picta, a strikingly colored and unmistakable species) is imperative. To clarify the taxonomy and distribution of the hardwickii-complex in China, 40 additional specimens collected from South China were studied through detailed morphological comparisons, multivariate statistical methods and phylogenetic inference. Our results evidently classified these specimens as K. furva instead of K. titania or K. hardwickii sensu stricto and together with the critical review of literature data indicate that all previous Chinese records of the two latter species were based on either misidentifications or outdated taxonomy. K. furva have so far been recorded in the Chinese provinces of Chongqing, Fujian, Guangdong, Guangxi, Hainan, Hunan, Jiangxi, Yunnan and Taiwan, but more field surveys are needed to confirm whether it could be found in other nearby provinces.

Activation of phosphatase and tensin homolog (PTEN) is known to induce cell apoptosis. MicroRNA-374a (miR-374a), which can suppress PTEN expression, has been found abnormally expressed in inflammatory bowel disease (IBD). Fortunellin is a citrus flavonoid that is a potential anti-inflammation agent in inflammatory diseases. The present study investigated the effects and mechanisms underlying fortunellin-induced inhibition of PTEN in IBD. Colitis was established in rats by the intracolonic administration of 2,4,6-trinitrobenzene sulfonic acid to mimic human ulcerative colitis, which is the main type of IBD. miR-374a expression was measured by quantitative real-time polymerase chain reaction, and the regulation of PTEN by miR-374a was evaluated by dual luciferase reporter assay. Western blotting was used to measure the corresponding protein expression. Fortunellin ameliorated colitis symptoms, including excessive inflammation and oxidative stress. Fortunellin decreased epithelial cell apoptosis through inhibiting PTEN expression in colitis. Fortunellin-induced downregulation of PTEN could be counteracted by miR-374a depletion. Moreover, knockdown of miR-374a in vivo partly inhibited the effects of fortunellin on rat colitis. In conclusion, PTEN inhibition contributes to the amelioration effects of fortunellin on colitis. It was confirmed that fortunellin targets miR-374a, which is a negative regulator of PTEN. This study provides novel insights into the pathological mechanisms and treatment alternatives of colitis.

Color variation in sea cucumber is one of the most crucial traits affecting price and taste in East Asian countries. However, the relationship and taxonomic status of the three color variants are still unclear. We used 14 samples that covered all three color variants and their geographic distributions, to construct the first phylogeny for the color variants based on the complete mitochondrial genome sequence and a number of tree-building methods (maximum parsimony (MP), maximum likelihood (ML), and Bayesian inference (BI)). The divergence times within color variants were estimated by the Bayesian molecular clock approach using the BEAST program. Our results showed that the color variants were not monophyletic in the well-resolved phylogenetic tree, which strongly refuted their separate species status. The molecular dating estimate revealed that the sea cucumber was a young group, which originated in the early Miocene period (22.03 mya) and rapidly diverged after the late Miocene period. It is interesting that individuals within each variant or geographic distribution were not always closely related and thus did not share a common origin. We propose that although they differ in body color, the three color morphs all belong to a single species of Apostichopus japonicus and the historical marine climate and the hydrographic complexity of the ocean currents could be responsible for their present distribution patterns.

Stretchable electronic devices have great potential for serving as bioelectrical interfaces due to their better deformability and modulus match with biological organs. However, surface modification, which is usually applied to enhance the capability of sensing and stimulating, as well as biocompatibility, may cause problems since their stretchability highly depends on the surface structure. In this work, stretchable nanocrack gold (SNCG) electrodes were fabricated, which can be stretched by a maximum 120% uniaxial strain while maintaining their electrical conductivity. We found that the electrodes lost their stretchability after surface modification of an additional continuous platinum layer, which was found to selectively weld or fully cover the nanocracks, consequently eliminating its crack structure. To address this issue, we designed a complex structure of dispersed, porous nanoislands landing on the SNCG film, which was further demonstrated as capable of maintaining the stretchability of electrodes while allowing the reshaping of cracks. Moreover, stretchable microelectrode arrays were then developed with this complex structure. Animal experiments demonstrated their capability of conformally wrapping on a rat brain cortex and effectively monitoring an intracranial electroencephalogram under deformation. In addition, their impedance can be precisely controlled by modulating the dispersity, diameter, and aspect ratio of individual nanoislands. This complex structure has great potential for developing highly stretchable, multiplexing sensors, allowing stiff materials to land on a stretchable conducting surface with maintenance of stretchability and controllable functional area.

Three distinct bamboo bat species (Tylonycteris) are known to inhabit tropical and subtropical areas of Asia, i.e., T. pachypus, T. robustula, and T. pygmaeus. This study performed karyotypic examinations of 4 specimens from southern Chinese T. p. fulvidus populations and one specimen from Thai T. p. fulvidus population, which detected distinct karyotypes (2n=30) compared with previous karyotypic descriptions of T. p. pachypus (2n=46) and T. robustula (2n=32) from Malaysia. This finding suggested a cryptic Tylonycteris species within T. pachypus complex in China and Thailand. Morphometric studies indicated the difficulty in distinguishing the cryptic species and T. p. pachypus from Indonesia apart from the external measurements, which might be the reason for their historical misidentification. Based on 623 bp mtDNA COI segments, a phylogeographic examination including T. pachypus individuals from China and nearby regions, i.e., Vietnam, Laos, and Cambodia, was conducted to examine the population genetic structure. Genealogical and phylogeographical results indicated that at least two diverged lineages existed in these regions (average 3.4 % of Kimura 2-parameter distances) and their population structure did not match the geographic pattern. These results suggested that at least two historical colonizations have occurred by the cryptic species. Furthermore, through integration of traditional and geometric morphological results, morphological differences on zygomatic arches, toothrows and bullae were detected between two lineages in China. Given the similarity of vegetation and climate of Guangdong and Guangxi regions, we suggested that such differences might be derived from their historical adaptation or distinct evolutionary history rather than the differences of habitats they occurred currently.

The complete mitochondrial genome of the maritime striped squirrel (Tamiops maritimus) was first sequenced and characterized. The genome was 16 523 bp in length, and the composition and the arrangement of genes were analogous to other rodents. The sequence of T. maritimus was used to construct phylogenetic tree with additional mitochondrial genomes of seven sciurid species available on GenBank. Phylogenetic result indicated that T. maritimus has a close relationship with T. swinhoei. Our mitochondrial genome data may provide information for species identification, diversity evaluation, and other studies about this genus.

The complete mitochondrial genome of the greater bandicoot rat (Bandicota indica) was first sequenced and characterized. The genome was 16 326 bp in length, the composition and arrangement of its genes were analogous to other rodents. To confirm the phylogenetic position of B. indica, the mitochondrial nucleotide sequence data of other 20 Rodentia species were used to construct phylogenetic tree by maximum likelihood. Phylogenetic analysis demonstrated that genera Bandicota and Rattus were sister taxa, and B. indica was closer to the genus Rattus than to genera Niviventer and Leopoldamys. The mitochondrial genome of B. indica presented in this study can provide useful information for species delimitation, taxonomic and phylogenetic analyses as well as other studies of the species.

Karyotypes and chromosomal data are presented for eight bat species representing two families (Rhinolophidae and Vespertilionidae) from Hainan Island, China. The species investigated were Rhinolophus lepidus (2n = 62, FN = 60), R. pusillus (2n = 62, FN = 60), R. affinis (2n = 62, FN = 60), R. sinicus (2n = 36, FN = 60), Myotis horsfieldi (2n = 44, FN = 52), Pipistrellus abramus (2n = 26, FN = 44), Miniopterus australis (2n = 46, FN = 50) and M. schreibersii (2n = 46, FN = 50). The karyotype of Rhinolophus lepidus is reported for the first time.

Rationally designed sinusoidal gold nanobelts are fabricated as stretchable electrodes, and they do not show obvious change of resistance under large deformation after 10,000 cyclic stretching/relaxing processes. As a proof of concept, they are successfully used to record intracranial electroencephalogram or electrocorticogram signals from rats.

Objective To understand the current epidemiological status of Angiostrongylus cantonensis in Hainan island and provide the scientific evidences for prevention and control of the disease. Methods The investigation on nature infection condition of intermediate hosts and final hosts of Angiostrongylus cantonensis were carried out in five counties, Hainan Province, and enzyme linked immunosorbent assay (ELISA) was employed to detect antibodies of Angiostrongylus cantonensis in human hosts. Results A total of 1612 molluscs were examined, 21.3% of which harbored L3 of Angiostrongylus cantonensis . Among them, the infection rates of Pomacea canaliculata, Achatina fulica, Cepaea and Phlegm bilineatus were 12.36%(64/518), 22.66%(121/534), 20.93%(9/43), and 28.24%(146/517), respectively. Of the 118 rats trapped, 13 Rattus norvegicus were found to be infected with Angiostrongylus cantonensis . A total of 459 serum samples were collected and tested. 92 serum samples were Angiostrongylus cantonensis antibody-positive. Conclusions The survey revealed a wide distribution of Angiostrongylus cantonensis in Hainan island. Pomacea canaliculata and Achatina fulica are main intermediate hosts of Angiostrongylus cantonensis . Rattus novegicus is a nature definitive host, indicating that a considerable number of people are at risk of angiostrongyliasis. Health education, rigorous food inspection and surveillance are all needed to prevent angiostrongyliasis outbreaks in future.

Accurate species delimitation in Lepus was often hindered by highly conserved morphology and frequent introgression. In this study, we used rigorous molecular species delimitation methods to evaluate the taxonomic status of Hainan hare (Lepus hainanus) which has been traditionally identified as a distinct species, or a subspecies of Burmese hare (L. peguensis). The genetic distance and phylogenetic network support L. hainanus and L. peguensis are conspecific. However, the phylogenetic species concept and Bayesian species delimitation analysis based on combined mtDNA supported they are different species. The discordance between different methods can be explained by different species criterion. By taking into account our conflict results, we hold the opinion that adoption of the phylogenetic species concept and Bayesian species delimitation analysis would increase the risk of taxonomic inflation of island biota or otherwise spatially isolated population. Conservatively, we suggest that L. hainanus and L. peguensis are conspecific based on the results of our genetic divergence and phylogenetic network exclusively.

The complete mitochondrial genome of Petaurista hainana from Hainan Island of China is 16,502 bp in length. Similar to most other mammals, it contains 13 protein-coding genes, 2 rRNAs, 22 tRNAs and 2 non-coding regions. The overall base composition of the H-strand is 31.77% A, 26.31% C, 13.31% G and 28.61% T. The base composition clearly showed the A-T skew. The mitochondrial genome of P. hainana presented in this report will be useful for species designation, conservation and phylogenetic study in Sciuridae.

To the Editor: Hendra virus (HeV) and Nipah virus (NiV), the only known members of the genus Henipavirus, are 2 emerging paramyxoviruses that are highly pathogenic in a variety of vertebrate animals, including humans (1). Since the initial discovery of the viruses in Australia and Malaysia (2,3), sporadic HeV outbreaks have been reported from 1995 to 2007 in Australia (4), and regular NiV outbreaks have occurred in Bangladesh (5) and India (6). Numerous frugivorous bat species (genus Pteropus), and some insectivous bat species have been found to be reservoir hosts of henipaviruses in Australia and Asian countries (7–9). In this study conducted during 2004–2007, bats were trapped within their natural habitat from 10 provinces in mainland People’s Republic of China. Serum, pharyngeal, and fecal swab samples were collected and stored as described previously (10). An ELISA was developed to detect antibodies to the NiV nucleocapsid (N) and attachment glycoprotein (G) proteins. For confirmation, ELISA-positive samples were tested by using Western blot against a recombinant NiV G fragment (aa 71–193) fused with the maltose-binding protein. Virus neutralization tests were conducted with live NiV and HeV under Biosaftey Level 4 containment in Australia. In addition, a surrogate neutralization test was developed by using recombinant env– HIV-1, pseudotyped with NiV G and F. RNA was extracted by using the QIA amp Viral RNA Mini Kit (QIAGEN, Hilden, Germany). Reverse transcription–PCR (RT-PCR) was performed by using primers against the NiV N gene as described previously (3). In total, 692 bat serum specimens were screened for antibody to NiV N or G protein (or both) by ELISA, and 33 were positive (Appendix Table). These specimens were from 9 of the 23 bat species examined in this study. Of the 33 serum samples reactive in ELISA, 25 with sufficient quantity left were further tested by Western blot, and 17 of 25 serum samples were reactive with MBP-NiV G fusion fragment, but not with the control MBP. None of the samples inhibited entry of NiV F/G-pseudotyped virus or neutralized either HeV or NiV. No NiV-specific RNA was detected by RT-PCR among 479 fecal swab samples and 67 throat swab samples tested; therefore, virus isolation was not attempted. This study systematically investigated NiV presence among bats in China. The detection of henipavirus antibody suggests that several bat species have been exposed to NiV or a closely related virus. The prevalence of antibody was especially prominent among Myotis species from Yunnan Province. Antibody was detected in samples from 3 of 4 Myotis species captured in the same location in 2006 and 2007. A relatively high prevalence of henipavirus antibody was also found among Rousettus leschenaultia samples from Hainan Province in 2007. Notably, Yunnan and Hainan are both located in southern China. Although pteropid bats are not found in China, these data suggest henipaviruses could be introduced into China by other susceptible bat species that overlap their habitat with pteropid bats in neighboring countries. Several possibilities may explain the failure to detect neutralizing antibodies. One might be the unique immune response among those nonpteropid bats, which results in a low level of neutralizing antibodies that are difficult to detect in the current assay systems. Alternatively, and perhaps more likely, >1 Nipah-like viruses could be circulating among the bat populations sampled in this study, producing antibodies that cross-react with, but do not neutralize, the prototype Malaysian NiV virus isolate. This phenomenon has been observed previously by our group for severe acute respiratory syndrome (SARS)–like viruses in horseshoe bats, whose sera cross-reacted with, but did not neutralize, the SARS virus in humans (10). Obtaining serologic evidence of viruses in bat populations is typically more successful as a screening tool than either nucleic acid based assays or virus isolation; this is likely attributable to the often low-level of virus replication, the transient nature of the infection in bats, or both. The inability to amplify NiV sequences may have been attributable to the viral RNA present among these samples being below the threshold of detection in our assay or to the absence of infection in the population at the time of sampling. In addition, the primers used in the PCR may target regions of the NiV N protein that exhibit substantial sequence divergence in a Nipah-like virus. Bat species in the genera Rousettus, Myotis, Miniopterus, and Hipposideros naturally reside in trees, buildings, and caves that can be in close proximity to human residential areas, which increases the potential of transmission of zoonotic pathogens from bats to humans. The increased risk for these zoonotic infections to spread from bats to humans in areas of cohabitation is best illustrated by the repeated spillover events involving NiV in Bangladesh (5). Given the present initial evidence of exposure among bats in mainland China shown here, there is an urgent need to continue and expand surveillance studies for henipaviruses in China and elsewhere on the Asian continent.

Based on 203 specimens belonging to the Rhinolophus “pusillus group” (Mammalia: Chiroptera: Rhinolophidae), univariate and multivariate morphometric analyses using 19 characters were performed to assess the confused species taxonomy. The results indicated that R. pusillus (including calidus, parcus, and szechuanus) in the continental region and Hainan Island of China and “R. cornutus” in Japan are morphologically divergent species. Rhinolophus cornutus should be further split into R. cornutus (including orii, pumilus, and miyakonis) in the main islands of Japan, the Amami and Okinawa Group of the central Ryukyu Archipelago, and Miyako Group of the southern Ryukyus; and R. perditus and R. imaizumii from the Yaeyama Group in the southern Ryukyus. Rhinolophus monoceros from Taiwan is morphologically more similar to species in Japan than to R. pusillus. In addition to R. pusillus, another form that is morphologically similar to species in Japan was recognized from Langzhong in Sichuan Province; this may represent an undescribed species, and further examination is necessary to determine its taxonomic status. Specimens from Guang’an in Sichuan Province, China, are also different from the others, and are characterized by the smallest skull size. Although further studies are required, these specimens were tentatively identified as R. subbadius.

The complete sequence of the mitochondrial genome of the mandarin vole (Lasiopodomys mandarinus) was completed and annotated in this study. The circular genome is 16,375 bp in length and contains the typical 37 genes that are arranged in the same order as that of the putative ancestor of vertebrate, 13 protein-coding genes, 22 tRNA genes, 2 rRNA genes and 2 non-coding regions. This study will provide genetic resource to clarify the taxonomic position of genus Lasiopodomys.

The complete mitochondrial genome of black giant squirrel (Ratufa bicolor) from Hainan Island was sequenced and characterized in detail. The 16,563 bp genome was composed of 13 protein-coding genes, 22 tRNAs, 2 rRNAs and 2 non-coding regions. The mitochondrial genome of R. bicolor presented in this report will be useful for species identification, conversation and clarifying the controversial taxonomic status of genus Ratufa.

We investigated the menstrual cycle of wild fulvous fruit bats (Rousettus leschenaulti), focusing on changes in the endometrial and ovarian structure and pituitary and steroid hormones. The menstrual cycle lasts for 33 days in bats studied in their natural habitat and in captivity. Vaginal bleeding was restricted to a single day (Day 1). A preovulatory follicle was found in the ovary on Day 18 when the levels of LH and FSH reached their maxima, accompanied by a thickened endometrium. On Day 24, serum levels of progesterone and estradiol-17 were also maximal, and uterine glands increased in size. After that, the levels of progesterone dropped precipitously, leading to menstrual bleeding. Both the morphologic and hormonal changes observed in fulvous fruit bats during the menstrual cycle resemble similar changes in humans. Fulvous fruit bats may be useful nonprimate laboratory models to study menstruation and menstrual dysfunction.

To explore a new method in the gene therapy of human laryngocarcinoma.The NP, HN and F gene of the Newcastle disease virus were cloned into the pcDNA3 by RT-PCR. The human laryngeal squamous carcinoma model in nude mice had been established by transplantation of the Hep-2 cell line. The nude mice had been grouped two in the third week of the Hep-2 cell line had been transplanted. The admixture of the pcDNA3-NP and the pcDNA3-HN and pcDNA3-F were injected into the tumor in the experimental group, the equal weights pcDNA3 was injected into the tumor in the control group. The microstructural and ultra-micro-structural changes of carcinoma were observed under light and electron microscopes. The chromatosome DNA were extracted from all tissue of the nude mice, the DNA was tested to do template by PCR, whether or not exist the NP gene. The nude mice serums were examined by the method of enzyme-linked immunosorbent assay (ELISA) for the antigen of the DNA V4.There were statistically significant difference in the mice weight, the tumor weight, the tumor volume and the restrained tumor rate between the experimental group and the control group (each P0.01). A nude mouse died in the control group, the others were very asthenic. The tumor looked like nodosity and the tumor surface were defeated. The tumor in the experimental group were smaller than that in the control group, the tumors were round shape and hard and not adhered on the skin and deep part constitution. The necrosis and apoptosis of the tumor cell had been found under light and electron microscopes. The NP gene had not been explored in all tumor tissue by PCR in the experimental group. The experimental group was positive in ELISA, The control group was negative.The NP, HN and F gene could play a role in treating tumor.

Background In the island of Hainan, the great majority of malaria cases occur in mountain worker populations. Using the behavioral change communication (BCC) strategy, an interventional study was conducted to promote mountain worker malaria prevention at a test site. This study found the methods and measures that are suitable for malaria prevention among mountain worker populations. Methods During the Plasmodium falciparum elimination stage in Hainan, a representative sampling method was used to establish testing and control sites in areas of Hainan that were both affected by malaria and had a relatively high density of mountain workers. Two different methods were used: a BCC strategy and a conventional strategy as a control. Before and after the intervention, house visits, core group discussions, and structural surveys were utilized to collect qualitative and quantitative data regarding mountain worker populations (including knowledge, attitudes, and practices [KAPs]; infection status; and serological data), and these data from the testing and control areas were compared to evaluate the effectiveness of BCC strategies in the prevention of malaria. Results In the BCC malaria prevention strategy testing areas, the accuracy rates of malaria-related KAP were significantly improved among mountain worker populations. The accuracy rates in the 3 aspects of malaria-related KAP increased from 37.73%, 37.00%, and 43.04% to 89.01%, 91.53%, and 92.25%, respectively. The changes in all 3 aspects of KAP were statistically significant (p 0.05). Furthermore, in the testing areas, both the percentage testing positive in the serum malaria indirect fluorescent antibody test (IFAT) and the number of people inflicted decreased more significantly than in the control sites (p