Molecular epidemiology of enteroviruses associated with severe hand, foot and mouth disease in Shenzhen, China, 2014-2018.

In this study, we investigated the epidemiology and molecular characteristics of enteroviruses associated with severe hand, foot and mouth disease (HFMD) in Shenzhen, China, during 2014-2018. A total of 137 fecal specimens from patients with severe HFMD were collected. Enterovirus (EV) types were determined using real-time reverse transcription polymerase chain reaction (RT-PCR), RT nested PCR, and sequencing. Sequences were analyzed using bioinformatics programs. Of 137 specimens tested, 97 (70.8%), 12 (8.8%), and 10 (7.3%) were positive for EV-A71, coxsackievirus A6 (CVA6), and CVA16, respectively. Other pathogens detected included CVA2 (2.9%, 4/137), CVA10 (2.9%, 4/137), CVA5 (0.7%, 1/137), echovirus 6 (E6) (0.7%, 1/137) and E18 (0.7%, 1/137). The most frequent complication in patients with proven EV infections was myoclonic jerk, followed by aseptic encephalitis, tachypnea, and vomiting. The frequencies of vomiting and abnormal eye movements were higher in EV-A71-infected patients than that in CVA6-infected or CVA16-infected patients. Molecular phylogeny based on the complete VP1 gene revealed no association between the subgenotype of the virus and disease severity. Nevertheless, 12 significant mutations that were likely to be associated with virulence or the clinical phenotype were observed in the 5'UTR, 2Apro, 2C, 3A, 3Dpol and 3'UTR of CVA6. Eight significant mutations were observed in the 5'UTR, 2B, 3A, 3Dpol and 3'UTR of CVA16, and 10 significant mutations were observed in the 5'UTR, VP1, 3A and 3Cpro of CVA10. In conclusion, EV-A71 is still the main pathogen causing severe HFMD, although other EV types can also cause severe complications. Potential virulence or phenotype-associated sites were identified in the genomes of CVA6, CVA16, and CVA10.

Molecular surveillance of coxsackievirus A16 reveals the emergence of a new clade in mainland China.

Coxsackievirus A16 (CV-A16) of the genotypes B1a and B1b have co-circulated in mainland China in the past decades. From 2013 to 2017, a total of 3,008 specimens from 3,008 patients with mild hand, foot, and mouth disease were collected in the present study. Viral RNA was tested for CV-A16 by a real-time RT-PCR method, and complete VP1 sequences and full-length genome sequences of CV-A16 strains from this study were determined by RT-PCR and sequencing. Sequences were analyzed using a series of bioinformatics programs. The detection rate for CV-A16 was 4.1%, 25.9%, 10.6%, 28.1% and 12.9% in 2013, 2014, 2015, 2016 and 2017, respectively. Overall, the detection rate for CV-A16 was 16.5% (497/3008) in this 5-year period in Shenzhen, China. One hundred forty-two (142/155, 91.6%) of the 155 genotype B1 strains in the study belonged to subgenotype B1b, and 13 (13/155, 8.4%) strains belonged to subgenotype B1a. Two strains (CVA16/Shenzhen174/CHN/2017 and CVA16/Shenzhen189/CHN/2017) could not be assigned to a known genotype. Phylogenetic analysis of these two strains and other Chinese CV-A16 strains indicated that these two CV-A16 strains clustered independently in a novel clade whose members differed by 8.4%-11.8%, 8.4%-12.1%, and 14.6%-14.8% in their nucleotide sequences from those of Chinese B1a, B1b, and genotype D strains, respectively. Phylogenetic analysis of global CV-A16 strains further indicated that the two novel CV-A16 strains from this study grouped in a previously uncharacterized clade, which was designated as the subgenogroup B3 in present study. Meanwhile, phylogenetic reconstruction revealed two other new genotypes, B1d and B4, which included a Malaysian strain and two American strains, respectively. The complete genome sequences of the two novel CV-A16 strains showed the highest nucleotide sequence identity of 92.3% to the Malaysian strain PM-15765-00 from 2000. Comparative analysis of amino acid sequences of the two novel CV-A16 strains and their relatives suggested that variations in the nonstructural proteins may play an important role in the evolution of modern CV-A16.

In Situ Bioorthogonal Metabolic Labeling for Fluorescence Imaging of Virus Infection In Vivo.

Optical fluorescence imaging is an important strategy to explore the mechanism of virus-host interaction. However, current fluorescent tag labeling strategies often dampen viral infectivity. The present study explores an in situ fluorescent labeling strategy in order to preserve viral infectivity and precisely monitor viral infection in vivo. In contrast to pre-labeling strategy, mice are first intranasally infected with azide-modified H5N1 pseudotype virus (N3 -H5N1p), followed by injection of dibenzocyclooctyl (DBCO)-functionalized fluorescence 6 h later. The results show that DBCO dye directly conjugated to N3 -H5N1p in lung tissues through in vivo bioorthogonal chemistry with high specificity and efficacy. More remarkably, in situ labeling rather than conventional prelabeling strategy effectively preserves viral infectivity and immunogenicity both in vitro and in vivo. Hence, in situ bioorthogonal viral labeling is a promising and reliable strategy for imaging and tracking viral infection in vivo.

Genomic characteristics of coxsackievirus A8 strains associated with hand, foot, and mouth disease and herpangina.

Coxsackievirus A8 (CV-A8), a member of the genus Enterovirus of the family Picornaviridae, can cause a variety of infectious diseases, such as hand, foot and mouth disease (HFMD), herpangina (HA), encephalitis, paralysis, myelitis, and meningitis. This is a first report of complete genome sequences of CV-A8 strains associated with HFMD/HA since the prototype strain Donovan was identified in 1949. The complete genome sequences of eight new CV-A8 strains showed 19.2 %-20.6 % nucleotide differences when compared to the prototype strain Donovan, and 81.5 %-99.9 % similarity to each other. The topology of a polyphyletic tree based on complete capsid protein gene sequences indicated that the new CV-A8 strains and Donovan are monophyletic. However, seven CV-A8 strains clustered with CV-A10 and CV-A2 in the 5'UTR and P2 region, respectively. In the P3 region, three and four CV-A8 strains grouped with CV-A6 and CV-A2, respectively. Seven CV-A8 strains segregated from Donovan and grouped in a separate lineage in the 3'UTR. The strain CVA8/SZ266/CHN/2014 was most similar to EV71 in the nonstructural proteins regions. Phylogenetic analysis classified worldwide CV-A8 isolates into four distinct clusters, and almost all Chinese and Thai CV-A8 strains evolved independently in their respective lineages, which indicated geographical evolution of CV-A8.

Development of Lateral Flow Immunoassay for Antigen Detection in Human Angiostrongylus cantonensis Infection.

Angiostrongyliasis is difficult to be diagnosed for the reason that no ideal method can be used. Serologic tests require specific equipment and are not always available in poverty-stricken zone and are time-consuming. A lateral flow immunoassay (LFIA) may be useful for angiostrongyliasis control. We established a LFIA for the diagnosis of angiostrongyliasis based on 2 monoclonal antibodies (mAbs) against antigens of Angiostrongylus cantonensis adults. The sensitivity and specificity were 91.1% and 100% in LFIA, while those of commercial ELISA kit was 97.8% and 86.3%, respectively. Youden index was 0.91 in LFIA and 0.84 in commercial ELISA kit. LFIA showed detection limit of 1 ng/ml of A. cantonensis ES antigens. This LFIA was simple, rapid, highly sensitive and specific, which opened an alternative approach for the diagnosis of human angiostrongyliasis.

Genome-wide screen of promoter methylation analysis of ES cells and ES derived epidermal-like cells.

Embryonic stem cells (ESCs) are a population of pluripotent cells which can differentiate into different cell types. However, there are few reports with regard to differentiate ESCs into epidermal cells in vitro. In this study, we aimed to investigate differentially methylated promoters involved in process of differentiation from ESCs into epidermal-like cells (ELCs) induced by human amnion. We successfully induced ESCs into ELCs, which expressed the surface markers of CK19, CK15 and ß1-integrin. With MeDIP-chip arrays, we identified 3435 gene promoters to be differentially methylated, involving 894 HCP (high CpG-containing promoter), 974 ICP (intermediate CpG-containing promoter) and 1567 LCP (low CpG-containing promoter) among all the 17,500 DNA methylation regions of gene promoters in both ESCs and ELCs. Gene oncology and pathway analysis demonstrated that these genes were involved in all the three categories of GO enrichment analysis, including biological process, molecular function and cellular component. All these data suggested that embryonic stem cells can differentiate into epidermal-like cells and promoter methylation is of great importance in this process.

Derivation of human embryonic stem cell lines without any exogenous growth factors.

Human embryonic stem cell (hESC) lines are traditionally derived through immunosurgery. Their maintenance in culture requires the presence of mouse embryonic fibroblasts (MEFs) as feeder cells and media supplemented with basic fibroblast growth factor (bFGF) or other growth factors-both of which might introduce animal-derived culture components. The drawbacks associated with immunosurgery, MEF co-culture, and the cost of growth factors necessitate the exploration of a xeno-free method to maintain the self-renewal capacity of hESCs. Here, we describe an isolation method for the human inner cell mass (ICM), which was then cultured in the absence of exogenous growth factors and in the presence of human foreskin fibroblasts (HFFs) as feeder cells. Three hESC lines were obtained from poor-quality embryos by this near-xeno-free protocol. After culturing for more than 10 months, the hESCs retained normal morphology, expressed all expected cell surface markers, could differentiate to embryoid bodies upon culture in vitro, and formed teratomas in vivo. Furthermore, secretion of bFGF by HFFs was observed. In conclusion, this is the first study to describe an inexpensive, xeno-free culture system for the isolation and maintenance of hESCs that does not require bFGF supplementation.

[Insulin-like growth factor-1 gene therapy improves the levels of mRNA and protein of endothelial nitric oxide synthase in aging related erectile dysfunction in rats].

OBJECTIVE: To explore the effects of gene transfer of insulin like growth factor-1 (IGF-1) on the penis of senile rats and the altered levels of mRNA and protein of endothelial nitric oxide synthase (eNOS). METHODS: Ten young (4 months) and 20 senile (24 months) Sprague-Dawley male rats were selected. The senile rats were divided into 2 groups: phosphate buffer solution (PBS)-only (n = 10) and 100 µg IGF-1 plasmid treatment group (n = 10). After a 4-week injection of IGF-1, the responses of intracavernous pressure (ICP) with electrical stimulation to the cavernous nerve and systemic mean arterial pressure (MAP) were evaluated. In the control and transfected senile rats, the levels of eNOS mRNA and protein were examined by reverse transcription-polymerase chain reaction (RT-PCR) and Western blot respectively. RESULTS: The ICP/MAP and total ICP were significantly higher in the young control group versus the PBS-only group at Week 4 (P < 0.05). The ICP/MAP and total ICP were significantly higher in the young control group and the 100 µg IGF-1 treatment group versus the PBS-only group at Week 4 (P < 0.05). The levels of mRNA and protein of eNOS were higher in the 100 µg IGF-1 treatment group versus the PBS-only group at Week 4 (0.62 ± 0.16 vs 0.25 ± 0.08, 0.71 ± 0.19 vs 0.27 ± 0.09, both P < 0.05, respectively). CONCLUSION: The gene therapy of IGF-1 can ameliorate erectile functions and improve the levels of mRNA and protein of eNOS in senile rats.

The effect of influenza A (H1N1) pdm09 virus infection on cytokine production and gene expression in BV2 microglial cells.

Acute influenza infection has been reported to be associated with neurological symptoms such as influenza-associated encephalopathy (IAE). Although the pathophysiology of this condition remain unclear, neuroinflammation and associated alterations in the central nervous system (CNS) are usually induced. Microglia (MGs), CNS-resident macrophages, are generally the first cells to be activated in response to brain infection or damage. We performed reverse transcriptase droplet digital PCR (RT-ddPCR) and luminex assays to investigate virus proliferation and immune reactions in BV2 MGs infected with influenza A(H1N1)pdm09 virus. Furthermore, isobaric tags for relative and absolute quantitation (iTRAQ)-based quantitative proteomics methods were used to investigate the dynamic change in the protein expression profile in BV2 MGs to gain insight into the CNS response to influenza A (H1N1) pdm09 infection. Our results showed that the influenza A(H1N1)pdm09 virus was replicative and productive in BV2 MG cells, which produced cytokines such as interleukin (IL)-1ß, IL-6, tumour necrosis factor (TNF)-α and monocyte chemoattractant protein (MCP)-1. The expression of osteopontin (OPN) in the influenza A (H1N1) pdm09-infected BV2 MGs was upregulated at 16 and 32 h post-infection (hpi) compared to that in the control group, resulting in aggravated brain damage and inflammation. Our study indicates that OPN signalling might provide new insights into the treatment of CNS injury and neurodegenerative diseases in IAE.

Specific detection of Angiostrongylus cantonensis in the snail Achatina fulica using a loop-mediated isothermal amplification (LAMP) assay.

Angiostrongylus cantonensis, a rat lungworm, can cause eosinophilic meningitis and angiostrongyliasis in humans following ingestion of contaminated foods or intermediate/paratenic hosts with infective larvae. The snail Achatina fulica is one of the important intermediate hosts of A. cantonensis and is commonly eaten by humans in some countries. In the present study, we developed a loop-mediated isothermal amplification (LAMP) method for the specific detection of A. cantonensis in Ac. fulica. Primers for LAMP were designed based on the first internal transcribed spacer (ITS-1) of nuclear ribosomal DNA (rDNA) of A. cantonensis. Specificity tests showed that only the products of A. cantonensis were detected when DNA samples of A. cantonensis and the heterologous control samples Anisakis simplex s.s, Trichuris trichiura, Toxocara canis, Trichinella spiralis and Ascaris lumbricoides were amplified by LAMP. Sensitivity evaluation indicated that the LAMP assay is 10 times more sensitive than the conventional polymerase chain reaction (PCR) assay. The established LAMP assay is rapid, inexpensive and easy to be performed. It can be used in clinical applications for rapid and sensitive detection of A. cantonensis in snails, which has implications for the effective control of angiostrongyliasis.

Angiostrongylus cantonensis: identification and characterization of microRNAs in male and female adults.

Angiostrongylus cantonensis causes eosinophilic meningitis and eosinophilic pleocytosis in humans and is of significant socio-economic importance globally. microRNAs (miRNAs) are endogenous small non-coding RNAs that play crucial roles in gene expression regulation, cellular function and defense, homeostasis and pathogenesis. They have been identified in a diverse range of organisms. The objective of this study was to determine and characterize miRNAs of female and male adults of A. cantonensis by Solexa deep sequencing. A total of 8,861,260 and 10,957,957 high quality reads with 20 and 23 conserved miRNAs were obtained in females and males, respectively. No new miRNA sequence was found. Nucleotide bias analysis showed that uracil was the prominent nucleotide, particularly at positions of 1, 10, 14, 17 and 22, approximately at the beginning, middle and the end of the conserved miRNAs. To our knowledge, this is the first report of miRNA profiles in A. cantonensis, which may represent a new platform for studying regulation of genes and their networks in A. cantonensis.

A 25-bp ancient spliceosomal intron in the TvRab1a gene of Trichomonas vaginalis.

Spliceosomal introns play a key role in eukaryotic genome evolution and protein diversity. A large Rab GTPase family has been identified in a unicellular eukaryote Trichomonas vaginalis. However, the characteristics of introns in Rab genes of T. vaginalis have not been investigated previously. In this study, we identified a 25-bp spliceosomal intron in the T. vaginalis Rab1a (TvRab1a) gene, the smallest intron in T. vaginalis to be characterized to date. This intron contains a canonical splice site at both 5' (GT) and 3' (AG) ends, and a putative branch-point sequence (TCTAAC) that matches the Trichomonad consensus sequence of ACTAAC except for the first nucleotide. The position and phase of the TvRab1a intron are evolutionarily conserved in Rab1 homologous genes across at least five eukaryotic supergroups, including Opisthokonta, Amoebozoa, Excavata, Chromalveolata, and Plantae. These results strongly suggest that the TvRab1a intron is likely to be an ancient spliceosomal intron, and it can therefore be used as a phylogenetic marker to evaluate particular eukaryotic groupings. Identification and characterization of the TvRabla intron may provide an insight into the evolution of the large Rab repertoire in T. vaginalis.

FOXO3a is involved in the apoptosis of naked oocytes and oocytes of primordial follicles from neonatal rat ovaries.

Inhibition of the forkhead transcription factor, FOXO3a, can promote the transition from primordial to primary follicle and subsequent follicle development in mammalian ovaries. Stem cell factor (SCF) initiates anti-apoptotic signaling from its membrane receptor, c-kit, to Bcl-2 family members through PI3K/AKT in oocytes of primordial follicles. However, whether FOXO3a mediates the apoptosis of naked oocytes and oocytes of primordial follicles remains unknown. In the present study, oocytes from nests and primordial follicles from neonatal rat ovaries were cultured, and oocyte apoptosis was examined using the TUNEL technique. The pro-apoptotic action of FOXO3a and the potential signal transduction pathways were investigated using RT-PCR, Western blot, and immunocytochemistry. Culturing oocytes in the presence of SCF did not affect the level of total FOXO3a protein, but rapidly elevated the level of phosphorylated FOXO3a (indicating functional suppression). As phosphorylated FOXO3a increased, oocyte apoptosis was inhibited. The specific PI3K/Akt inhibitor, LY 294002, abolished the phosphorylation of FOXO3a and the anti-apoptotic action of SCF. SCF down-regulated the expression of p27KIP1 and pro-apoptotic factors such as Bim, Bad, and Bax, and this activity was reversed by LY 294002. SCF up-regulated the expression of MnSOD, which was also inhibited by LY 294002. However, SCF had no effect on Bcl-2 protein. These results suggest that FOXO3a is involved in oocyte apoptosis in the neonatal rat ovary, and the SCF-PI3K/Akt-FOXO3a signaling pathway mediates oocyte apoptosis and primordial follicle formation.

Selective molecularly imprinted stationary phases for bisphenol A analysis prepared by modified precipitation polymerization.

Bisphenol A (BPA)-imprinted polymeric microspheres were synthesized by modified precipitation polymerization (MPP) method. Influences of cross-linker, monomer, porogen volume, and agitation on polymerization were investigated. Proper amount of cross-linker ethyleneglycol-dimethacrylate (EGDMA) was critical to achieve narrowly dispersed microspheres. For template BPA, monomer 4-vinylpyridine (4-VP) was better than MAA to get the best imprinted effects. The optimum template/monomer ratio was 1:6. Increasing porogen volume increased size dispersity and decreased binding characters. Agitation increased coagulation and resulted in irregular particles. Microspheres with the best binding characters were used as selective stationary phase of chromatographic column to detect BPA in milk, pig urine, and chicken meat. Under optimal chromatographic conditions, the calibration graph was linear with R2 = 0.9994 in the range of 3-50 micromol/L. The LOD and LOQ were 1 and 3 micromol/L, respectively. When large amounts (20 mL or 20 g) of samples were analyzed, the recoveries ranged from 70.2 to 87.3% with RSD less than 4.85% in all samples spiked with 0.05-0.2 micromol/L BPA. The intra-day and inter-day RSD were less than 1.83 and 3.96%, respectively. Microspheres prepared by MPP are successfully used in molecularly imprinted polymer (MIP)-based analytical column to detect trace BPA in different biologic samples with acceptable accuracy and repeatability.

Differentially expressed genes between female and male adult Anopheles anthropophagus.

The aim of the present study was to identify sex-specific genes in adult Anopheles anthropophagus. As the major malaria vector and Brugia malayi vector in the Asian continent, female Anopheles mosquitoes take blood meals and transmit pathogens through this pathway, while males are nectar feeders. This complex behavior is controlled at several levels, but is probably initiated by the genetic background difference between these two groups. In our study, a subtractive cDNA library for female A. anthropophagus was constructed using the suppression subtractive hybridization (SSH) technique and then 3,074 clones from the female SSH library were analyzed using a microarray-based survey. Genes that were expressed differentially according to sex in A. anthropophagus were screened using real-time polymerase chain reaction and reverse transcription polymerase chain reaction. In our results, we report a series of genes which may be involved in female-specific mosquito behavior, including an inorganic phosphate transporter, a serine protease, the salivary protein GP35-2, and the D7 cluster salivary protein. These findings will provide clues to the nature of insect vectors and open up unprecedented opportunities to develop novel strategies for the control of mosquito-borne diseases.

Near-Complete Genome Sequences of 12 Coxsackievirus Group A Strains from Hand, Foot, and Mouth Disease and Herpangina Cases with Different Clinical Symptoms.

Coxsackievirus group A (CV-A) strains are important pathogens of hand, foot, and mouth disease and herpangina. We report here the near-complete genome sequences of 12 CV-A strains isolated from infants and children with different clinical diseases. The presented data will be very useful for future genome-based epidemiological studies.

Food-borne parasitic zoonoses in China: perspective for control.

Food-borne parasitic zoonoses (FBPZs) cause death and serious diseases in humans and animals worldwide, and are of both public health significance and socioeconomic importance. The FBPZ problem is severe in mainland China, where approximately 150 million people are suffering from FBPZs and more people are at risk. Here, the current status of the FBPZ problem in mainland China is reviewed and strategies and measures for effective control of FBPZs are proposed. Major parasitic zoonoses transmitted through consumption of infected or contaminated meat, fish, plants and/or water will be discussed.

Association of platelet-derived growth factor-D gene polymorphism with ischemic stroke in a Chinese case-control study.

Stroke is a multiple genetic disease. Platelet-derived growth factor-D has been found to be involved in the pathogenesis of atherosclerosis, suggesting possible association between platelet-derived growth factor-D and the development of ischemic stroke. However, little information on the relationship between platelet-derived growth factor-D and stroke is currently available. The aim of this study was to investigate the association between platelet-derived growth factor-D genetic variation and the risk of ischemic stroke in a Chinese population. We conducted a case-control study with 309 ischemic stroke patients and 309 sex and age (<5 years)-matched controls. DNA was extracted from the whole blood of each participant. Platelet-derived growth factor-D C/G polymorphism at position +3166 (rs7950273) was detected by TaqMan SNP genotyping assay. Overall, the combined rates of platelet-derived growth factor- D CG and GG are 51% in patients in contrast with 46% in controls. There were no significant differences in the genotype frequencies of platelet-derived growth factor-D +3166 polymorphisms between the patients and controls with history or family history of hypertension or diabetes (P = 0.770). However, among people without history or family history of hypertension or diabetes, platelet-derived growth factor-D CG/GG is significantly more frequently expressed in patients (60%) than in controls (43%) (odds ratio 1.97; 95% confidence interval 1.19-3.26). This significant association holds after adjustment for age, sex, smoking and alcohol intaking (odds ratio 1.86; 95% confidence interval 1.11-3.10) (P = 0.018). Our study found that the G allele of rs7950273 of the platelet-derived growth factor-D gene is associated with higher risk of ischemic stroke in a Chinese population without history or family history of hypertension or diabetes. Future studies with larger and ethnically diverse populations are needed to further evaluate the platelet-derived growth factor-D polymorphism and stroke association, as well as its pathophysiological mechanisms.

Complete Genome Sequence of an Enterovirus A71 Strain Isolated in 2006 from a Patient in Shenzhen, Southern China, with a Lethal Case of Enterovirus Infection.

The whole-genome sequence of an enterovirus A71 strain (EV71/SHENZHEN001/2006) isolated in 2006 from a patient with a fatal case of enterovirus infection was determined. Phylogenetic analysis based on the complete VP1 gene classified this strain as subgenotype C4a.

Identification and characterization of the causative triatomine bugs of anaphylactic shock in Zhanjiang, China.

BACKGROUND: Two health concerns primarily related to triatomine bugs are transmission of Trypanosoma cruzi through infective feces, and allergic reactions induced by triatomine bites. In the Southwestern United States, reduviid bugs bites commonly cause insect allergy. In South China, four cases of anaphylactic shock have been reported after this bite exposure. To further classify the species of these bugs and confirm the sensitization of the triatomine saliva, we caught triatomine bugs from the region where the bites occurred and performed phylogenetic and immunohistochemical (IHC) analysis. METHODS: Triatomine bugs were collected in Donghai Island of Zhanjiang City in South China. The genomic DNA was extracted from three legs of the bugs. The fragments of mitochondrial 16S rRNA, cytochrome c oxidase subunit I (COI) gene and nuclear ribosomal 18S and 28S rRNA genes were obtained by PCR and sequenced. A phylogenetic tree was constructed based on the sequence of 16S rRNA gene using a maximum likelihood method with MEGA 7.0 software. Trypanosomal specific fragments and vertebrate COI genes were amplified from the fecal DNA to detect the infection of trypanosomes and analyze the blood feeding patterns, respectively. Paraffin-embedded sections were then prepared from adult triatomines and sent for IHC staining. RESULTS: We collected two adult triatomine bugs in Donghai Island. Morphological and molecular analyses indicated that the triatomines were Triatoma rubrofasciata. No fragments of T. cruzi or other trypanosomes were detected from the fecal DNA. Mitochondrial gene segments of Homo sapiens and Mus musculus were successfully amplified. The allergens which induced specific IgE antibodies in human serum were localized in the triatomine saliva by IHC assay. CONCLUSIONS: The two triatomine bugs from Donghai Island were T. rubrofasciata. They had bitten humans and mice. Their saliva should contain the allergens related to the allergic symptoms and even anaphylactic shock of exposed residents. Great consideration should be given to this triatomine bugs due to their considerable distribution and potential threat to public health in South China.