Hydrocarbon-Degrading Bacteria and Paraffin from Polluted Seashores 9 Years after the Nakhodka Oil Spill in the Sea of Japan

Pollution of petroleum hydrocarbons, in particular oil spills, has attracted much attention in the past and recent decades. Oil spills influence natural microbial community, and physical and chemical properties of the affected sites. The biodegradation of hydrocarbons by microorganisms is one of the primary ways by which oil spill is eliminated from contaminated sites. One such spill was that of the Russian tanker the Nakhodka that spilled heavy oil into the Sea of Japan on January 2, 1997. The impact of the Nakhodka oil spill resulted in a viscous sticky fluid fouling the shores and affected natural ecosystems. This paper describes the weathering of hydrocarbon-degrading bacteria （genus Pseudomonas） and crystallized organic compounds from the Nakhodka oil spill-polluted seashores after nine years. The Nakhodka oil has hardened and formed crust of crystalline paraffin wax as shown by XRD analysis （0.422, 0.377, and 0.250 nm d-spacing） in association with graphite and calcite after 9 years of bioremediation. Anaerobic reverse side of the oil crust contained numerous coccus typed bacteria associated with halite. The finding of hydrocarbon-degrading bacteria and paraffin wax in the oil crust may have a significant effect on the weathering processes of the Nakhodka oil spill during the 9- year bioremediation.

Cloning and Sequencing Characteristics of rDNA ITS Region of the Pathogen Causing Black Spot Disease on Tumorous Stem Mustard

Taking tumorous stem mustard infected by black spot disease as the research material, the ribosomal 5.8S rDNA and its flanking ITS region were cloned, sequenced and aligned in the study. The results showed that the base sequence of pathogens collected from five different sites was almost identical with that of Alternaria brassicae, and the similarity degree reached 99.68%, without base difference greater than 3 bp. It had obvious differences with the sequences of A. brassicicola and A. japonica, and the base difference was greater than 3 bp with a lot of fragment deletions. It was preliminarily determined that the pathogen cau- sing black spot disease on tumorous stem mustard was A. brassicae.

Expression of mitogen-activated protein kinase double-stranded RNA in cucumber has no apparent effect on the diversity of rhizosphere archaea

The expression of mitogen-activated protein kinase （mapk） double-stranded RNA in cucumber is effective in controlling infestations of the root-knot nematode Meloidogyne incognita. However, little is known about the ecological effects of transgenic plants. Here, we analyzed the diversity of 16S rDNA genes derived from the rhizosphere archaea of transgenic cucumber plants as an indicator of ecological change. A total of 17 and 18 operational taxonomic units were detected in the rhizospheres of non-transgenic cucumber and mapk dsRNA-expressing plants, respectively. No significant difference was observed between the two groups according to Shannon and Simpson indices. In soil samples of the two rhizospheres, the dominant group was Crenarchaeota at the phylum level, with Staphylothermus, Methanococcus, Pyrodictium and Sulfolobus the abundant taxa at the genus level. These results suggest that expressing mapk double-stranded （ds） RNA in cucumber has no apparent effect on the diversity of rhizosphere archaea, and provide powerful evidence for the ecological safety of transgenic cucumber expressing mapk dsRNA.

A New Species of Cat Snake(Serpentes:Colubridae) Morphologically Similar to Boiga cynodon from the Nusa Tenggara Islands,Indonesia

We describe a new cat snake species of the genus Boiga from the Nusa Tenggara Islands, Indonesia. The new species is superficially similar to Boiga cynodon, as it was identified previously. It differs from the latter species by the following combination of characteristics: only reaching half of the size of B. cynodon(up to 1250 mm SVL), higher number of dorsal scales; lower ventral and subcaudal counts and having only a very fine postorbital stripe.

The Use of Cyanobacteria Arthrospira platensis and Cladoceran Daphnia magna as Complementary Protein and Lipid Sources in Transitional Diet for Common Carp (Cyprinus carpio L.) Nursery

This study was conducted to evaluate the use of cyanobacteria Arthrospira platensis and cladoceran Daphnia magna biomass as complementary protein and lipid sources in diet supplementation for common carp (Cyprinus carpio L.) nursery. Three experimental diets containing A. platensis and D. magna meal at different concentrations were compared to the commercial (control) diet. Each experimental diet (ED) was set to contain D. magna and A. platensis meal at a specific combination: 2% and 5%, 2% and 7%, and 4% and 5% for ED1, ED2 and ED3, respectively. The protein and lipid content of the experimental diets ranged from 43.20% to 44.60% dry weight (DW) and 10.64% to 13.42% DW, respectively;while the protein and lipid content of the control diet were 43.00% DW and 6.72% DW, respectively. After 20 days of feeding period, ED3 group obtained the highest final body weight (BW) (58.18 ± 35.24 mg), total biomass (1936 ± 1625 mg), food conversion rate (1.34 ± 0.04), and specific growth rate (12.86 ± 0.03% BW day–1) among all treatment groups (P > 0.05);while ED1 group obtained the highest survival (75.5% ± 7.47%) among all treatment groups (P > 0.05). The total bacterial count and total pathogenic Aeromonas sp. in the culture water of the treatment groups (ranged from 2.00 to 2.65 × 105 CFU mL–1 and 3.50 to 4.12 × 103 CFU mL–1, respectively) were lower compared to the water of the control group fed (3.73 × 105 and 4.70 × 103 CFU mL–1, respectively). No significant differences in physicochemical water quality parameters were observed among treatments (P > 0.05). The current study suggests that the combination of A. platensis and D. magna biomass can be used as complementary protein and lipid sources in diet supplementation for common carp larvae and can result in a comparable fish growth, survival and feed utilization in common carp culture at the nursery phase.

Effects of dietary supplementation with tropical microfungi Mucor circinelloides and Trichoderma harzianum powder on growth performance,bacterial communities,and the biomass nutritional profile of white shrimp grow-out culture(Litopenaeus vannamei)

Microfungi have been explored further as supplement constituents in shrimp culture due to their rich protein,lipid,essential amino acids,fatty acids,and various bioactive compounds.The researchers have evaluated the effects of two microfungi powders(Mucor circinelloides and Trichoderma harzianum)as feed supplements on growth performance,bacterial communities,and the biomass nutritional profile of white shrimp(Litopenaeus vannamei).The results showed that commercial/artificial feed(Feng-Li)with supplementation microfungi powder had a different effect on white shrimp growth performance than control after 62 days of the grow-out period.Supplementation with 1.5%M.circinelloides powder(M-2)could improve the growth performance of white shrimp,especially the survival rate(81.33%),final body weight(5.21 g),and shrimp biomass(211.35 g).In contrast,supplementation with 1.5%T.harzianum powder(T-2)did not improve the growth performance of white shrimp compared to control(p>0.05).Furthermore,it also affected the profile of the bacterial community in culture water as well as in shrimp intestines.Supplementation with both microfungi could reduce the total number of bacteria in culture water at the end of the experiment,where the lowest number was obtained in treatment T-2(3.9×10^(5)CFU/mL),followed by M-2(9.9×10^(5)CFU/mL)and control(4.4×10^(6)CFU/mL).In the culture water,20 isolates of culturable bacteria were found during the trial,with the dominant bacteria in M-2:Bacillus cereus,Bacillus velezensis,Gordonia polyisoprenivorans,T-2:Acinetobacter sp.,Gordonia polyisoprenivorans,Micrococcus luteus.Based on average well colour development(AWCD),the highest bacterial activity was in the M-2(0.297-0.707),followed by T-2(0.226-0.717)and control(0.094-0.694).The highest substrate group utilised in M-2 and T-2 were polymers(0.896 and 1.348)and control:amines(0.100).The highest Shannon diversity index was obtained in the M-2(2.508),followed by control(2.286)and the T-2(2.157).As many as 11 isolates of culturable bacteria were found in the shrimp intestine,with dominant bacteria in M-2:Bacillus velezensis,Micrococcus luteus,Bacillus cereus,T-2:Bacillus velezensis,Micrococcus sp.2,Demequina activiva.The highest bacterial activity was in M-2(2.259),followed by control(2.020)and T-2(0.050).The highest substrate group utilisation was in M-2:polymers(2.564),T-2:carbohydrates(0.094),and control:polymers(2.185).The highest Shannon diversity index was in the M-2(3.234),followed by control(3.230)and the T-2(0.751).In addition,1.5%M.circinelloides and T.harzianum supplementation increased white shrimp muscle protein content up to 79.21%and 80.40%,respectively,compared to control(76.73%).In conclusion,using Mucor circinelloides powder as a feed supplement can improve the growth performance of white shrimp,increase the functional/probiotic bacteria:B.cereus and B.velezensis in cultur and shrimp’s intestine,and increase the protein content in shrimp muscle tissue.However,more in-depth research is needed,especially on anti-nutritional compounds from fungi that affect shrimp growth.

Genome evolution trend of common carp (Cyprinus carpio L.) as revealed by the analysis of microsatellite loci in a gynogentic family

Genome evolution arises from two main ways of duplication and reduction. Fish specific genome duplication （FSGD） may have occurred before the radiation of the teleosts. Common carp （Cyprinus carpio L.） has been considered to be a tetraploid species, because of its chromosome numbers （2n=100） and its high DNA content. Using 69 microsatellite primer pairs, the variations were studied to better understand the genome evolution （genome duplication and diploidization） of common carp from a gynogenetic family. About 48% of primer pairs were estimated to amplify duplicates based on the number of PCR amplification per individual. Segregation patterns in the family suggested a partially duplicated genome structure and disomic inheritance. This indicates that the common carp is tetraploid and polyploidy occurred by allotetraploidy. Two primer pairs （HLJ021 and HLJ332） were estimated to amplify reduction based on the number of PCR amplification per individual. One allele in HLJ002 locus and HLJ332 locus was clearly lost in the gynogenetic family and the same as in six wild populations. Segregation patterns in the family suggested a partially diplodization genome structure. A hypothesis transition （dynamic） and equilibrium （static） were proposed to explain the common carp genome evolution between genome duplication and diploidization.

A Resource for Inactivation of MicroRNAs Using Short Tandem Target Mimic Technology in Model and Crop Plants

microRNAs (miRNAs)are endogenous small non-coding RNAs that bind to mRNAs and target them for cleavage and/or translational repression,leading to gene silencing.We previously developed short tandem target mimic (STTM)technology to deactivate endogenous miRNAs in Arabidopsis.Here,we created hundreds of STTMs that target both conserved and species-specific miRNAs in Arabidopsis,tomato,rice,and maize,providing a resource for the functional interrogation of miRNAs.We not only revealed the functions of several miRNAs in plant development,but also demonstrated that tissue-specific inactivation of a few miRNAs in rice leads to an increase in grain size without adversely affecting overall plant growth and development.RNA-seq and small RNAseq analyses of STTM156/157 and STTM165/166 transgenic plants revealed the roles of these miRNAs in plant hormone biosynthesis and activation,secondary metabolism,and ion-channel activity-associated electrophysiology,demonstrating that STTM technology is an effective approach for studying miRNA functions.To facilitate the study and application of STTM transgenic plants and to provide a useful platform for storing and sharing of information about miRNA-regulated gene networks,we have established an online Genome Browser (https://blossom.ffr.mtu.edu/designindex2.php) to display the transcriptomic and miRNAomic changes in STTMinduced miRNA knockdown plants.

Toxicity of Sodium Fluoride to Caenorhabditis elegans

Objective To investigate the toxic effect of sodium fluoride（NaF） on the nematode Caenorhabditis elegans（C.elegans）.Methods Adult C.elegans were exposed to different concentrations of NaF（0.038 mmol/L,0.38 mmol/L,and 3.8 mmol/L） for 24 h.To assess the physiological effects of NaF,the brood size,life span,head thrashes,and body bend frequency were examined.Reactive oxygen species（ROS） and cell apoptosis were detected as parameters of biochemical response.The gene expressions were determined by real‐time polymerase chain reaction（PCR） to assess the molecular‐level response.Results At the physiological level,the brood size of C.elegans exposed to 0.038 mmol/L,0.38 mmol/L,and 3.8 mmol/L concentrations of NaF were reduced by 6%,26%,and 28% respectively in comparison with the control group.The maximum life spans of C.elegans exposed to 0.038 mmol/L,0.38 mmol/L,and 3.8 mmol/L concentrations of NaF were reduced by 3 days and 5 days,respectively.Head thrashes and body bend frequency both decreased with increasing concentrations of NaF.At the biochemical level,the production of ROS and the incidence of cell apoptosis increased with increasing concentrations of NaF（P0.05）.At the molecular level,different concentrations of NaF exposure raised the expression of stress‐related genes,such as hsp16.1,sod‐3,ctl‐2,dhs‐28,gst‐1,and cep‐1.Conclusion NaF exposure could induce multiple biological toxicities to C.elegans in a concentration‐dependent manner.These toxicities may be relevant to the oxidative stress induced by increased ROS production and accumulation in C.elegans.

Identification of Perkinsus-like parasite in Manila clam, Ruditapes philippinarum using DNA molecular marker at ITS region

Genomic DNA was extracted from hypnospores of Perkinsus-like parasite of Manila clam Ruditapes philippinarum collected at the fishing grounds in Huanghai Sea coast Shicheng Island and East China Sea coast Ningbo, China. The internal transcribed spacer（ITS） in rDNA was PCR-amplified, cloned, sequenced, and compared with that of five Perkinsus species in GenBank. The fragment amplified from DNA of parasite of either Shicheng Island or Ningbo contained 649 bp, including partial ssrRNA（51 bp） and ITS（＋5.8 S） （598 bp） regions. The ITS（＋5.SS） sequences of Perkinsus-like parasite of both Shicheng Island and Ningbo were all 99% identical to those ofPerkinsis atlanticus, and were not more than 95% identical to those of other four Perkinsus species including P. marinus, P. andrewsi, P. qugwadi and P. medierraneus.The ITS （＋5.8S） sequence of Perkinsus-like parasite of Shicheng Island was 99% identical to that of Ningbo. These facts about nucleotide sequences suggested that the Perkinsus-like parasite in Manila clam, Ruditapes philippinarum collected from either the Huanghai Sea coast or the East China Sea coast was P. atlanticus, and might reflect P. atlanticus strains of distinct geographic distribution.

Toxicity of Naphthalene on Caenorhabditis elegans

Naphthalene is a common environmental contaminant substance. The toxic effects of naphthalene on Caenorhabditis elegans were investigated at the molecular, biochemical and physiological levels. To assess the molecular-level effect, stress-related gene expression was investigated such as those of hsp-16.1, sod-3, ctl-2, cep-1, cyp35a2, ugt-44, gst-1 and dhs-28. Cell apoptosis was assessed at the biochemical level. Life span, locomotion behaviors and brood size were investigated at the physiological level. The results indicate that naphthalene exposure could not only induce the expression of stress-related genes such as hsp16.1, sod-3, ctl-2 and cep-1 but also reduce the life span of Caenorhabditis elegans. At the same time, naphthalene exposure could result in cell apoptosis and interfere in the locomotion behaviors of Caenorhabditis elegans. These data suggest that naphthalene has multiple toxicity on Caenorhabditis elegans.

Reduction of 4-nitrophenol catalyzed by nitroreductase

nitroreductase 面对 NADH 催化的 4-nitrophenol 的减小在这篇论文被调查。4-Amino-phenol 和 4-hydroxylamino-phenol 在减少的产品被发现。在反应周期和减少的比率之间的关系被学习。4-nitrophenol 的类似的减少比率在氧气、厌氧的条件下面被获得。结果显示氧感觉迟钝的反应在 4-nitrophenol 的减小被继续， nitroreductase 是氧感觉迟钝的酶。4-nitrophenol 的减少的产品被 HPLC 和 MS 决定。

New Cell Viability Measurement of SH-SY5Y Cells in the PD Research-Alamarblue^(TM)

The traditional MTT assay requires destructive analyses and is not convenient in continuous monitory of cell viability. However, a new cell model was developed in this research, by using the oxidation-reduction (redox) indicator alamarblue? instead of the MTT assay. The alamarblue?does no harm to cells and provide a much more safe and convenient methods of measurement. Firstly, cell apoptosis was induced by different concentration of 6-OHDA, then the cell viability was tested by the alamarblue?at a serial of time points. Finally, the optimism cells density, 6-OHDA concentration and testing time point were gained to set a wonderful SH-SY5Y cell model in our research. And when it is applied in the study of neuroprotection effects of NAC, GSH and Catalase, the new model reveals undeniable advantages.

Performance evaluation of IoT-based service system for monitoring nutritional deficiencies in plants

This study aimed to develop and evaluate the performance of a service system platform based on the Internet of Things(IoT)for monitoring nutritional deficiencies in plants and providing fertilizer recommendations.There are two distinct differences between this work and previous ones;namely,this service system platform has been developed based on IoT using a system engineering approach and its performance has been evaluated using dependability.We have successfully developed and integrated a service system platform and chlorophyll meter that is based on IoT.We have also successfully tested the performance of the service system platform using the JMeter software.The dependability value measured from the five tested variables(reliability,availability,integrity,maintainability,and safety)showed a value of 0.97 which represents a very good level of system confidence in not failing to deliver services to users under normal operational conditions.From a future perspective,this platform can be used as an alternative service to monitor nutrient deficiencies in plants and provide fertilization recommendations to increase yields,reduce fertilizer costs,and prevent the use of excessive fertilizers,which can cause environmental pollution.

Microbiological synthesis of nanophase PbS by Desulfotomaculum sp.

The uniform nanoparticles PbS with diameters about 13 nm could be synthesized by Desulfotomaculum sp. under mild condition. The effects of the processing vari-ables such as pH and temperature were investigated. The obtained products were in detailed by means of transmission electron microscopy(TEM) and X-ray diffraction(XRD) ,respectively. The results revealed that the PbS crystallites were identical in structure,shape and size under different temperatures while their morphology changed from rod to spheroidal with pH increasing. In the biological synthetic process for PbS nanoparticles,Desulfotomaculum sp. can use sulfate as a terminal electron acceptor to produce sulfide which acts as the source of sulfur for the formation of PbS nanoparticles.

Enhancement of Essential Oil Yield from Melaleuca Leucadendra L.Leaves by Lignocellulose Degradation Pre-Treatment Using Filamentous Fungi

The essential oil from Melaleuca leucadendra L.leaves has been widely used as a perfume and traditional remedy,cosmetics and pharmaceutical products ingredient since many years ago.The common technology to recover the oil is hydro-distillation and steam-distillation.However,all oil can not be fully extracted from the leaves by this method due to the recalcitrant structure of leaves that hindrance the access of the solvent.Adding a submerged fermentation as a pretreatment step prior to the extraction process helped to loosen the lignocellulose structure and enhance oil release in the extraction process.In this study,the raw materials were collected from the natural forest in Buru Island,Maluku,Indonesia.The biological agents applied in these processes were Phanerochaete chrysosporium ITBCC136 and Trichoderma viride ITBCC143.The oil extraction process was conducted by method of steam-distillation,the oil was analysed using gas chromatography-mass spectroscopy(GC-MS),and the lignocellulose content in the biomass was measured by the fractionation method.The treatment using T.viride provided the highest increase in yield percentage up to 3.47%as compared with control of 1.45%,with the lowest percentages of the remained cellulose,while the fermentation with the presence of P.chrysosporium did not affect the oil yield even the lignin content was decrease as much as 21%.The percentages of 1,8-cineole in the oil were almost unchanged,which was about 20%of the oil.