Analgesic Effects of Tetrodotoxin Combined with Acetylsalicylic Acid on Acetic Acid-induced Abdominal Constriction and Formalin Test in Mice

Aim To study the effects of tetrodotoxin （TTX） combined with acetylsalicylic acid （ASA） on nociceptive stimulus in mice. Methods To assess the antinociceptive effects of TTX, ASA or TTX plus ASA, the acetic acid-induced abdominal constriction test and formalin pain test were used. Results TTX （0.5 - 4.0 μg· kg^-1 ） or ASA （25 - 200 mg· kg^-1 ） im produced a significant inhibition of acetic acid-induced abdominal constriction. The median inhibitory doses （ID508） were 2.1 μg· kg^-1 for TTX（ and 64 mg· kg^-1 for ASA. TTX and ASA also showed a dose-dependent inhibition of the second phase response in the formalin pain model, the ID508, being 2.3μg·kg^-1 and 74.2 mg· kg^-1, respectively. The ihteraction between TTX and ASA was synergistic, as evidenced by the fact that （1） when ASA alone compared with the combination of TTX （0.79 μg · kg^-1 or 0.39μg· kg^-1 ） and ASA, the ID508, of ASA reduced from 64.0 mg· kg^-1 to 5.8 mg· kg^-1 or 12.6 mg· kg^-1, and from 74.2 mg· kg^-1 to 7.4 mg· kg^-1 or 13.0 mg· kg^-1 on tile two models of nociceptive tests, respectively; and that （2） synergism in the analgesic effects was shown by isobiolographic analysis. Conclusion TTX, ASA and the combination of the two drags produce analgesic effects in acetic acid-induced abdominal constriction test and formalin-induced pain test. The interactions between TTX and ASA may be useful in developing novel analgesic agents.

Diversity of filamentous fungi in organic layers of two forests in Zijin Mountain

A study was conducted to evaluate the cultivable filamentous fungal diversity in organic layers (L, F, and H layers) and A1 layer of two main forest types, Pinus massoniana and Liguidambar formasana mixed forest and Quercus variabilis forest, in Zijin Mountain(325?N, 11848?E), Nanjing, China. A total of 67 taxa comprising 56 Deuteromycetes, 3 Zygomycetes, 5 Asco-mycetes and 3 unidentified fungi were recognized from samples from the forest floor of the two forest types. The most abundant group was Deuteromycetes. The dominant genera in both forests were Alternaria sp., Aspergillus sp., Cladosporium sp., Mucor sp., Penicillium sp., Rhizopus sp., Gliocladium sp. and Trichoderma spp. The fungal diversity was higher in the mixed forest than that in Q. variabilis forest. For both forest types, the maximum fungal diversity was found in layer F and there existed significantly different in fungal diversity between layer F and layer L. In the mixed forest, richness of fungi isolated from needle litter (P. massoniana) was lower than that from leaf litter (L. formasana). The richness of fungi from needle litter increased with the in-crease of forest floor depth, but for leaf litter, the fungal diversity decreased with the depth of forest floor. The co-species of fungi from the two forest types, as well as from two kinds of litters in mixed forest, increased with the depth of the forest floor. The succession of fungi along with the process of decomposition was discussed here. The results also showed that litter quality was a critical factor affecting fungal diversity.

Inactivation of infectious bursal disease virus by binary ethylenimine and formalin

In this experiment conducted to study the inactivation dynamics of infectious bursal disease virus （IBDV） by binary ethylenimine （BEI） in comparison with formalin, IBDV was isolated from the bursa of infected chickens and its confirmation was done by agar gel precipitation test. Viral suspensions were subjected to inactivation with BEI and formalin for pre-set time in- tervals. BEI was employed at concentrations of 0.001 and 0.002 mol/L while formalin was used at 0.1% and 0.2%. Sampling was done at 6, 12, 24, 36 and 48 h of incubation and samples were tested for their inactivation status in 9-day-old embryonated eggs and 3-week-old broiler chickens. IBDV was completely inactivated by 0.001 and 0.002 mol/L BEI after 36 h of incubation at 37℃, whereas formalin at 0. 1% and 0.2% concentrations inactivated IBDV in 24 h.

Responses of Earthworms to Organic Matter at Different Stages of Decomposition

This study was conducted to examine the responses of earthworms to soil organic matter and litter at different decomposition stages and their contributions in litter decomposition processes in southern subtropical areas of China. Two plantations were selected as the study sites: Site I was dominated by the exotic endogeic earthworm species Ocnerodrilus occidentalis; Site II was dominated by epigeic species Amynthas corticis. After the fallen litter and earthworms were removed or expelled, four treatments were set up as: reserving the top soil (0–5 cm, equal to H layer) (H), removing the top soil and adding fresh litter (Le), removing the top soil and adding semi-decomposed litter (Li), and a control with no top soil nor any litter (CK). Five randomized blocks that were enclosed with nylon nets on the top were set up in each site, and then the four treatments were arranged randomly in each block. After 2–3 months, earthworms were collected using the formalin method. The results showed that Ocnerodrilus occidentalis preferred Treatment H though it was found in Treatments Le and Li as well; Amynthas corticis preferred Treatment Li though sometimes it also appeared in Treatment H; and Amynthas sp., another epigeic species, was mainly present under Treatment Le and only appeared in Treatment H occasionally. These findings confirmed that earthworm species belonging to different ecological groups had different responses to organic matter at different decomposition stages. The impacts of earthworm communities dominated by O. occidentalis mainly appeared at the later periods of litter decomposition.

Detection of KRAS G12D in colorectal cancer stool by droplet digital PCR

AIMTo assess KRAS G12D mutation detection by droplet digital PCR (ddPCR) in stool-derived DNA from colorectal cancer (CRC) patients.METHODSIn this study, tumor tissue and stool samples were collected from 70 patients with stage I-IV CRC diagnosed by preoperative biopsy. KRAS mutational status was determined by pyrosequencing analysis of DNA obtained from formalin-fixed paraffin-embedded (FFPE) tumor tissues. The KRAS G12D mutation was then analyzed by ddPCR in FFPE tumors and stool-derived DNA from patients with this point mutation. Wild-type (WT) tumors, as determined by pyrosequencing, were included as controls; analysis of FFPE tissue and stool-derived DNA by ddPCR was performed for these patients as well.RESULTSAmong the total 70 patients included, KRAS mutations were detected by pyrosequencing in 32 (45.71%), whereas 38 (54.29%) had WT tumors. The frequency of KRAS mutations was higher in left-sided tumors (11 located in the right colon, 15 in the left, and 6 in the rectum). The predominant point mutation was KRAS G12D (14.29%, n = 10), which was more frequent in early-stage tumors (I-IIA, n = 7). In agreement with pyrosequencing results, the KRAS G12D mutation was detected by ddPCR in FFPE tumor-derived DNA, and only a residual number of mutated copies was found in WT controls. The KRAS G12D mutation was also detected in stool-derived DNA in 80% of all fecal samples from CRC patients with this point mutation.CONCLUSIONddPCR is a reliable and sensitive method to analyze KRAS G12D mutation in stool-derived DNA from CRC patients, especially at early stages. This non-invasive approach is potentially applicable to other relevant biomarkers for CRC management.

Effects of Lugol's iodine solution and formalin on cell volume of three bloom-forming dinoflagellates

Fixatives are traditionally used in marine ecosystem research. The bias introduced by fixatives on.the dimensions of plankton cells may lead to an overestimation or underestimation of the carbon biomass. To determine the impact of traditional fixatives on dinoflagellates during short- and long-term fixation, we analyzed the degree of change in three bloom-forming dinoflagellates （Prorocentrum rnicans, Scrippsiella trochoidea and Nocfiluca scintillans） brought about by Lugol＇s iodine solution （hereafter Lugol＇s） and formalin. The fixation effects were species-specific. P. micans cell volume showed no significant change following long-term preservation, and S. trochoidea swelled by approximately 8.06% in Lugol＇s and by 20.97% in formalin as a percentage of the live cell volume, respectively. N. scintillans shrank significantly in both fixatives. The volume change due to formalin in N. scintillans was not concentration-dependent, whereas the volume shrinkage ofN. scintillans cells fixed with Lugol＇s at a concentration of 2% was nearly six-fold that in cells fixed with Lugol＇s at a concentration of 0.6%-0.8%. To better estimate the volume of N. scintillans fixed in formalin at a concentration of 5%, we suggest that the conversion relationship was as follows： volume of live cell=volume of intact fixed cell/0.61. Apart from size change, damage induced by fixatives on N. scintillans was obvious. Lugol＇s is not a suitable fixative for N. scintillans due to high frequency of broken cells. Accurate carbon biomass estimate ofN. scintillans should be performed on live samples. These findings help to improve the estimate of phytoplankton cell volume and carbon biomass in marine ecosystem.

Acute Toxicities of Potassium Permanganate, Formalin, and Lugol's Iodine Solution to a Marine Ciliate, Pleuronema coronatum (Ciliophora, Scuticociliatida)

Acute toxicities of potassium permanganate, formalin, and Lugol’s iodine solution to a commonly occurred marine ciliate Pleuronema coronatum (Ciliophora, Scuticociliatida) were measured. Linear regression analysis of the results highlighted the close relationships between doses of the medicines and mortalities of the organisms, thus providing a capability to predict toxicity effects from the dose. Toxic effects of the medicines on the ciliates were described in the present paper, and the median lethal concentrations (LC 50 values) were given. Results of measurements indicated that 2 h-LC 50 and 12 h-LC 50 values of formalin on P. coronatum were 59.00×10 -6 and 43.57×10 -6, while those of Lugol’s solutions were 90.13 and 67.84×10 -6 respectively. The tolerance of P. coronatum to formalin is apparently lower than that to Lugol’s iodine solution and potassium permanganate is a suitable medicine to kill ciliates in short time.

Expression of 6 MicroRNAs in Prostate Cancer and Its Significance

OBJECTIVE Numerous microRNAs (miRNAs) are deregulatedin human cancers. The experimental evidence supports thatmiRNAs plays a role in the initiation and progression of humanmalignancies.The present study was undertaken to evaluatethe differential expression of 6 miRNAs as biomarker for earlydetection of prostate cancer, and then to determine whether theexpression profiling of these miRNAs could predict the prognosisof prostate cancer.METHODS The expression profilings of these 6 miRNAs wereinvestigated using the method of locked nucleic acid (LNA)-modified oligonucleotide in situ hybridization (ISH). And thetechnology of tissue microarray (TMA) was employed using theformalin-fixed, paraffin-embedd (FFPE) specimens taken from52 patients with prostate carcinoma (PCa) and 38 patients withbenign prostatic hyperplasia (BPH).RESULTS The rates of positive expression for 6 miRNAs (miR-15b, miR-16, let-7g, miR- 96,miR-182 and miR-183) were 26.92%,15.38%, 15.38%, 67.31%, 61.54% and 71.15% in the specimens ofprostate cancer, and 57.89%, 76.32%, 68.42%, 44.74%, 31.58%,47.37% in the tissues of benign prostatic hyperplasia, respectively.The expressions of all 6 miRNAs between the prostate cancer andbenign prostatic hyperplasia tissues were significantly different(P < 0.05). The positive rate of these 6 miRNAs was significantlyrelated to the Gleason Grading of prostate cancer (P < 0.01). Therewas no significant correlation between the expression of thesemiRNAs and age and the concentration of serum PSA of thepatient (P >0.05). We also found that the expression of miR-15b,miR-96 and miR-182 correlated with clinical stages of tumor (P <0.05). The expression of miR-96 correlated with lobus prostatae oftumor invasion (P < 0.01), but the expressions of the remaining fivemiRNAs were not correlated with that (P >0.05). In addition, theexpression of miR-15b was negatively related to that of miR-96,miR-182 and miR-183, respectively (P < 0.01, r < 0.00).There wasa positive correlation among the expressions of miR-96, miR-182and miR-183 in prostate cancer (P < 0.01, r >0.00). The expressionof miR-16 was positively related to that of miR-let-7g (P < 0.01, r >0.00).CONCLUSION The results suggest that miRNA expressionprofiling could have relevance to the biological and clinicalbehavior of prostate cancer,and they might be importantbiomarkers for early detection and prognostic assessment ofprostate cancer.

Multi-temporal forest cover dynamics in Kashmir Himalayan region for assessing deforestation and forest degradation in the context of REDD+ policy

The role of forests is being actively considered under the agenda of REDD＋ （Reducing Emissions from Deforestation and Forest Degradation plus） aimed at reducing emissions related to changes in forest cover and forest quality. Forests in general have undergone negative changes in the past in the form of deforestation and degradation, while in some countries positive changes are reported in the form of conservation, sustainable management of forests and enhancement of carbon stock. The present study in the Kashmir Himalayan forests is an effort to assess historical forest cover changes that took place from 1980 to 2009 and to predict the same for 2030 on the basis of past trend using geospatial modeling approach. Landsat data （Multispectral Scanner （MSS）, Thematic Mapper （TM） and Enhanced Thematic Mapper （ETM＋）） was used for the years 1980, 199o and （2001, 2009） respectively and change detection analysis between the dates was performed. The maps generated were validated through ground truthing. The study area （3375.62 km^2） from 1980-2009 has uffered deforestation and forest degradation of about 126 km^2 and 239.02 km^2 respectively which can be claimed under negative options of REDD＋, while as the area that experienced no change （1514 km^2） can be claimed under conservation. A small area （23.31 km^2） observed as positive change can be claimed under positive options. The projected estimates of forest cover for 2030 showed increased deforestation and forest degradation on the basis of trend analysis using Cellular Automata （CA） Markov modeling. Despite the fact that country as a whole has registered a net positive change in the past few decades, but there are regions like Kashmir region of western Himalaya which have constantly undergoing deforestation as well as degradation in the past few decades.

Effects of Thinning and Litter Fall Removal on Fine Root Production and Soil Organic Carbon Content in Masson Pine Plantations

Soils play a critical role in the global carbon cycle, and can be major source or sink of CO2 depending upon land use, vegetation type and soil management practices. Fine roots are important component of a forest ecosystem in terms of water and nutrient uptake. In this study the effects of thinning and litter fall removal on fine root production and soil organic carbon content were examined in 20-year-old Masson pine （Pinus resinosa） plantations in Huitong, Hunan Province of China in the growing seasons of 2004 and 2005. The results showed that fine root production was significantly lower in the thinning plots than in the control plots, with a decrease of 58% and 14% in 2004 and 2005 growing seasons, respectively. Litter fall removal significantly increased fine root production by 14% in 2004. Soil temperature （Tsoil） and soil moisture （Msoil） were higher in the thinning plots than those in the controls. Litter fall removal had significant effects on Tsoil and Msoil. Soil organic carbon content was higher in the thinning plots but was lower in the plots with litter fall removal compared with that in the controls. Our results also indicated that annual production of fine roots resulted in small carbon accumulation in the upper layers of the soil, and removal of tree by thinning resulted in a significant increase of carbon storage in Masson pine plantations.

A Taxonomic Study of the Genus Tinocladia (Chordariales, Phaeophyta) in China Coasts I. Two new species, T. eudesmoides sp.nov. and T. zhangii sp.nov.

Two new species of the genus Tinocladia Kylin (1940) T. eudesmoides Ding et Lu sp.nov. and Tinocladia zhangii Ding et Lu sp.nov., are described. T. eudesmoides is mainly characterized by its longitudinally and transversely produced sub-cortical filaments from the medulla, its short uniseriate assimilating filaments, usually composed of 6-10 cells with slightly swollen and narrow ultimate cells, and its unilocular sporangia (60-115)×55-70μm, variable in size. T. zhangii is mainly characterized by its very solid frond, not rip apart by hands, its evident longer primary branches, its short and slender secondary branches, with very evident sub-cortical layer, and filaments divided repeatedly, and its large and small unilocular sporangia, with the latter borne on an evidently swollen pedicel.

Acute Toxicity of Formalin to the Red-tide Dinoflagellate Prorocentrum minimum Schiller(Protozoa,Mastigophora)

The acute toxic levels of formalin to a marine red-tide dinoflagellate Prorocentrum minimum Schiller (Mastigophora, Dinoflagellida) were determined by linear regression analysis. The data obtained revealed that there is a close regression relationship between logarithmic concentrations of formalin and mortality probit scales of the organism when exposure times are 1, 2, 6, 12, 24h. The median lethal concentrations (LC50 values) obtained from correlation analysis for these exposure times (with 95% confidence intervals), were 11.83, 6.76, 4.37, 4.27 and 3.98mgL-1, respectively. A toxicity curve was obtained to connect the exposure time and LC50 value from the correlation between them. The results indicated that P. minimum could be killed or fixed by formalin in the concentration range of 4-12mgL-1 within 1-24h.

A Forest Fire Risk Assessment Using ASTER Images in Peninsular Malaysia

Based on the physical concept of heat energy of pre-ignition,a new fire susceptibility index (FSI) is used to estimate forest fire risk. This physical basis allows calculation of ignition probabilities and comparisons of fire risk across eco-regions. The computation of the index requires inputs of fuel temperature and fuel moisture content (FMC),both of which can be estimated using remote sensing data. While ASTER data for land surface temperatures (LST) was used as proxys for fuel temperatures,fuel moisture content is estimated by regression technique utilizing the ratio NDVI/LST of ASTER data. FSIs are computed in peninsular Malaysia for nine days before the fires of 2004 and 2005 and validated with fire occurrence data. Results show that the FSI increases as the day approaches the fire day. This trend can be observed clearly about four days before the day of fire. It suggests that FSI can be a good estimator of fire risk. The physical basis provides a more meaningful FSI,allows calculation of ignition probabilities and facilitates the development of a future class of fire risk models. FSI can be used to compare fire risk across different eco-regions and time periods. FSI retains the flexibility to be localized to a vegetation type or eco-regions for improved performance.

Distributional Data of the Lizard Fauna （Sauria） of the Maringa-Lopori-Wamba Landscape, Democratic Republic of the Congo

This paper presents the composition of the lizard fauna of Maringa-Lopori-Wamba landscape in the western portion of the central Congo basin in the Democratic Republic of the Congo and its weak similarity with the lizard fauna of Gaboon and Ituri faunal areas. It also fills a gap of knowledge in a region where no previous research on Sauria has been undertaken and signals the presence ofLeptosiaphos hylophilus and Lygodactylus n. sp. The paper mentions the finding of a subspecies of Gastropholis echinata. These data are important since collected for the first time in a spot where no herpetological research on lizards has previously been undertaken.

Use of Residual Material in Biobanking： Solidarity, Common Good, and Informed Consent

Biological residual materials can be obtained from surgical activities or from pathological waste material collected for analysis and stored in formalin. This material can be stored in biobanks with the purpose of future research. Formalin-fixed tissue and also formalin-fixed paraffin embedded tissues are not suitable for all kinds of genetic studies on DNA or RNA, as formalin solution is well known for damaging nucleic acids. Therefore, for the purpose of conducting genetic studies, samples should be frozen in order to maintain a good quality of DNA/RNA over time Biobanks, in which waste samples are frozen, are undoubtedly expensive to maintain; however, it could be useful and important to consider their possible implication in particular research, regarding for example the tumor cells growth process, or when the procurement of samples is difficult. Regarding the relationship between informed consent and tissue collection, storage and research, two choices are possible： irreversible or reversible sample anonymization. These procedures involve different approaches and possible solutions that we will seek to define. Also, an important ethical aspect in regard to the role of biobanks is encouraging sample donation. For donors, seeing human sample being kept rather than discarded and seeing them become useful for research highlight the importance of the human body and improve the attitude towards donation. This process might also facilitate the giving of informed consent more trustfully and willingly.

Investigation Food Diet of Cobitis taenia in Talar River （Mazandaran Province）

Perch sweeper fish with scientific name Cobitis taenia is one of the Cobitidae family. Since this kind of fish lives in clean waters and in the high areas of Talar River, it can be an index on water implution. This survey on the diet of this specimen is done on two stations （Kaselian and Tajoon） and shallow and high areas of Talar River in Mazandaran （Southern part of Caspian sea） and the duration of this survey is from October 2008 till September 2009. We should mention that fishing was just done by electro-shocker with voltage 300 V. And this fish after fishing is fixation in 10% formalin solution. So 410 perch sweeper fish were examined and the average of relatively length of gut （RLG） was 17.35 cm. The index of vacated intestine （VI） was calculated 43.35% for all samples. From intense of diet points of view （GSI）, the most frequency of full intestines was considered in summer and spring. On the other hand, the smallness of relatively length of gut index （RLG 〈 1） shows that perch sweeper fish＇s diet manner is inclined to herbivority. In the examination of containing of digestive track, overally 10 kinds of foodstuff were known. Based on amount of this index, 2.67% Chironomidae is known as main bait in all sampling period.

Species Richness, Diversity and Density of Understory Vegetation along Disturbance Gradients in the Himalayan Conifer Forest

We investigated whether species richness, diversity and density of understory herbaceous plants differed along logging(gap) and grazing(primarily by cattle) disturbance gradients, and sought to identify drivers of richness, diversity and density of understory vegetation of logged sites. A factorial experiment was conducted in the mixed conifer forest of Gidakom in Western Bhutan. Levels of the logging treatment included small(0.15 – 0.24 ha), medium(0.25 – 0.35 ha) and large(0.36 – 1.31 ha) gaps. The grazing treatment included grazed(primarily by cattle) and ungrazed(where herbivores were excluded by a fence) plots nested within each gap. Data were collected from 12 gaps(4 replicates at each level of logging) using the point intercept method. Shannon Weaver Diversity and Margalef's indices were used to estimate species diversity and describe species richness, respectively. Soil samples were analyzed for pH and nutrients. The interaction effect of logging and grazing was significant(p≤0.001) only on species diversity. Relative to ungrazed areas, species diversity was significantly higher(0.01≤p≤0.05) in medium grazed gaps. Under grazed conditions, soil P was negatively correlated with gap size and species diversity. While species diversity was positivelycorrelated(0.01≤p≤0.05) with soil N in grazed plots species richness was positively correlated(0.001≤p≤0.01) with soil N in ungrazed plots. Relative density of Yushania microphylla and Carex nubigena were higher under ungrazed conditions. Our study suggests that the combined effect of cattle grazing and logging results in higher species diversity of understory vegetation in medium and grazed gaps in mixed conifer forests of Bhutan,whereas increase or decrease in relative density of major species is determined primarily by the independent effects of grazing and logging. From management perspective, forest managers must refrain from creating large gaps to avoid loss of nutrients(mainly P and N), which may eventually affect tree regeneration. Managers intending to maintain understory vegetation diversity must consider the combined effects of grazing and logging, ensuring low to moderate grazing pressure.

Optimal Use of Skin Thiersch Graft

The most popular and most widely used mode for short-term storage of grafts was curing them in a standard fridge at a temperature of＋4 ℃. According to the literature, the usability of the same ranges from 10 clays to 2! days. The aim was to determine the usefulness of such a maximum time preserved grafts. This study comprised 50 patients, of which, take parts of skin with the size of 3 cm × 5 cm. It started from the 11th day after the 5 preparation were divided, one of which is a part applied to a suitable surface for accepting the transplant, the second part of the extended conservation 10 more days （to be transplanted） and the third placed in formalin and sent to histopatological analysis. Then, a series of five preparations were made, in which each subsequent extended period of conservation by one day longer, ending their 20th day. Usability over 50% of grafts preserved in NaCl solutions up to 20 days, and then felt. It is possible that a transplant is used with 30 days of preservation, but this is the exception, not the usual state.