Downstream processing of pullulan from fermentation broth

pullulan, a water soluble extracellular polysaccharide, was produced by downstream fermentation employing the strain Aureobasidium pullulans. To obtain pure biopolymer from the fermentation broth, it is necessary to harvest cells, heat the broth, remove the melanin pigments co-produced during fermentation, concentration, precipitate and dry. Centrifugation of the fermentation broth at 10,000 rpm for 15 min gave cell pellets that were discarded and a green–black supernatant containing melanin pigment was subjected to the heat treatment at 80 °C for 20 min in order to remove the protein in the fermentation broth. The supernatant was demelanized by oxidation with hydrogen peroxide, concentrated under vacuum, precipitated with ethanol and dried at 60 °C for 30 min. This procedure produced high purity pullulan that was comparable in color and texture to the commercial samples.     

Identification of black muntjac (Muntiacus crinifrons) in Tibet, China, by cytochrome b analysis

Full cytochrome b gene sequence of mtDNA was used to identify and analyze four skin samples collected from Tibet, China in this research. By searching for highly similar sequences (megablast) on NCBI, we found all four samples have the highest similarities with the published sequence: AY239042 of the black muntjac (Muntiacus crinifrons). By comparing our sequences to those available on GenBank, all four samples were identified as the black muntjac (Muntiacus crinifrons) by high sequence similarity. We therefore record two new localities for the black muntjac and it is a new distribution in Tibet and hope this study will not only promote more advanced researches on the evolution and phylogeny about this species, but also enhance the conservation work for this species and local biodiversity.     

Microbiological and physicochemical factors affecting <Emphasis Type="Italic">Aspergillus</Emphasis> section <Emphasis Type="Italic">Flavi</Emphasis> incidence in Cavendish banana (<Emphasis Type="Italic">Musa cavendishii</Emphasis>) chips production in Southern Philippines

Microbiological and physicochemical factors affecting the incidence of Aspergillus section Flavi in dried Cavendish banana (Musa cavendishii) chips production in Southern Philippines were examined. The average counts of Aspergillus section Flavi (AFC) in fresh and dried Cavendish bananas from 10 production batches of the Philippine Agro-Industrial Development Cooperative in Davao del Norte, Southern Philippines were 1.2 × 10² and 1.6 × 10² cfu/g, respectively. Isolates from both samples were identified to be Aspergillus flavus based on spore type and conidial structure of isolates. An increasing trend in the AFC of Cavendish bananas was observed during dried banana chips processing. Variability in the AFC between production batches was attributed to differences in aerobic and fungal populations and physicochemical characteristics of the fruits, peel damage of the raw materials, concentration of AFC in the air and food-contact surfaces of the production area, and temperature and relative humidity (RH) conditions of the environment during production and storage. Physicochemical characteristics of Cavendish bananas from the receipt of raw materials up to the first day of drying were within the reported range of values allowing growth and toxin production by aflatoxigenic fungi. Air-borne AFC varied depending on the section of the production area examined. The close proximity of the waste disposal area from the production operation to the preparation, drying and storage areas suggests that cross-contamination, probably air-borne or insect-borne was a likely occurrence. The hands of workers were also identified as AFC sources. Results of this study highlight the need for the development of strategies to control aflatoxigenic fungi and aflatoxin contamination in Philippine dried Cavendish bananas.     

Food resource availability for American black ducks wintering in southern New Jersey

Midwinter waterfowl survey data indicates a long-term decline in the number of wintering American black ducks (Anas rubripes), potentially due to habitat limitations. In order for future estimates of carrying capacity to be determined, it is critical that regional food availability is estimated. We collected pairs of habitat core samples (n = 510) from 5 habitat types in southern New Jersey, USA, during October, January, and April 2006–2008 to estimate resource availability and variability. We collected upper gastrointestinal tracts from hunter-killed birds (n = 45) and late season collections (n = 19) to identify food items and limited our estimates of resource availability to only winter food items; thereby reducing the availability of seed foods found in our core samples by 38% and animal foods by 96%. We did not detect differences in years or sampling period, but did between habitat types. Mudflat habitat had the greatest availability of invertebrate and vertebrate food items and appeared to supply the bulk of energy to black ducks wintering in southern New Jersey. We suggest conservation efforts to be focused on restoring or enhancing mudflat habitat as an integral component of an ecologically functioning salt marsh to increase food availability. © 2011 The Wildlife Society.     

The trade in bear parts from Myanmar: an illustration of the ineffectiveness of enforcement of international wildlife trade regulations

We assessed the effectiveness of national and international wildlife trade regulations in Asia by surveying four wildlife markets in Myanmar for bears and bear parts. Bears are protected in Myanmar and neighbouring countries, and are included in CITES Appendix I, precluding international trade. Three of the four wildlife markets were situated at the border with neighbouring countries (China and Thailand) whereas the fourth, situated in Myanmar’s interior, also catered to international markets. During seven checks (1999–2006) we recorded 1,200 bear parts, representing a minimum of 215 individual bears. Most items were from Asiatic black bears Ursus thibetanus but also sun bear Helarctos malayanus parts were offered for sale. There were significant temporal and spatial differences in what items were offered for sale. Prices were low (USD 4–40 per item) and the total monetary value of the items for sale was USD 6,500–9,500 (not including gall bladders). Carcasses, skulls, canines, paws, claws, whole skins, pieces of skin, gall bladders and derivates, were openly displayed, with vendors being frank about prices, origin, and potential buyers. Only in the interior were prices quoted in the local currency; at the other three markets currencies of the neighbouring countries were used. Legal (international) trade in bears or bear parts from Myanmar is virtually non-existent, and the observed trade in bear parts strongly indicates a serious lack of enforcement effort. International trade in bear parts from Myanmar is significant, and open, and we conclude that the enforcement of wildlife trade regulations, at least when they concerns bear species, have by and large failed.     

河北省冬枣黑斑病病原菌的分离鉴定及生物防治初探

冬枣黑斑病是冬枣重要病害之一,目前多以化学农药进行防治,给自然环境和人类健康带来了严重的威胁。河北省是冬枣种植大省,进行冬枣黑斑病病原菌的分离鉴定对冬枣黑斑病的有效防治具有重要意义。2014年8月至10月从河北省沧州、衡水、邯郸、邢台等地采集冬枣黑斑病病果,采用PDA培养基进行病原菌分离,从病样中共分离出2株分离频率较高的真菌,经过回接和再分离实验筛选出河北省冬枣黑斑病的致病菌株,经形态学和ITS序列分析初步确定该菌为细交链格孢(Alternaria alternata)。以枯草芽胞杆菌J18进行冬枣采后黑斑病的防治,浓度为1×108cfu/m L的菌液对病害的防效为80.67%。     

苏云金芽孢杆菌芽孢萌发相关基因gerM的克隆及功能研究

采用土壤平板法和稀释平板法,自湖北神农架自然保护区的14份土壤样品中分离得到40个暗色丝孢菌分离物,鉴定为16属23种。其中,湖北弯孢Curvularia hubeiensis为一新种,嗜热革节孢Scytalidium thermophilum和禾色串孢Torula graminis为中国新记录种。对新种及中国新记录种作了详细的描述。其余20种为国内已报道种。所有研究菌株的干制培养物标本与活菌种均存放在山东农业大学植物病理学标本室(HSAUP)。     

枯草芽孢杆菌(Bacillus subtilis)Tpb55灌根与烟草根围细菌多样性变化的相关性北大核心CSCD

【目的】利用454高通量测序技术分析生防菌株枯草芽孢杆菌(Bacillus subtilis)Tpb55对烟草根围土壤细菌群落的影响;同时测定施加Tpb55处理对烟草黑胫病的大田防效。【方法】试验设置Tpb55菌剂108 CFU/m L灌根和空白对照2个处理,分别在处理0、10、22 d采集烟草根围土壤,提取土壤细菌总DNA,扩增细菌16S r DNA V1-V3区,对扩增产物进行454高通量测序,使用Qiime软件分析不同处理的细菌群落结构。【结果】对照和处理样品测序后共获得了41207个优质序列,鉴定属于细菌的25个门。所有样品中优势菌群均为放线菌门(Actinobacteria)、变形菌门(Proteobacteria)和酸杆菌门(Acidobacteria)。在病情发展过程中,放线菌门的细菌丰度逐渐下降,变形菌门含量呈上升趋势。施用Tpb55后,酸杆菌门含量明显上升并高于对照。对照中芽孢杆菌科(Bacillaceae)和多样性指示菌草酸杆菌科(Oxalobacteraceae)的细菌丰度均明显下降,Tpb55处理的样品中芽孢杆菌科含量不断上升,草酸杆菌科含量相对稳定。Chao1、ACE和Shannon指数分析表明,Tpb55处理的样品中细菌多样性和丰富度不断提高且高于对照。Tpb55处理后10 d和22 d,OTU序列数据库中与Tpb55 16S r DNA V1-V3区PCR产物高度同源OTU数目分别为31和45。Tpb55处理的烟草黑胫病病情指数(5.29)显著低于对照(38.52)。【结论】施用Tpb55可以提高根围土壤细菌多样性和群落结构稳定性,这可能是其发挥良好生防作用的重要机制之一。     

Extraction of fucoxanthin and polyphenol from <Emphasis Type="Italic">Undaria pinnatifida</Emphasis> using supercritical carbon dioxide with co-solvent

Extraction of brown seaweed (Undaria pinnatifida) oil was carried out by using supercritical carbon dioxide (SCO₂) and ethanol as co-solvent. The flow rate of ethanol was 3.0% (v/v) as compared to that of SCO₂. Experiments were performed in a semi-batch flow apparatus on dried samples at temperatures from 303 to 333 K and pressures from 80 to 300 bar. Fucoxanthin and polyphenol were quantitatively analyzed by using HPLC and UV-spectrometer. The highest yields of fucoxanthin and polyphenol were shown at 200 bar, 323 K and 250 bar, 333 K, respectively. The solubility of fucoxanthin in SCO₂ agreed well with the Chrastil model.     

Pigmentation in Black-boned sheep (Ovis aries): association with polymorphism of the MC1R gene

Variations in vertebrate skin and hair color are due to varied amounts of eumelanin (brown/black) and phaeomelanin (red/yellow) produced by the melanocytes. The melanocortin 1 receptor (MC1R) is a regulator of eumelanin and phaeomelanin production in the melanocytes, and MC1R mutations causing coat color changes are known in many vertebrates. We have sequenced the entire coding region of the MC1R gene in Black-boned, Nanping indigenous and Romney Marsh sheep populations and found two silent mutation sites of A12G and G144C, respectively. PCR-RFLP of G144C showed that frequency of allele G in Black-boned, Nanping indigenous and Romney Marsh sheep was 0.818, 0.894 and 0, respectively. Sheep with GG genotype had significantly higher (P < 0.05) tyrosinase activity than sheep with CC genotype in the all investigated samples. Moreover, there was significant effect of MC1R genotype on coat color, suggesting that MC1R gene could affect coat color but not black traits. There would be merit in further studies using molecular techniques to elucidate the cause of black traits in these Black-boned sheep.     

Gas chromatographic method using nitrogen–phosphorus detection for the measurement of tramadol and its O-desmethyl metabolite in plasma and brain tissue of mice and rats

A method that allows the measurement of plasma and brain levels of the centrally-acting analgesic tramadol and its major metabolite (O-desmethyl tramadol) in mice and rats was developed using gas chromatography equipped with nitrogen–phosphorus detection (GC–NPD). Plasma samples were extracted with methyl tert.-butyl ether (MTBE) and were injected directly into the GC system. Brain tissue homogenates were precipitated with methanol, the resulting supernatant was dried then acidified with hydrochloric acid. The aqueous solution was washed with MTBE twice, alkalinized, and extracted with MTBE. The MTBE layer was dried, reconstituted and injected into the GC system. The GC assay used a DB-1 capillary column with an oven temperature ramp (135 to 179°C at 4°C/min). Dextromethorphan was used as the internal standard. The calibration curves for tramadol and O-desmethyl tramadol in plasma and brain tissue were linear in the range of 10 to 10 000 ng/ml (plasma) and ng/g (brain). Assay accuracy and precision of back calculated standards were within ±15%.     

Species-specific RAPD fingerprints for the closely related Picea mariana and P. rubens

Species-specific molecular markers were designed to assist in the identification of closely related black spruce (Picea mariana [B.S.P.] Mill.) and red spruce (P. rubens Sarg.) in northeastern North America. Trees from six provenances of black spruce and three provenances of red spruce were sampled from outside the sympatric zone. They were first classified using a composite index of five qualitative morphological traits. The species-specific genetic markers were developed using random amplified polymorphic DNAs (RAPD) and a combination of bulk sample and individual tree analyses. Each species bulk sample was constructed from DNAs obtained from 12 trees that were from outside the sympatric zone and showed a morphological composite index specific of each species. A total of 161 primers were screened with the bulk samples. From these, 52 primers showing segregating fingerprints were further screened with the individual trees. Most of the markers observed were shared by the two species, and there was less diversity in P. rubens. A small number of markers were found to be monomorphic or nearly monomorphic and specific to either P. mariana or P. rubens. These markers remained species-specific when F₁ progenies derived from independent intraspecific crosses were screened, and they were subsequently found to co-segregate in hybrids derived from independent interspecific crosses here used as controls.     

Fungal contamination of some imported spices

Seventeen imported raw spice samples obtained from retail outlets were examined for spoilage mould profile. A total of 665 fungal isolates, representing 14 species, were recovered and identified from dried and ground spice samples on several media using standard dilution plate method. Moisture content varied greatly among various samples and were generally high. The most heavily contaminated spice samples examined were observed in red chili and black pepper in order of magnitude of 1580 and 1120 cfu/g, respectively. The most predominant fungal genera encountered were Aspergillus, Penicillium, Rhizopus, Cladosporium and Trichoderma. Yeasts were also frequently recovered, but not identified. Relative occurrence values of taxa disclosed ranged between 36.4% for A. flavus and 0.6% for A. parasiticus and Absidia corymbifera. Samples obtained from gunny bags encounter higher colony counts and contamination frequency than other packing methods. The present magnitude of contamination and spectra of mycobiota approximate those reported for similar spice samples. Although several potentially mycotoxigenic fungi were isolated during the present study, neither the foodstuff nor the fungi were assayed for the presence of these toxins.     

Rapid detection of Pythium porphyrae in commercial samples of dried Porphyra yezoensis sheets by polymerase chain reaction

The fungal parasite Pythium porphyrae is the causative organism of red rot disease in Porphyra cultivation farms. The detection of P. porphyrae from dried Porphyra yezoensis sheets was achieved using the species-specific primers PP-1 (5′-TGTGTTCTGTGCT-CCTCTCG-3′) and PP-2 (5′-CCCAAATTGGTGTTGCCTCC-3′) with the polymerase chain reaction (PCR). The DNA sequence (707 bp) of PCR product was found to be identical to that amplified from ITS rDNA extracted from a type species of P. porphyrae (IFO 30800, The Institute of Fermentation, Osaka, Japan). Quantities of the product amplified varied with the time when samples were harvested after the occurrence of red rot disease in Porphyra farms. This simple, rapid, and inexpensive method should have great applications in furthering quality control and determination of quality ranking in the Porphyra processing industry.     

水淹条件下灰化薹草和虉草活体、枯落物分解过程的比较

为了揭示鄱阳湖湿地活体植物和自然枯落物在水淹条件下分解过程的差异,以鄱阳湖湿地优势植被物种灰化薹草(Carex cinerascens)和虉草(Phalaris arundinacea)为研究对象,采用分解袋法开展室内模拟实验。植物样品设置新鲜和风干两种处理,分别模拟活体植物和自然枯落物在水下的分解过程。研究结果表明:(1)在180 d的淹水实验过程中,两种处理下的灰化薹草和虉草的分解过程都具有阶段性,前期分解速率较快,后期较慢。(2)两种植物枯落物的分解速率与植物C/N比有显著相关性,而活体植物的分解速率与植物C/N比相关性不强。(3)活体植物和自然枯落物的水下分解速率与物种有关,虉草活体比虉草枯落物分解速率快,而薹草活体比薹草枯落物分解速率慢。(4)模拟实验结束时活体植物仍有34%—43%未分解,推测鄱阳湖丰水期退水后大量洲滩植被消失的原因并非是植被在水淹下完全分解,而是一部分植被被水生动物啃噬或被水流冲走。     

Seasonal occurrence and feeding habits of black marlin, <Emphasis Type="Italic">Istiompax indica</Emphasis>, around Yonaguni Island,southwestern Japan

The seasonal occurrence of black marlin, Istiompax indica, around Yonaguni Island, southwestern Japan, was analyzed. Black marlin was abundant from April to July, and the majority of landed fish were female (97%). None of these females was reproductively mature. The condition factor of females increased from April to July. Of 56 black marlin examined, 21 individuals contained 12 prey species; the most important prey species was skipjack tuna, Katsuwonus pelamis, and one neonatal silky shark, Carcharhinus falciformis, was also found. These results indicate that the occurrence of black marlin off Yonaguni Island was associated with feeding and not with spawning.     

甘肃河西走廊土壤中的暗色丝孢菌

从甘肃河西走廊的25份土样中分离出39个暗色丝孢菌分离物,经鉴定分别属于16个属的27个种。其中厚垣链格孢Alternaria chlamydospora、黑小枝顶孢Acremoniella atra和疣小枝顶孢Acremoniella verrucosa为中国新记录种:士栖葡萄穗霉Stachybotrys terrestris为新种。研究过的标本(干制培养物)与活菌种均保存在山东农业大学植物病理学标本室(HSAUP)。     

Survival ofXanthomonas campestris pv.vesictoria in pepper seeds and roots in symptomless and dry leaves in non-host plants and in the soil

Summary A population ofXanthomonas campestris pv.vesicatoria developed as endophytes in the leaves and rhizosphere of apparently symptomless plants grown under mist but not under dry conditions. The pathogen survired for long periods on, and could be isolated from, the surface of infested dried seeds, inoculated sandy loam soil, dried leaves, and the rhizosphere of pepper and of other non-host plants. In addition, small numbers of the pathogen survived for 18 months in a field previously cropped with pepper diseased with bacterial scab. Healthy nursery or mature plants developed symptoms while growing in soil containing infested leaves, either buried or placed on the soil surface.     

Incidence and significance of opportunistic fungi in leukemia patients in India

Twenty-four patients with acute leukemia were investigated for the incidence of opportunistic fungi. Culture isolations of the sputum and urine samples revealed significant levels of Candida in 14 patients; Candida albicans, C. tropicalis and C. pseudotropicalis were the predominant ones isolated. Aspergillus flavus was isolated from blood in two cases and C. albicans and a black yeast from the blood of another two. Serological studies showed fungal antibodies in seven patients; precipitins against Candida were detected in five and Aspergillus in two. Both of the Aspergillus positive cases and two patients who had rising antibodies against Candida died during the course of investigation. In this study 13 of 24 patients developed oral candidiasis.