Cloning, sequencing and mutational analysis of the cytochrome c552 gene (cycB) from Bradyrhizobium japonicum strain 110

We report the cloning and nucleotide sequence analysis of the cytochrome c552 gene (cycB) of Bradyrhizobium japonicum strain 110. The gene was identified with help of an oligonucleotide that was designed on the basis of the amino acid sequence determined for purified cytochrome c552 of B. japonicum strain CC705. The cycB gene product has an N-terminal 23-amino acid signal peptide that is missing in the mature cytochrome c552 protein. A B. japonicum cycB insertion mutant was constructed which had no observable phenotypic defects in denitrification and symbiotic nitrogen fixation. Thus, the function of c552 remains unknown.     

The regulation of mortality and fecundity in Schistosoma mattheei following a single experimental infection in sheep.

The regulation of mortality and fecundity of Schistosoma mattheei in sheep was examined using a series of mathematical models applied to data culled from the literature. Parasite mortality (μ) was found to be an increasing linear function of the magnitude of the initial infection over the ranges of doses examined (200–91, 000 cercariae) where μ = 9.78 × 10⁻³ + 3.476 × 10^−7* infection dose. Parasite fecundity (λ) was found to be inversely related to the duration of the infection. The best fit model for parasite fecundity was one in which fecundity decreased exponentially with time since initial infection, λ = λ₀e^−δ1−r. There was no evidence for density-dependent regulation of fecundity.     

Ancient Human Parasites in Ethnic Chinese Populations

Whilst archaeological evidence for many aspects of life in ancient China is well studied, there has been much less interest in ancient infectious diseases, such as intestinal parasites in past Chinese populations. Here, we bring together evidence from mummies, ancient latrines, and pelvic soil from burials, dating from the Neolithic Period to the Qing Dynasty, in order to better understand the health of the past inhabitants of China and the diseases endemic in the region. Seven species of intestinal parasite have been identified, namely roundworm, whipworm, Chinese liver fluke, oriental schistosome, pinworm, Taenia sp. tapeworm, and the intestinal fluke Fasciolopsis buski. It was found that in the past, roundworm, whipworm, and Chinese liver fluke appear to have been much more common than the other species. While roundworm and whipworm remained common into the late 20th century, Chinese liver fluke seems to have undergone a marked decline in its prevalence over time. The iconic transport route known as the Silk Road has been shown to have acted as a vector for the transmission of ancient diseases, highlighted by the discovery of Chinese liver fluke in a 2,000 year-old relay station in northwest China, 1,500 km outside its endemic range.     

The Substrate-free and -bound Crystal Structures of the Duplicated Taurocyamine Kinase from the Human Parasite Schistosoma mansoni

The taurocyamine kinase from the blood fluke Schistosoma mansoni (SmTK) belongs to the phosphagen kinase (PK) family and catalyzes the reversible Mg²⁺-dependent transfer of a phosphoryl group between ATP and taurocyamine. SmTK is derived from gene duplication, as are all known trematode TKs. Our crystallographic study of SmTK reveals the first atomic structure of both a TK and a PK with a bilobal structure. The two unliganded lobes present a canonical open conformation and interact via their respective C- and N-terminal domains at a helix-mediated interface. This spatial arrangement differs from that observed in true dimeric PKs, in which both N-terminal domains make contact. Our structures of SmTK complexed with taurocyamine or l-arginine compounds explain the mechanism by which an arginine residue of the phosphagen specificity loop is crucial for substrate specificity. An SmTK crystal was soaked with the dead end transition state analog (TSA) components taurocyamine-NO₃²⁻-MgADP. One SmTK monomer was observed with two bound TSAs and an asymmetric conformation, with the first lobe semiclosed and the second closed. However, isothermal titration calorimetry and enzyme kinetics experiments showed that the two lobes function independently. A small angle x-ray scattering model of SmTK-TSA in solution with two closed active sites was generated.     

A Crystallin Fold in the Interleukin-4-inducing Principle of Schistosoma mansoni Eggs (IPSE/α-1) Mediates IgE Binding for Antigen-independent Basophil Activation

The IL-4-inducing principle from Schistosoma mansoni eggs (IPSE/α-1), the major secretory product of eggs from the parasitic worm S. mansoni, efficiently triggers basophils to release the immunomodulatory key cytokine interleukin-4. Activation by IPSE/α-1 requires the presence of IgE on the basophils, but the detailed molecular mechanism underlying activation is unknown. NMR and crystallographic analysis of IPSEΔNLS, a monomeric IPSE/α-1 mutant, revealed that IPSE/α-1 is a new member of the βγ-crystallin superfamily. We demonstrate that this molecule is a general immunoglobulin-binding factor with highest affinity for IgE. NMR binding studies of IPSEΔNLS with the 180-kDa molecule IgE identified a large positively charged binding surface that includes a flexible loop, which is unique to the IPSE/α-1 crystallin fold. Mutational analysis of amino acids in the binding interface showed that residues contributing to IgE binding are important for IgE-dependent activation of basophils. As IPSE/α-1 is unable to cross-link IgE, we propose that this molecule, by taking advantage of its unique IgE-binding crystallin fold, activates basophils by a novel, cross-linking-independent mechanism.     

Schistosoma mansoni-Related Hepatosplenic Morbidity in Adult Population on Kome Island,Sengerema District,Tanzania

Schistosomiasis is one of the important neglected tropical diseases (NTDs) in Tanzania, particularly in Lake Victoria zone. This baseline survey was a part of the main study of integrated control of schistosomiasis and soil-transmitted helminths (STHs) aimed at describing morbidity patterns due to intestinal schistosomiasis among adults living on Kome Island, Sengerema District, Tanzania. Total 388 adults from Kome Islands (about 50 people from each village) aged between 12 and 85 years, were examined by abdominal ultrasound according to the Niamey protocol. Liver image patterns (LIPs) A and B were considered normal, and C-F as distinct periportal fibrosis (PPF). The overall prevalence of PPF was 42.2%; much higher in males than in females (47.0% in male vs 34.4% in females, P=0.007). Abnormal increase of segmental branch wall thickness (SBWT) and dilated portal vein diameter (PVD) were also more common in males than in females. Hepatosplenomegaly was frequently encountered; 68.1% had left liver lobe hepatomegaly and 55.2% had splenomegaly. Schistosoma mansoni-related morbidity is quite high among adults in this community justifying the implementation of integrated control strategies through mass drug administration, improved water supply (pumped wells), and health education that had already started in the study area.     

抗日本血吸虫pVAX1／SjHGPRT·SDISP多价疫苗的构建方法探讨

目的：探讨抗日本血吸虫生殖产卵编码基因多价疫苗pVAX1／SjHGPRT·SDISP的构建方法并从基因与蛋白水平验证该构建方法是否成功。方法：分别以pcDNA3．0／SjHGPRT、pcDNA3．0／SjSDISP为模板,设计引物扩增SjHGPRT、sjSDISP,采用overlap方法扩增全长SjHGPRT·SDISP。将纯化后的产物sjHGPRT·SDISP以及pVaxl质粒采用KpnI和XbaI双酶切,1％低熔点胶回收目的DNA片段以及酶切后Pvaxl载体片段双胶连。连接产物转化至DH5a细胞。挑选阳性克隆采用双酶切以及测序方法从基因水平鉴定是否构建成功。将构建产物免疫观察动物,采用免疫组化方法从蛋白水平鉴定疫苗pVAX1／SjHGPRT·SDISP是否构建成功。结果：酶切方法以及测序结果均显示重组质粒的插入序列分别与目的基因完全一致,昆明小鼠肌肉组织酶免疫组织化学染色显示免疫组小鼠肌细胞中呈现较强的棕黄色颗粒,而阴性对照组肌细胞呈阴性。结论：真核重组质粒pVAX1／SjHG—PRT·SDISP构建成功,且在昆明鼠肌肉细胞内能够获得良好的表达。     

Non-random organization of the Biomphalaria glabrata genome in interphase Bge cells and the spatial repositioning of activated genes in cells co-cultured with Schistosomamansoni

Biomphalaria glabrata is a major intermediate host for the parasitic trematode Schistosoma mansoni, a causative agent of human schistosomiasis. To decipher the molecular basis of this host-parasite interaction, the Bge embryonic cell line provides a unique in vitro model system to assess whether interactions between the snail and parasite affect the cell and genome biology in either organism. The organization of the B. glabrata genome in Bge cells was studied using image analysis through positioning territories of differently sized chromosomes within cell nuclei. The snail chromosome territories are similar in morphology as well as in non-random radial positioning as those found in other derived protostome and deuterostome organisms. Specific monitoring of four gene loci, piwi, BgPrx, actin and ferritin, revealed non-random radial positioning of the genome. This indicates that specific parts of the snail genome reside in reproducible nuclear addresses. To determine whether exposure to parasite is reflected in genome organization, the interphase spatial positioning of genes was assessed after co-culturing Bge cells with either normal or irradiation attenuated miracidia for 30 min to 24 h. The loci of actin and ferritin, genes that are up-regulated in the snail when subjected to infection, were visualized by fluorescence in situ hybridisation (FISH) and their radial nuclear positions i.e. their position in the interphase nucleus with respect to the nuclear edge/envelope, mapped. Interestingly, large scale gene repositioning correlated to temporal kinetics of gene expression levels in Bge cells co-cultured with normal miracidia while irradiated parasites failed to elicit similar gene expression or gene loci repositioning as demonstrated using the ferritin gene. This indicates that normal but not attenuated schistosomes provide stimuli that evoke host responses that are reflected in the host’s nuclear architecture. We believe that this is not only the first time that gene-repositioning studies have been attempted in a mollusc but also demonstrates a parasite influencing the interphase genome organization of its host.     

抗日本血吸虫pVAX1／SjHGPRT-SDISP多价疫苗对昆明小鼠的保护作用及其机制探讨

目的：探讨抗日本血吸虫生殖产卵编码基因多价疫苗pVAX1／sjHGPRToSDISP对昆明小鼠的保护作用及其机制。方法：选取昆明小鼠30只,分别使用pVAX1／sjHGPRToSDISP、pVAX1以及生理盐水,每只小鼠100ug或等量经左腿股四头肌注射。处理2周后,采集动物模型血样检测IgG、IL-2,IL-4,IL-10以及INF-Y表达量。处理4周后,以20±1条尾蚴贴腹感染,感染6周后检测减虫率、减卵率。结果：尾蚴攻击动物后6周,pVAX1／sjHGPRToSDISP免疫组的肝脏减虫率为42．2％,子宫与肝脏减卵率分别为68．04％以及72．96％。与对照组比较,差异有显著性。pVAX1／sjHGPRToSDISP免疫组虫体内IgG、IL-4以及INF．V表达量明显升高．与对照组比较差异有显著性（P〈0．05）。结论：pVAX1／sjHGPRToSDISP多价疫苗具有较好的免疫保护作用,且该类作用的机制与IgG、IL-4以及INF—v的表达升高存在关联。