Risk transboundary transmission areas and driving factors of brucellosis along the borders between China and Mongolia.

OBJECTIVE: Brucellosis is a common and neglected zoonotic infectious disease worldwide caused by Brucella. However, transboundary transmissions among countries, particularly those with high incidences, are seldom investigated. In the present study, by taking China and Mongolia as examples, we aim to identify transboundary transmission risk and driving factors of brucellosis along borders. METHODS: 167 brucellosis outbreak locations along the border between China and Mongolia were collected. Wildlife distribution and cross-border activities were mapped. Maximum entropy approach modeling was conducted to predict the potential risk of prevalence of brucellosis with meteorological factors, geographical environment, economic development, living habits et al. The accuracy of the models was assessed by the area under the receiver operating characteristic (ROC) curve (AUC), Kappa test, and correctly classified instances (CCI). RESULTS: The spatial model performed excellent predictive performance with the predictor variables of soils, pastures, goat density, mean precipitation of the wettest month, temperature seasonality, and population density, which with the contribution and permutation important in 27.2 %, 31.9; 23.3 %, 6.8; 18.0 %, 17.2; 11.2 %, 18.1; 10. 3 %, 15.2; 10.0 %, 10.8. The calculated AUC, SD, Kappa, and CCI are 0.870, 0.001, 0.882, and 0.883, respectively. The distribution map of brucellosis showed high-risk areas along the borders. CONCLUSIONS: Our study identified high-risk areas and the driving effect of brucellosis along the borders between China and Mongolia. Moreover, there is the possibility of cross-border wildlife activities in high-risk areas, which increases the risk of cross-border brucellosis transmission. The funding provides clues for cooperative prevention and control of brucellosis by reducing transboundary transmission.

Driving effect of multiplex factors on human brucellosis in high incidence region, implication for brucellosis based on one health concept.

Brucellosis is a typical zoonosis driven by various risk factors, including environmental ones. The present study aimed to explore the driving effect of environmental factors on human brucellosis in a high incidence rate area, which provides understanding and implications in mitigating disease transmission risk in a multi-system between the human-animal-environment interface for preventing and controlling brucellosis based on the One Health concept. Based on the monthly time series data of human brucellosis and environmental variables, a Seasonal Autoregressive Integrated Moving Average Model with explanatory variables (SARIMAX) was applied to assess the association between environmental indicators and human brucellosis incidence (IHB). The results indicated distinct seasonal fluctuation during the study duration, tending to climb from April to August. Atmospheric pressure, precipitation, relative humidity, mean temperature, sunshine duration, and normalized difference vegetation index significantly drive IHB. Moreover, the well-fitting and predicting capability were performed and assessed in the optimal model was the SARIMAX (0,1,1) (0,1,1)12 model with the normalized difference vegetation index (ß = 0.349, P = 0.036) and mean temperature (ß = 0.133, P = 0.046) lagged in 6 months, and the precipitation lagged in 1 month (ß = -0.090, P = 0.004). Our study suggests the association between environmental risk factors and human brucellosis infection, which can be contributed to mitigating the transmission risk in the environmental drivers in a multi-system interface through comprehensive prevention and intervention strategies based on the One Health concept.

Identification and a full genome analysis of novel camel hepatitis E virus strains obtained from Bactrian camels in Mongolia.

Hepatitis E virus (HEV) infects humans and a wide variety of other mammalian hosts. Recently, HEV strains belonging to genotype 8 (G8) within the Orthohepevirus A species of the Hepeviridae family, were identified in Bactrian camels (Camelus bactrianus) in China. The Bactrian camel (also known as the Mongolian camel) is native to the steppes of Central Asia. However, the HEV strains of Mongolian camels have not been examined. Among 200 serum samples from domestic Bactrian camels raised on 6 farms, in 6 soums in 3 provinces; 71 (35.5 %) were positive for anti-HEV IgG, with prevalence differing by farm (soum) (4.2-75.0 %); and 2 camels (1.0 %) that had been raised in Bogd, Bayankhongor Province, which had the highest seroprevalence among the six studied areas, were positive for HEV RNA. The two HEV strains (BcHEV-MNG140 and BcHEV-MNG146) obtained from the viremic camels in the present study shared 97.7 % nucleotide identity. They were closest to the reported G8 Chinese camel HEV strains but differed from them by 13.9-14.3 % over the entire genome, with a nucleotide difference of 24.0-26.5 % from the reported G1-G7 HEV strains. A phylogenetic tree indicated that the BcHEV-MNG140 and BcHEV-MNG146 strains were located upstream of a clade consisting of the Chinese camel HEV strains and formed a cluster with them, with a bootstrap value of 100 %, suggesting that they may represent a novel subtype within G8. These results indicate a high prevalence of HEV infection in Mongolian camels and suggest that the variability of camel HEV genomes is markedly high.

Putative Novel Serotypes '33' and '35' in Clinically Healthy Small Ruminants in Mongolia Expand the Group of Atypical BTV.

Between 2015 and 2018, we identified the presence of three so-far-unknown Bluetongue virus (BTV) strains (BTV-MNG1/2018, BTV-MNG2/2016, and BTV-MNG3/2016) circulating in clinical healthy sheep and goats in Mongolia. Virus isolation from EDTA blood samples of BTV-MNG1/2018 and BTV-MNG3/2016 was successful on the mammalian cell line BSR using blood collected from surveillance. After experimental inoculation of goats with BTV-MNG2/2016 positive blood as inoculum, we observed viraemia in one goat and with the EDTA blood of the experimental inoculation, the propagation of BTV-MNG2/2016 in cell culture was successful on mammalian cell line BSR as well. However, virus isolation experiments for BTV-MNG2/2016 on KC cells were unsuccessful. Furthermore, we generated the complete coding sequence of all three novel Mongolian strains. For atypical BTV, serotyping via the traditional serum neutralization assay is not trivial. We therefore sorted the 'putative novel atypical serotypes' according to their segment-2 sequence identities and their time point of sampling. Hence, the BTV-MNG1/2018 isolate forms the 'putative novel atypical serotype' 33, the BTV-MNG3/2016 the 'putative novel atypical serotype' 35, whereas the BTV-MNG2/2016 strain belongs to the same putative novel atypical serotype '30' as BTV-XJ1407 from China.

The first isolation and identification of canine parvovirus (CPV) type 2c variants during 2016-2018 genetic surveillance of dogs in Mongolia.

Canine parvovirus type 2 (CPV-2) causes a highly contagious and fatal disease, developing into acute hemorrhagic enteritis and myocarditis, in dogs. CPV-2 has evolved, generating antigenic variants CPV-2a/2b/2c that are globally distributed. However, investigating molecular characterization of CPV-2 among dog populations in Mongolia has been limited. Herein, 42 stool samples were collected from dogs with clinical signs of infection, and conventional PCR assays were employed to detect CPV-2 in 23. Our results indicated that during 2016-2018, the new CPV-2a and 2c subtypes were detected in 34.7% of the samples, and the new CPV-2b subtype was detected in 30.4% of samples. VP2 protein sequence analysis and next-generation sequencing of the complete viral genome confirmed these antigenic types. However, sequence analysis indicated new and unreported mutations, Pro580Thr, and Tyr584His in the CPV-2c subtype. From a PCR-positive sample, CPV-2c was successfully isolated, and we performed an immunofluorescence assay for antigen detection. Additionally, we performed genetic characterization and phylogenetic analysis to investigate genetic diversity among isolates from the region, resulting in high CPV-2 genetic diversity in the Mongolian dog population. Striking similarities were also observed between sequences of the strains isolated from Mongolia and China over a similar time span.

Detection and molecular characterization of equine infectious anemia virus in Mongolian horses.

The genetic characterization and actual prevalence of EIAV in Mongolian horse in the disease endemic region is currently unknown. Here, 11 of 776 horse serum samples from four Mongolian provinces tested positive on agar gel immunodiffusion test. Genomic DNA extracted from all seropositive samples was subjected to nested PCR assay. Among these, three samples tested positive with nested PCR assay and were identified by sequencing analysis based on long termination repeat and tat gene of the virus. Two of the three sequences were identical, with 94.0% identity with the third. These two independent Mongolian EIAV sequences were retained functional motifs, with no dramatic changes but some variability in the U5 region; they were clustered with genotypes from European countries but not with those from China, U.S.A., or Japan.

CD8+ T cells induce thyroid epithelial cell hyperplasia and fibrosis.

CD8(+) T cells can be important effector cells in autoimmune inflammation, generally because they can damage target cells by cytotoxicity. This study shows that activated CD8(+) T cells induce thyroid epithelial cell hyperplasia and proliferation and fibrosis in IFN-γ(-/-) NOD.H-2h4 SCID mice in the absence of CD4(+) T cells. Because CD8(+) T cells induce proliferation rather than cytotoxicity of target cells, these results describe a novel function for CD8(+) T cells in autoimmune disease. In contrast to the ability of purified CD8(+) T cells to induce thyrocyte proliferation, CD4(+) T cells or CD8 T cell-depleted splenocytes induced only mild thyroid lesions in SCID recipients. T cells in both spleens and thyroids highly produce TNF-α. TNF-α promotes proliferation of thyrocytes in vitro, and anti-TNF-α inhibits development of thyroid epithelial cell hyperplasia and proliferation in SCID recipients of IFN-γ(-/-) splenocytes. This suggests that targeting CD8(+) T cells and/or TNF-α may be effective for treating epithelial cell hyperplasia and fibrosis.

Comparison of sensitivity of Th1, Th2, and Th17 cells to Fas-mediated apoptosis.

Following activation through the TCR, CD4+ T cells can differentiate into three major subsets: Th1, Th2, and Th17 cells. IL-17-secreting Th17 cells play an important role in the pathogenesis of several autoimmune diseases and in immune responses to pathogens, but little is known about the regulation of apoptosis in Th17 cells. In this study, the sensitivity of in vitro-polarized Th1, Th2, and Th17 cells to Fas-mediated apoptosis was compared directly by different methods. The order of sensitivity of T cell subsets to Fas-mediated apoptosis is: Th1 > Th17 > Th2. The greater sensitivity of Th17 cells to Fas-mediated apoptosis compared with Th2 cells correlated with their higher expression of FasL and comparable expression of the antiapoptotic molecule FLIP. The decreased sensitivity of Th17 compared with Th1 cells correlated with the higher expression of FLIP by Th17 cells. Transgenic overexpression of FLIP in T cells protected all three subsets from Fas-mediated apoptosis. These findings provide new knowledge for understanding how survival of different subsets of T cells is regulated.

Proteome-based analysis of serologically defined tumor-associated antigens in cutaneous lymphoma.

Information on specificities of serological responses against tumor cells in cutaneous lymphoma patients is relatively restricted. To advance the knowledge of serological immune responses against and to assess the scope of tumor antigenicity of cutaneous lymphoma, 1- and 2-dimensional Western blot analyses with sera from patients were combined with proteomics-based protein identification. Testing sera from 87 cutaneous lymphoma patients by 1-dimensional Western blot analysis, 64 cases of seroreactivity against lymphoma cells were found. The positive responses were relatively weak, restricted to few antigens in each case, and heterogeneous. To identify the antigens, proteins of the mycosis fungoides cell line MyLa and primary tumor cells were separated by 2-dimensional gel electrophoresis, Western-blotted and probed with heterogeneous and autologous patient sera. The antigens were identified from silver-stained replica gels by MALDI-TOF mass spectrometry. 14 different antigens were assigned and identified with this proteome-serological approach. Only one, vimentin, had been reported before, the other 13 are new antigens for cutaneous lymphomas.

Immunosuppressive mechanisms in cancer: consequences for the development of therapeutic vaccines.

Recent investigations revealed strong immunosuppressive mechanisms in tumors that may block anti-tumor T cells and be responsible for failures of immunotherapies. Current attempts to overcome this immunosuppression include blockade of co-inhibitory factors on T cells. Reports from the respective trials indicate that the strategy can improve efficacy of therapeutic vaccination, but at the cost of severe inflammatory and autoimmune reactions. We tried to circumvent tumor-associated immunosuppression by mimotope vaccination to broaden reactive anti-tumor T cell repertoires to include T cells that have not been rendered anergic by the tumor. Initial clinical observations suggest that this strategy bears considerable promise.

Mimotope vaccines for cancer immunotherapy.

Cancer vaccines need to be designed to effectively induce tumor-specific CD8(+) T cells, the key effector cells in immune responses against tumors. These T cells recognize peptides generated from cellular proteins by limited proteolysis, and bound and presented at cell surfaces by MHC class I molecules. Mimotopes, mimetics of T cell epitopes, have been derived from known epitopes by sequence modification, or developed de novo using combinatorial peptide libraries to scan the entire sequence space for peptides that induce the desired T cell responses. Mimotopes of both types have been tested in clinical vaccination trials for treatment of cancer.

Identification of noncanonical melanoma-associated T cell epitopes for cancer immunotherapy.

The identification of tumor-associated T cell epitopes has contributed significantly to the understanding of the interrelationship of tumor and immune system and is instrumental in the development of therapeutic vaccines for the treatment of cancer. Most of the known epitopes have been identified with prediction algorithms that compute the potential capacity of a peptide to bind to HLA class I molecules. However, naturally expressed T cell epitopes need not necessarily be strong HLA binders. To overcome this limitation of the available prediction algorithms we established a strategy for the identification of T cell epitopes that include suboptimal HLA binders. To this end, an artificial neural network was developed that predicts HLA-binding peptides in protein sequences by taking the entire sequence context into consideration rather than computing the sum of the contribution of the individual amino acids. Using this algorithm, we predicted seven HLA A*0201-restricted potential T cell epitopes from known melanoma-associated Ags that do not conform to the canonical anchor motif for this HLA molecule. All seven epitopes were validated as T cell epitopes and three as naturally processed by melanoma tumor cells. T cells for four of the new epitopes were found at elevated frequencies in the peripheral blood of melanoma patients. Modification of the peptides to the canonical sequence motifs led to improved HLA binding and to improved capacity to stimulate T cells.

Tumour-dendritic hybrid cell vaccination for the treatment of patients with malignant melanoma: immunological effects and clinical results.

Hybrid cell vaccines of autologous tumour cells fused with allogenic dendritic cells (DC) combine the tumour's antigenicity with the immune-stimulatory capacity of mature dendritic cells and allogenic MHC class II molecules to activate T cell help and induce tumour-specific cytotoxic T cells. This concept was tested in a clinical trial with melanoma stage III and IV patients. Seventeen patients were evaluated: one experienced complete, one partial response and six stable disease with long survival times. Eleven of fourteen patients, clinical responders and non-responders alike, mounted high-frequency T cell responses to various tumour-associated antigens. Failing clinical responses correlated with loss of antigenicity.