Mixed Rare-Earth Chalcogenide Borate Eu9-xRExMgS2B20O41 (RE = Sm, Gd) Featuring a 3D {[B20O41]22-}∞ Framework Connected by [B6O9(O0.5)6]6- and [B7O13(O0.5)3]8- Clusters.

Rare-earth (RE) chalcogenide borates are very rarely discovered in view of the difficulties in synthesis though they have demonstrated attractive physical performances. Here, the first mixed RE chalcogenide borates Eu5.4Sm3.6MgS2B20O41 (1) and Eu3Gd6MgS2B20O41 (2) are synthesized by combining RE, sulfur, and borate ions into one structure. They crystallize in the centrosymmetric hexagonal space group P63/m, and their 3D honeycomb-like {[B20O41]22-}∞ open frameworks are built by [B6O9(O0.5)6]6- and [B7O13(O0.5)3]8- polyanionic clusters and consolidated by Mg2+ ions; both of which are formed by BO4 tetrahedra and BO3 planar triangles. The coordination modes of RE ions are rare REO6S2 bicapped trigonal prisms and REO8S irregular polyhedra, and their band gaps are determined to be 2.25 and 2.22 eV, respectively. They exhibit antiferromagnetic interactions and distinct photocurrent responses. The corresponding theoretical calculations are also performed. The study of 1 and 2 perhaps stimulates interest in exploring new functional RE chalcogenide borates.

Seed Germination and Seed Bank Dynamics of Eruca sativa (Brassicaceae): A Weed on the Northeastern Edge of Tibetan Plateau.

As a versatile cruciferous species, Eruca sativa is widely cultivated, but in some areas, it has become an invasive weed. There are few studies on its seed dormancy and soil seed bank. This research examined seed dormancy, germination, and dynamics of the soil seed bank of E. sativa, with a view to provide support for its prevention and control. We tested the effects of temperature, light, storage, water, and salinity stress on seed germination and burial depth on seedling emergence of E. sativa. Dynamics of the soil seed bank were determined with a 24 month in situ seed-burial study. Seeds of E. sativa can germinate in a temperature range of 5-35°C; moreover, they exhibited non-deep physiological dormancy (NDPD) at maturity, which can be broken by dry storage or exposure to low temperature in winter. Germination of E. sativa seeds was sensitive to water and salinity stress, and most seeds did not germinate at -0.3 MPa. When buried in soil in the field, seeds exhibited an annual dormancy/non-dormancy cycle and formed at least a short-term persistent soil seed bank. Seeds buried deeper than 5 cm can hardly emerge. Seeds of E. sativa have a wide germination temperature range and exhibited dormancy cycling, which promotes the formation of a persistent soil seed bank and enables it to better adapt to the harsh low-temperature climate of the Qinghai-Tibet Plateau. No-tillage would be a good management strategy for this species.

Comparative short-term outcomes of robotic-assisted surgery for older patients with non-small cell lung cancer: a propensity matched study.

To investigate comparative short-term outcomes of robotic-assisted surgery (RAS) versus video-assisted thoracoscopic surgery (VATS) for older non-small cell lung cancer (NSCLC) patients. Patients ≥ 65 years old with stage I-III NSCLC operated with RAS or VATS from 2016 to 2018 were consecutively included. Propensity score-matched (PSM) method was implemented to balance inter-group biases. Totally 376 participants (224 with VATS and 152 with RAS) were included. After PSM, a cohort (144 with VATS and 107 with RAS) was generated with balanced baseline characteristics. RAS was significantly superior over VATS in the majority of perioperative outcomes, such as operating time (120.8 vs. 165.1 min), conversion rate (0.0% vs. 19.4%), and length of stay (8.6 vs. 10.8 days). RAS versus VATS was significantly associated with comparable rates of postoperative complications (OR 0.642, 95% CI 0.311-1.327), except the rate of pneumonia (OR 0.161, 95% CI 0.048-0.544). RAS leads to analogous postoperative complications and seemingly accelerates the recovery time of older NSCLC patients compared with VATS.

[Construction and Expression of Eukaryotic and Prokaryotic Recombinant Vectors of Pepck and Gp63 Dominant Epitopes of Leishmania infantum].

OBJECTIVE: To construct eukaryotic and prokaryotic recombinant vectors containing Pepck- Gp63 and to achieve protein expression by selecting the dominant epitope genes of Pepck and Gp63 of Leishmania infantum. METHODS: The secondary structure and HLA epitopes of phosphoenolpyruvate carboxylase (PEPCK) were predicted by in silico analysis, and the dominant epitopes were picked out. According to the analysis results of glycoprotein of 63×10 3(GP63) epitopes identified by the same method in our laboratory, the dominant epitope genes of Pepck and Gp63 were used to construct pET32a- Pepck- Gp63 and pVAX1- Pepck- Gp63 by overlapping PCR and enzyme reaction. Then, for protein expression, the prokaryotic vectors were transfected into E.coil while the eukaryotic vectors were transfected into NIH3T3 cells by liposome transfection. RESULTS: There were multiple dominant epitopes in Pepckand there were Pepck-Gp63 sequences in the polyclonal site of expression vector. The expression of Pepck-Gp63 in E.coil appeared in inclusion form and led to 74 kDa band in SDS-PAGE. The immunofluorescence results of NIH3T3 cells transfected by pVAX1- Pepck-Gp63 were positive. CONCLUSION: The recombinant prokaryotic expression plasmids pET32a- Pepck-Gp63 and eukaryotic expression plasmids pVAX1- P epck -Gp63 were successfully constructed, and it was shown that the recombinant plasmids were able to express the corresponding target proteins in E. coli and NIH3T3 cells, respectively, providing a preliminary experimental basis for the subsequent study of immunization strategies.

Smoking history and the efficacy of immune checkpoint inhibitors in patients with advanced non-small cell lung cancer: a systematic review and meta-analysis.

BACKGROUND: Some studies imply a strong correlation between smoking history and the efficacy of immune checkpoint inhibitors (ICIs) in patients with advanced non-small cell lung cancer (NSCLC). Hence, a systematic review and meta-analysis was conducted to comprehensively investigate this correlation. METHODS: Three online databases including PubMed, Embase and Cochrane Library were searched. Abstracts and presentations from European Society of Medical Oncology (ESMO) and American Society of Clinical Oncology (ASCO) were also reviewed. The deadline of search was Nov 9, 2019. Randomized clinical trials (RCT) of ICIs that reported hazard ratio (HR) for overall survival (OS) or progressive-free survival (PFS) by the smoking status of NSCLC patients were eligible for our study. We focused on publications issued in English. A random effects model was implemented in the synthesis, and a two-step interaction test was used to investigate the difference of ICIs efficacy among patients with different smoking histories. RESULTS: Twelve RCTs involving 6,497 NACLC patients [5,569 (85.72%) current/former smokers and 928 (114.28%) never smokers] were eligible for our systematic review and meta-analysis. The pooled HRs [95% confidential interval (CI)] of OS and PFS were 0.74 (0.67, 0.81) and 0.72 (0.59, 0.88) respectively for current/former smokers in the experimental group with ICIs versus those in the control group. The pooled HRs (95% CI) of OS and PFS were 0.81 (0.60, 1.08) and 0.92 (0.55, 1.54) respectively for never smokers in the experimental group with ICIs compared with those in the control group. The difference of ICIs efficacy in terms of OS between current/former and never smokers was insignificant [interaction HR (95% CI), 0.77 (0.69, 0.86), I2=25.4%, P_hetero=0.21]. CONCLUSIONS: The efficacy of ICIs in patients with smoking history is seemingly superior over patients without smoking history, but insignificantly. The difference can be explained by several factors such as insufficient sample size of non-smokers, and confounding factors. We suggest that smoking history cannot be recognized as a predictor of immune therapy in advanced NSCLC.

Seed dormancy release of Halenia elliptica in response to stratification temperature, duration and soil moisture content.

BACKGROUND: Although the effect of cold stratification on seed dormancy release has been extensively studied for many species, knowledge of the role of stratifying temperature, soil moisture content and duration of stratification on seed dormancy release at the population level is limited. Here, we aimed to determine the response of seed dormancy release to these factors in six populations of Halenia elliptica. RESULTS: Seed dormancy release was more responsive to low than high temperatures, and no dormancy break occurred at 8 °C. Seed germination percentage increased first and then remained unchanged as stratifying soil moisture content increased from 0 to 24%. Seed dormancy release of populations from low altitude was more sensitive to increased stratifying temperature and decreased soil moisture content than those from high altitudes. CONCLUSIONS: Temperature and soil moisture changes resulting from global warming could affect seed dormancy release and consequently seedling establishment. Thus, incorporating data on seed dormancy release involving temperature, soil moisture content and stratification duration is beneficial for predicting plant species regeneration, migration and coexistence in a scenario of climate change.

Evaluation of in vitro antileishmanial efficacy of cyclosporin A and its non-immunosuppressive derivative, dihydrocyclosporin A.

BACKGROUND: New therapeutic drugs are urgently needed against visceral leishmaniasis because current drugs, such as pentavalent antimonials and miltefosine, produce severe side effects and development of resistance. Whether cyclosporine A (CsA) and its derivatives can be used as therapeutic drugs for visceral leishmaniasis has been controversial for many years. METHODS: In this study, we evaluated the efficacy of CsA and its derivative, dihydrocyclosporin A (DHCsA-d), against promastigotes and intracellular amastigotes of Leishmania donovani. Sodium stibogluconate (SSG) was used as a positive control. RESULTS: Our results showed that DHCsA-d was able to inhibit the proliferation of L. donovani promastigotes (IC50: 21.24 µM and 12.14 µM at 24 h and 48 h, respectively) and intracellular amastigotes (IC50: 5.23 µM and 4.84 µM at 24 and 48 h, respectively) in vitro, but CsA treatment increased the number of amastigotes in host cells. Both DHCsA-d and CsA caused several alterations in the morphology and ultrastructure of L. donovani, especially in the mitochondria. However, DHCsA-d showed high cytotoxicity towards cells of the mouse macrophage cell line RAW264.7, with CC50 values of 7.98 µM (24 h) and 6.65 µM (48 h). Moreover, DHCsA-d could increase IL-12, TNF-α and IFN-γ production and decrease the levels of IL-10, IL-4, NO and H2O2 in infected macrophages. On the contrary, CsA decreased IL-12, TNF-α, and IFN-γ production and increased the levels of IL-10, IL-4, NO and H2O2 in infected macrophages. The expression of L. donovani cyclophilin A (LdCyPA) in promastigotes and intracellular amastigotes and the expression of cyclophilin A (CyPA) in RAW 264.7 cells were found to be significantly downregulated in the CsA-treated group compared to those in the untreated group. However, no significant changes in LdCyPA and CyPA levels were found after DHCsA-d or SSG treatment. CONCLUSIONS: Our findings initially resolved the dispute regarding the efficacy of CsA and DHCsA-d for visceral leishmaniasis treatment. CsA showed no significant inhibitory effect on intracellular amastigotes. DHCsA-d significantly inhibited promastigotes and intracellular amastigotes, but it was highly cytotoxic. Therefore, CsA and DHCsA-d are not recommended as antileishmanial drugs.

Pathogenic Leishmania spp. detected in lizards from Northwest China using molecular methods.

BACKGROUND: Leishmaniosis, a disease caused by pathogenic Leishmania parasites, remains an unresolved health problem in the New World and the Old World. It is well known that lizards can be infected by a subgenus of Leishmania parasites, i.e. Sauroleishmania, which is non-pathogenic to humans. However, evidence suggests that lizards may also harbor pathogenic Leishmania species including the undetermined Leishmania sp., discovered in our previous work. Leishmania DNA in lizard blood can be detected by using molecular methods, such as the polymerase chain reaction (PCR). RESULTS: Three hundred and sixteen lizards, representing 13 species of four genera, were captured for blood samples collection in Northwest China. Two reliable molecular markers (cytochrome b and heat shock protein 70 genes) were used for detection in the lizard blood samples, to confirm a widespread presence of pathogenic Leishmania parasites and the distribution pattern of Leishmania spp. in lizards from Northwest China. The PCR data indicated positive detection rate for Leishmania in all the tested lizards with an overall prevalence of 57.91% (183/316). Apart from lizard parasites like Leishmania tarentolae and Leishmania sp., several pathogenic Leishmania including L. turanica, L. tropica and L. donovani complex were identified by using phylogenetic analysis. Co-existence of different haplotypes was observed in most Leishmania DNA-positive lizards with an overall rate of 77.6% (142/183). Even mixed infections with different Leishmania species appeared to occur in the lizards with an overall rate of 37.7% (69/183). CONCLUSIONS: Lizards can harbor pathogenic Leishmania spp. Co-existence of different haplotypes or even species of Leishmania indicates mixed infections in natural lizard host. Lizards may contribute to the spread of Leishmania parasites. The pathogenic Leishmania species detected in lizards from Northwest China may be of great eco-epidemiological importance.

An Investigation on the Molecular Characteristics and Intracellular Growth Ability among Environmental and Clinical Isolates of Legionella pneumophila in Sichuan Province, China.

OBJECTIVE: To investigate the molecular characteristics and intracellular growth ability of Legionella pneumophila (L. pneumophila) strains from 1989 to 2016 in Sichuan Province, China. METHODS: Seventy-nine isolates of L. pneumophila were collected from environmental and clinical sources, including cooling towers, hot springs, bath water, fountains, and patients, and identified with 16S rRNA gene analysis and serum agglutination assay. The isolates were then typed by Sequence-Based Typing (SBT), and Genotyping of forty-two LP1 strains were analyzed by means of multiple-locus VNTR analysis with 8 loci (MLVA-8). All strains were further analyzed for two virulence genes: Legionella vir homologue (lvh) and repeats in structural toxin (rtxA). The intracellular growth ability of 33 selected isolates was determined by examining their interaction with J774 cells. RESULTS: All isolates were identified to L. pneumophila including 11 serogroups, among which the main serogroup were LP1, accounting for 54.43%. Thirty-three different sequence types (STs) from five main clonal groups and five singletons were identified, along with 8 different MLVA patterns. Both the lvh and rtxA loci were found in all 79 strains. Thirty isolates showed high intracellular growth ability in J774 cells. CONCLUSION: L. pneumophila is a potential threat to public health, and effective control and prevention strategies are urgently needed.

Direct and Indirect Inhibition Effects of Resveratrol against Toxoplasma gondii Tachyzoites In Vitro.

Toxoplasma gondii is one of the most widespread obligatory parasitic protozoa and infects nearly all warm-blooded animals, leading to toxoplasmosis. The therapeutic drugs currently administered, like the combination of pyrimethamine and sulfadiazine, show high rates of toxic side effects, and drug resistance is encountered in some cases. Resveratrol is a natural plant extract with multiple functions, such as antibacterial, anticancer, and antiparasite activities. In this study, we evaluated the inhibitory effects of resveratrol on tachyzoites of the Toxoplasma gondii RH strain extracellularly and intracellularly. We demonstrate that resveratrol possesses direct antitoxoplasma activity by reducing the population of extracellularly grown tachyzoites, probably by disturbing the redox homeostasis of the parasites. Moreover, resveratrol was also able to release the burden of cellular stress, promote apoptosis, and maintain the autophagic status of macrophages, which turned out to be regulated by intracellular parasites, thereby functioning indirectly in eliminating T. gondii In conclusion, resveratrol has both direct and indirect antitoxoplasma effects against RH tachyzoites and may possess the potential to be further evaluated and employed for toxoplasmosis treatment.

The first successful report of the in vitro life cycle of Chinese Leishmania: the in vitro conversion of Leishmania amastigotes has been raised to 94% by testing 216 culture medium compound.

Chinese Leishmania isolate MHOM/CN/90/SC10H2 (L. H2), which was obtained from the spinal cords of patients from the Sichuan province of China, is an uncharacterized, pathogenic species closely related to Leishmania tarentolae. The in vitro transformation rate of L. H2 promastigotes into amastigotes has not been studied. This study is the first to successfully define the in vitro life cycle of L. H2 by investigating the percent conversion of L.H2 promastigotes to amastigotes in vitro under 216 different culture conditions. The highest proportion of L. H2 amastigotes observed (94%) was significantly higher than that previously reported. After conversion, the axenic amastigotes remained viable as verified by the levels of stage-specific genes (Gp46, A2 and ß-tubulin) detected by RT-PCR. Meanwhile, morphological and protein characterizations of these axenic amastigotes were carried out in order to confirm the successful conversion. Specific antibodies were only able to detect 46 kDa, 52 kDa and 75 kDa proteins in samples isolated from axenic amastigotes. Afterward, these converted axenic amastigotes were transformed into the promastigote form by altering the culture condition. These converted axenic promastigotes still have the ability to infect macrophages, and their morphology changed back to the amastigote form following infection. These findings will assist further investigations into the biological characteristics of the host-parasite relationship and the process of pathogenesis.

Molecular detection, identification and phylogenetic inference of Leishmania spp. in some desert lizards from Northwest China by using internal transcribed spacer 1 (ITS1) sequences.

Leishmaniasis caused by Leishmania is still endemic in Northwest China. It has been thought that reptiles could be a reservoir for mammalian leishmaniasis. However, data are still scarce on natural infection of lizards with Leishmania spp. in China. The present study deals with detection, identification and phylogenetic inference of Leishmania parasites at species and intraspecies levels isolated from six desert lizard species from 10 geographical locations in Northwest China using amplification and sequencing of ITS-rDNA. In total, 83 haplotypes were found among 137 ITS1 sequences obtained from up to 64.6% of all captured lizards. Representative sequences of Leishmania available in GenBank were compiled for comparison with the obtained haplotypes. Tree-based species delimitation was achieved by using Bayesian phylogenitc analyses and maximum parsimony approach. Phylogenetic trees congruently supported that the haplotypes were found to belong to three Leishmania species including L. (sauroleishmania) sp., Leishmania tropica and Leishmania donovani complex. A network approach revealed paraphyletic populations of L. (sauroleishmania) sp. and L. tropica at intraspecies level regarding geographical origin and low host specificity. Chinese L. tropica from lizards showed significant heterogeneity as the obtained haplotypes were distributed in different clusters from other countries. Common ancestry was observed between some sequences of L. tropica from lizards and other sequence types from clinical samples from other countries. This may lend support to the potential reservoir role of lizards for human leishmaniasis. Our results appear to be the first molecular evidence for natural infection of lizards in Northwest China with reptilian Leishmania and mammalian Leishmania species. Desert lizards may be considered as putative reservoir hosts for Leishmania in China. Further studies on persistence of the Leishmania parasites in lizards and sandflies are recommended for the better understanding of their epidemiological involvement.

[Construction and Expression of Vector with mip/flaA Advantages Epitope Genes of Legionella pneumophila].

OBJECTIVE: To generate and express fusion vector with mip/flaA advantages epitope genes of Legionella pneumophila by select mip and flaA advantages epitope genes for future research on Legionella pneumophila protein vaccine. METHODS: Following analysis of secondary structure and surface properties such as: physical and chemical properties, hydropathy, plasticity, antigen index and extracellular domain of Mip and FlaA proteins by bioinformatics methods, the region which active epitope may exist was selected as advantages epitope region. Then, the recombinant plasmid pET-mip, pET-flaA and pET-mip/flaA with advantages epitope genes were constructed by PCR amplification and T4 ligase connection, and induced the expression in E. coli. RESULTS: Many potential antigenic epitopes in Mip and FlaA were identified, and the selected advantages epitope regions were cloned and expressed successfully. Moreover, the mip/flaA two advantages associated epitope fusion proteins were also successfully expressed. CONCLUSION: DNA Star software and Expasy online analysis system can successfully predict antigenic epitopes for Legionella pneumophila Mip and FlaA. And prokaryotic expression vector pET-mip/flaA with advantages epitope genes has been successfully constructed and efficiently expressed.

Regulation of intrinsic apoptosis in cycloheximide-treated macrophages by the Sichuan human strain of Chinese Leishmania isolates.

Leishmania spp. are able to survive and proliferate inside mammals' mononuclear phagocytes, causing Leishmaniasis. Previous studies have noted that the regulation of apoptosis in host cells by these parasites may contribute to their ability to evade the immune system. However, current results remain unclear about whether the parasites can promote or delay the apoptotic process in host cells, because the regulatory effect of Leishmania was assumed to be strain-, species- and even infection time-dependent. The aim of this study was to investigate whether the Sichuan isolates of Chinese Leishmania (SC10H2) can alter the process of intrinsic apoptosis induced by cycloheximide in different types of macrophage cell lines and to determine in which steps of the signaling pathway the parasites were involved. Human THP-1 and mouse RAW264.7 macrophages were infected by SC10H2 promastigotes followed by cycloheximide stimulation to assess the alteration of intrinsic apoptosis in these cells. The results indicated that SC10H2 infection of human THP-1 macrophages could promote the initiation of intrinsic apoptosis, but completely opposite results were found in mouse RAW264.7 macrophages. Nevertheless, the expression of Bcl-2 and the DNA fragmentation rates were not altered by SC10H2 infection in the cell lines used in the experiments. This study suggests that SC10H2 promastigote infection is able to promote and delay the transduction of early apoptotic signals induced by cycloheximide in THP-1 and RAW264.7 macrophages, revealing that the regulation of intrinsic apoptosis in host cells by SC10H2 in vitro occurs in a host cell-dependent manner. The data from this study might play a significant role in further understanding the relationship between Leishmania and different host cells.

Prognostic value of stromal decorin expression in patients with breast cancer: a meta-analysis.

BACKGROUND: Numbers of studies have investigated the biological functions of decorin (DCN) in oncogenesis, tumor progression, angiogenesis and metastasis. Although many of them aim to highlight the prognostic value of stromal DCN expression in breast cancer, some controversial results still exist and a consensus has not been reached until now. Therefore, our meta-analysis aims to determine the prognostic significance of stromal DCN expression in breast cancer patients. METHODS: PubMed, EMBASE, the Web of Science and China National Knowledge Infrastructure (CNKI) databases were searched for full-text literatures met out inclusion criteria. We applied the hazard ratio (HR) with 95% confidence interval (CI) as the appropriate summarized statistics. Q-test and I(2) statistic were employed to estimate the level of heterogeneity across the included studies. Sensitivity analysis was conducted to further identify the possible origins of heterogeneity. The publication bias was detected by Begg's test and Egger's test. RESULTS: There were three English literatures (involving 6 studies) included into our meta-analysis. On the one hand, both the summarized outcomes based on univariate analysis (HR: 0.513; 95% CI: 0.406-0.648; P<0.001) and multivariate analysis (HR: 0.544; 95% CI: 0.388-0.763; P<0.001) indicated that stromal DCN expression could promise the high cancer-specific survival (CSS) of breast cancer patients. On the other hand, both the summarized outcomes based on univariate analysis (HR: 0.504; 95% CI: 0.389-0.651; P<0.001) and multivariate analysis (HR: 0.568; 95% CI: 0.400-0.806; P=0.002) also indicated that stromal DCN expression was positively associated with high disease-free survival (DFS) of breast cancer patients. No significant heterogeneity or publication bias was observed within this meta-analysis. CONCLUSIONS: The present evidences indicate that high stromal DCN expression can significantly predict the good prognosis in patients with breast cancer. The discoveries from our meta-analysis have better be confirmed in the updated review pooling more relevant investigations in the future.

Analysis of kinetoplast cytochrome b gene of 16 Leishmania isolates from different foci of China: different species of Leishmania in China and their phylogenetic inference.

BACKGROUND: Leishmania species belong to the family Trypanosomatidae and cause leishmaniasis, a geographically widespread disease that infects humans and other vertebrates. This disease remains endemic in China. Due to the large geographic area and complex ecological environment, the taxonomic position and phylogenetic relationship of Chinese Leishmania isolates remain uncertain. A recent internal transcribed spacer 1 and cytochrome oxidase II phylogeny of Chinese Leishmania isolates has challenged some aspects of their traditional taxonomy as well as cladistics hypotheses of their phylogeny. The current study was designed to provide further disease background and sequence analysis. METHODS: We systematically analyzed 50 cytochrome b (cyt b) gene sequences of 19 isolates (16 from China, 3 from other countries) sequenced after polymerase chain reaction (PCR) using a special primer for cyt b as well as 31 sequences downloaded from GenBank. After alignment, the data were analyzed using the maximum parsimony, Bayesian and netwok methods. RESULTS: Sequences of six haplotypes representing 10 Chinese isolates formed a monophyletic group and clustered with Leishmania tarentolae. The isolates GS1, GS7, XJ771 of this study from China clustered with other isolates of Leishmania donovani complex. The isolate JS1 was a sister to Leishmania tropica, which represented an L. tropica complex instead of clustering with L. donovani complex or with the other 10 Chinese isolates. The isolates KXG-2 and GS-GER20 formed a monophyletic group with Leishmania turanica from central Asia. In the different phylogenetic trees, all of the Chinese isolates occurred in at least four groups regardless of geographic distribution. CONCLUSIONS: The undescribed Leishmania species of China, which are clearly causative agents of canine leishmaniasis and human visceral leishmaniasis and are related to Sauroleishmania, may have evolved from a common ancestral parasite that came from the Americas and may have split off earlier than the other old world Leishmania. Our results also suggest the following: the isolates GS7, GS1 and XJ771 occur as part of the L. donovani complex; the JS1 isolate is L. tropica; and the isolate GS-GER20 identified as Leishmania gerbilli is close to KXG-2 which is L. turanica.

Phylogenic analysis of Chinese Leishmania isolates based on small subunit ribosomal RNA (SSU rRNA) and 7 spliced leader RNA (7SL RNA).

The leishmaniases are zoonotic diseases caused by protozoan parasites of the genus Leishmania. Leishmaniases are still endemic in China, especially in the west and northwest froniter regions. To revalue the preliminary phylogenetic results of Chinese Leishmania isolates, we amplified partial fragment of small subunit ribosomal RNA (SSU rRNA) and 7 spliced leader RNA (7SL RNA), then tested the phylogenetic relationships among Chinese Leishmania isolates and their relatives by analyzing SSU rRNA gene sequences and 7SL RNA gene sequences. 19 SSU RNA sequences and 9 7SL RNA sequences were obtained in our study, then analyzed with 42 SSU RNA sequences and 32 7SL RNA sequences retrieved from Genbank, respectively. In the Bayesian analysis of the SSU RNA gene, the isolate MHOM/CN/93/GS7 and the isolate IPHL/CN/77/XJ771 are members of Leishmania donovani complex, while the isolate MHOM/CN/84/JS1 clustered with Leishmania tropica. The other 11 Chinese Leishmania isolates (MHOM/CN/90/WC, MCAN/CN/90/SC11, MHOM/CN/80/XJ801, MHOM/CN/85/GS4, MHOM/CN/84/SD1, MCAN/CN/86/SC7, MHOM/CN/54/#3, MHOM/CN/83/GS2, MHOM/CN/90/SC10H2, MHOM/CN/89/GS6 and MHOM/CN/ 89/GS5) form an unclassified group, defined as Leishmania sp., and the most relative species to this group is L. tarentolae. In the Bayesian analysis of the 7SL RNA gene, 9 Chinese Leishmania isolates also formed an unclassified group with L. tarentolae, including canine isolate 10, MHOM/CN/85/GS4, MHOM/CN/84/SD1, MCAN/CN/86/SC7, MHOM/CN/54/#3, MHOM/ CN/83/GS2, MHOM/CN/90/SC10H2, MHOM/CN/89/GS6 and MHOM/CN/89/GS5. We concluded that: (1) Chinese Leishmania isolates are non-monophyly group; (2) an unclassified group may exist in China, and the most relative species to this group is L. tarentolae; (3) MHOM/CN/84/JS1, which was previously assigned as L. donovani, was most genetically related to L. tropica strain MHOM/SU/74/K27.

Prevalence of canine leishmaniasis in Beichuan County, Sichuan, China and phylogenetic evidence for an undescribed Leishmania sp. in China based on 7SL RNA.

BACKGROUND: Leishmaniasis is a vector-borne disease, which is still endemic in the west and northwest area of China. Canines are the major reservoirs of Leishmania, the etiological agent of human visceral leishmaniasis. Phlebotomus chinensis is the main transmission vector of zoonotic visceral leishmaniasis (ZVL). METHODS: In this study, rK39 dip-stick, ELISA and PCR methods were used to investigate the prevalence of canine leishmaniasis (CanL) in Beichuan County, Sichuan Province, China. RESULTS: Among the 86 dogs which were included in the study, 13 dogs were positive using the dip-stick test (15.12%), while 8 dogs were positive using ELISA (9.30%) and 19 dogs were positive for PCR (22.03%). In total, 32 dogs were positive for one or more tests (37.21%). Interestingly, phylogenetic analysis based on the partial 7SL RNA fragment provided evidence that an undescribed Leishmania species, which is clearly a causative agent of CanL and human visceral leishmaniasis, does exist in China. This result is consistent with our previous study. CONCLUSIONS: Our work confirmed that canine leishmaniasis is still prevalent in Beichuan County. Further control is urgently needed, as canine leishmaniasis is of great public health importance. The phylogenetic analysis based on 7SL RNA segment provides evidence for the existence of an undescribed Leishmania sp. in China.

Protective immunity against Legionnaires' disease in an A/J mouse model using a DNA vaccine composed of an outer membrane protein (29 kDa) and the pilE fusion protein.

In order to assess the protective effects of the DNA vaccine (pcDip/pilE) against Legionella pneumophila, the coding sequences of the 2 proteins were cloned into the pET32a(+) and pcDNA3.1(+) vectors. To provide an enhanced immunological response, the proteins were linked together. In this study, the A/J mouse model was used for examine the immunogenicity and protective efficacy of the DNA vaccines of pcDip, pcDpilE and pcDip/pilE. Our results showed that the total IgG titers were higher level increasing after the stimulation of pcDip/pilE than pcDip and pcDpilE immunization. The DNA vaccine (pcDip/pilE) can protect the A/J mouse against a higher dose (2 × 10(7)L. pneumophila cells) of L. pneumonia compared to the other single-DNA vaccine in our study, and the ratio of the survival reached 100% in 10 days after the last DNA vaccine immunization. Our study indicates that these findings provide experimental evidence to support the claim that pcDip/pilE may be an efficient DNA vaccine against Legionella pneumophila.