Molecular typing of sand fly species (Diptera, Psychodidae, Phlebotominae) from areas endemic for Leishmaniasis in Ecuador by PCR-RFLP of 18S ribosomal RNA gene

Surveillance of the distribution of sand fly species is important for prediction of the risk and expansion of Leishmania infection in endemic and surrounding areas. In the present study, a simple and reliable method of typing New World Lutzomyia species circulating in endemic areas in Ecuador was established by using polymerase chain reaction (PCR)-restriction fragment length polymorphism (RFLP) technique. PCR-RFLP of 18S ribosomal RNA (rRNA) genes with the restriction enzyme AfaI and subsequently HinfI successfully identified seven sand fly species in nine endemic areas in Ecuador. Although intraspecific genetic-diversity affecting the RFLP-patterns was detected in a species, the patterns were species specific. The method promises to be a powerful tool for the classification of New World Lutzomyia species.     

Molecular subtyping of feline immunodeficiency virus from cats in Melbourne

OBJECTIVE: To determine the subtypes of feline immunodeficiency virus (FIV) present in the domestic cat population in Melbourne. METHODS: Blood samples were collected from 42 cats that had serum antibodies against FIV. DNA was extracted and subjected to polymerase chain reaction (PCR) to amplify variable regions of the envelope (env) and group specific antigen (gag) genes of FIV. PCR products were directly sequenced or sequenced after cloning when direct sequencing yielded ambiguous results. Phylogenetic analysis was performed and comparisons made with representative sequences of different subtypes. RESULTS: The variable region of the env gene was successfully amplified by PCR from 41 of the 42 cats. All 41 were found to cluster with subtype A env sequences. The variable region of the gag gene was successfully amplified by PCR from all 42 cats. Forty-one were found to cluster with subtype A gag genes and one was found to cluster with subtype B sequences, suggesting that it may be derived from a recombinant env A/gag B virus. CONCLUSIONS: Subtype A is the predominant FIV type in Melbourne, although a subtype A/B recombinant was identified in the population of FIV positive cats. These results of env gene analysis were similar to those in a previous Australian study, suggesting that subtype A predominates in Australia. The results of the gag gene analysis show the importance of analysing multiple areas of the FIV genome when assigning FIV subtypes. Comparison with other major urban centres may provide useful information about the phylogenic diversity of FIV in Australia.     

Effects of resveratrol treatment on mitochondria and subsequent embryonic development of bovine blastocysts cryopreserved by slow freezing

This study evaluated the effects of cryopreservation by slow freezing on the mitochondrial function, DNA integrity, and developmental ability of bovine embryos and examined whether resveratrol treatment of the frozen‐thawed blastocysts improved embryonic viability. In vitro produced bovine embryos were subjected to slow freezing. After thawing, the ATP content and mitochondrial DNA integrity (mtDNA), determined by real‐time PCR targeting short and long mitochondrial sequences, was found to be lower in frozen‐thawed embryos than in fresh embryos, and mtDNA copy number was significantly reduced during the 24‐hr incubation post warming. Furthermore, immunostaining against double‐strand DNA revealed DNA damage in frozen‐thawed embryos. When frozen‐thawed embryos were incubated in the medium containing 0.5 µM resveratrol, SIRT1 expression, and survival rate of the embryos significantly improved compared with the vehicle‐treated embryos. In addition, cell‐free mtDNA content in medium was higher in case of resveratrol‐treated embryos than of vehicle‐treated embryos. In conclusion, slow freezing affects mitochondrial integrity and function in the blastocysts. In the frozen‐thawed embryos, mitochondria were removed during post‐thawing incubation and resveratrol enhanced the process, resulting in improved survivability of the embryos.     

Expression of recombinant feline serum amyloid A (SAA) protein.

Feline serum amyloid A (SAA) cDNA clone was isolated from a hepatic mRNA of a mixed-breed cat. The feline SAA cDNA clone contains 333 nucleotides and deduced amino acid sequence shows 87.4%, 73.9%, and 65.8% homology with dog, human and mouse SAA respectively. Relative to the human and mouse SAA proteins, an additional peptide of eight amino acids is specified in the feline cDNA clone. Recombinant feline SAA (rfSAA) was expressed at high levels using pGEX bacterial expression system. Cleavage from the fusion moiety, and purification using glutathione-sepharose yielded pure soluble form of rfSAA. Antibodies generated against rfSAA were specific for feline SAA and showed no cross-reactivity with human SAA. Furthermore, antibodies against human SAA did not react with feline SAA. These results indicate that antigenicity of feline SAA is totally different from human SAA.     

Selenoneine suppresses melanin synthesis by inhibiting tyrosinase in murine B16 melanoma cells and 3D-cultured human melanocytes

Fisheries Science - The novel organic selenium compound, selenoneine, is found in the blood of tuna and has metal-binding activity. In this report, selenoneine displays tyrosinase inhibitory...     

蒙古高原中部草地土壤冻融过程及土壤含水量分布

利用土壤剖面的温度、湿度观测数据,结合气象资料初步分析了蒙古高原中部典型针茅草原在季节转变过程中(2003～2004年)的土壤冻融过程和土壤含水量分布动态。研究表明,0～150cm深度范围的土壤完全冻结天数为154～160d。冻融日循环主要发生在表层0～5cm。0～30cm土层的土壤含水量变化剧烈,与地温有较好的一致性。0～10cm深度土壤含水量高于其他土层。随着深度的增加,土壤含水量季节波动性变小。冻结过程有利于保持土壤水分,有利于春季草地植被返青。     

Lesions in the Larynx of Wistar RccHan: WIST Rats

Specific regions in the rat larynx exhibit cellular changes in response to inhaled xenobiotics. These regions include the base of the epiglottis, ventral pouch, and medial surfaces of the vocal processes of the arytenoid cartilages. 1 , 2 In order to collect information on the usefulness of trimming techniques, the influence of different vehicles, the impact of different application routes in toxicity studies, and differences between induced vs. spontaneous lesions, the data obtained from a large number of inhalation and non-inhalation studies performed in Wistar RCCHan(TM): Wist rats at Harlan Laboratories Ltd Switzerland, all evaluated or reviewed by the same pathologist, were compiled for a detailed review. The value of different trimming techniques was deemed to be greatest for transverse and sagittolongitudinal section techniques, as compared to horizontolongitudinally section techniques. The comparison of lesions encountered in control rats of inhalation studies treated with different vehicles did not reveal differences in the type, distribution pattern, incidence and/or severity of spontaneous lesions. The types of lesions were also independent of different application routes in non-inhalation studies compared to inhalation studies. The pattern of spontaneous lesions in the rodent larynx was determined by degenerative and inflammatory lesions starting most often in the submucosal glands by desiccated secretion followed by mineralization and local inflammation or were induced by impacted foreign bodies. Squamous metaplasia was recorded in the respiratory epithelium overlaying the ventral gland as a spontaneous lesion in male Wistar rats from inhalation studies with a maxim of 20.0% in an inhalation oncogenicity study. Induced metaplastic changes recorded in the larynx were reversible. Other induced lesions in inhalation studies consisted of submucosal edema, necrosis, inflammation and/or granuloma. Induced lesions in non-inhalation studies were found to be exclusively related to reflux laryngitis or food impaction. It is concluded, that in rodents induced lesions of the larynx differ in type, distribution pattern, severity and incidence from spontaneous lesions.     

Origin identification of dried seaweed product “nori” by PCR–RFLP analysis of Pyropia yezoensis in the internal transcribed spacer ITS-1 region

Nucleotide sequences in internal transcribed spacer (ITS)-1 region derived from dried nori products produced in Japan, China, and the Republic of Korea were compared. Thalli contained in the Japanese products were genetically homogenous, and their nucleotide sequences in ITS-1 were identical to those of the reference strains of Pyropia yezoensis f. narawaensis. In Chinese products, the thalli were related to P. yezoensis strain Minomiasakusa. In contrast, the thalli in the Korean products were genetically heterogeneous, and several different P. yezoensis strains and other Pyropia spp. were used for dried nori products. In some thalli produced in both China and Korea, the DNA sequences of the ITS-1 region were identical with that of Japan, suggesting that the cultivar strains might have been transplanted from Japan to China in recent years. The 432-bp-long nucleotide sequences in the ITS-1 region of thalli derived from Japanese origin were cleaved to two restriction fragments at 154 and 278 bp by cleavage of PCR-amplified products using MspI. Conversely, almost all of the corresponding sequences derived from China and Korea were lacking MspI or other restriction patterns, except for nori products from some areas that cultivate a closely related strain to the Japanese cultivar.     

Origin identification method by multiple trace elemental analysis of intermuscular bones in grilled eel products produced in Japan, China, and Taiwan

Trace elemental composition in intermuscular bones of grilled eel fillets was analyzed to discriminate the geographic origin of eel products derived from Japan, China, and Taiwan. The intermuscular bones were decomposed with nitric acid-hydrogen peroxide, and twelve elements (Li, Ti, V, Mn, Ni, Co, Zn, Rb, Sr, Ba, Pb, and U) were analyzed by inductively coupled plasma mass spectrometry. Trace elemental composition including six elements: V, Co, Sr, Ba, Pb, and U, in intermusucular bones were significantly different between Japanese domestic eels and imported eels from China and Taiwan. The average content of lead in Japanese eels was 1/4 to 1/6 that in eels imported from China and Taiwan. The data of elemental content in intermuscular bones were used for linear discriminant analysis, and two discriminant models were constructed. In the Japan–China discriminant model, the discriminant probabilities between Japanese origin and Chinese origin were 82.5 and 93.3 %, respectively. In the Japan–Taiwan discriminant model, the discriminant probabilities between Japanese origin and Taiwanese origin were 87.5 and 87.0 %, respectively. Therefore, trace elemental analysis is effective for country-of-origin identification of eels processed as grilled eel fillets.     

Identifying the origin of Corbicula clams using trace element analysis

The trace element contents of Corbicula clam shells collected from Japan, Russia, China, and the Republic of Korea were analyzed to determine their geographic origin. The crushed shells were decomposed with nitric acid–hydrogen peroxide, and the concentrations of 14 elements (Li, Mg, V, Mn, Co, As, Rb, Mo, Ba, Ce, Pb, U, Sr, and Ca) were measured by inductively coupled plasma mass spectrometry and inductively coupled plasma optical emission spectrometry. Some of the elements identified in samples displayed a geographic trend. The average content of manganese in Japanese samples was twice that of Russian samples. Conversely, the arsenic content in Japanese samples was approximately half of that in Russian samples. Linear discriminant analysis was applied to the data from Japanese and Russian samples, and a discriminant model was constructed. The discriminant model was used to determine the geographic origin of Corbicula clams produced in Japan, with 89.8 % of those identified as Japanese and 92.2 % of those identified as Russian being classified correctly. Therefore, trace element analysis of the shells of Corbicula clams is a useful technique for the identification of their country of origin.