Exploration of Extracellular Vesicle Long Non-Coding RNAs in Serum of Patients with Cervical Cancer.

INTRODUCTION: Cervical cancer (CC) is the fourth most common cancer type and a leading cause of cancer-related deaths in women worldwide. Its underlying molecular mechanisms are unclear. Cancer cell-derived extracellular vesicles (EVs) are involved in cancer development and progression by delivering regulatory factors, including microRNAs and long non-coding RNAs (lncRNAs). METHODS: Here, we identified the EV lncRNA expression profiles associated with different developmental stages of CC using next-generation sequencing. EVs from the serum of patients with stages I-III CC and healthy donors were characterized using EV marker immunoblotting and transmission electron microscopy. RESULTS: The EV concentration increases with progression of the disease. Most particles had a 100-250-nm diameter, and their sizes were similar in all groups. We identified many lncRNAs that were uniquely and differentially expressed (DE) in patients with different stages of CC. The pathway analysis results indicated that the upregulated DE EV lncRNAs abundant in stages I and II were associated with cell proliferation and inflammation and cancer progression pathways, respectively. LINC00941, LINC01910, LINC02454, and DSG2-AS1 were highly expressed, suggesting poor overall survival of CC patients. Interestingly, DSG2-AS1 was associated with the human papillomavirus infection pathway through AKT3, DLG1, and COL6A2 genes. CONCLUSION: This is the first study that reports the levels of EVs and their lncRNA contents change during cancer development, demonstrating the existence of a unique vesicle-mediated cell-to-cell communication network underlying cancer progression.

Transcriptome profiling of banana shrimp (Fenneropenaeus merguiensis) ovaries and testes: Insights into FoxL2.

The banana shrimp is found in the Pacific and Indian Oceans. Female shrimp are preferred for consumption because they are larger than males. Understanding the mechanism of sex differentiation is important for developing techniques to increase the number of female shrimp for economic benefits. This study investigates the reproductive development of F. merguiensis using transcriptome analysis. Sxl2, dsx, AGH, FEM-1, and Nrg-X2 were classified as essential genes for testes development during the juvenile stage. Several genes were required for both juvenile and adult male development. Additionally, the expression of several genes was shown to be required for juvenile and adult ovarian development, including SOP1, SOP2, Ptgr1, EST, Vgr, Vmol1, and TR-beta A. Interestingly, high levels of FoxL2 expression were observed in the testes, in contrast to previous studies in humans and other mammals. The binding of FoxL2 to the Vtg promoter was demonstrated in silico with the highest relative binding score (RS = 0.89) using the JASPAR program. Knock-down of the FoxL2 gene with dsRNA significantly suppressed FoxL2 at 2, 4, and 6 d. As a result, Vtg expression increased when compared with the control at 2, 4, and 6 d, indicating that FoxL2 plays an important role in Vtg expression in the ovary. Our findings highlight the role of FoxL2 in banana shrimp reproduction and provide valuable information on the genes associated with the F. merguiensis reproductive system.

Differentially expressed genes in the testes from early to mature development of banana shrimp (Fenneropenaeus merguiensis).

Banana shrimp (Fenneropenaeus merguiensis) is an economically important species in Thailand owing to the high value of globally exported frozen brine shrimps. However, the regulatory mechanisms governing spermatogenesis and testicular development in this species are poorly understood. High-throughput RNA sequencing was used to investigate the mechanisms and regulated genes involved in testis development using transcriptome profiling of juvenile and adult banana shrimp testes. Differentially expressed genes (DEGs) in these two libraries were identified and quantified to confirm gene expression. DEGs were found in 7,347 genes, with 4,465 upregulated and 2,882 downregulated. Some of these genes were designated as candidate genes, and six specific DEGs, including PRM1, SPATA20, Sry, SSRF, Sxl, and Tra-2c, were selected to confirm the reliability of the RNA-seq data using qPCR. Moreover, six non-DEGs were chosen based on testis-specific and regulatory genes that support a specific function in spermatogenesis and testis development in this species, including Dsx, Gfra2, IAG, Sox9, Sox13, and Sox14A. Furthermore, Sry, Sox14A, Sox14B and SPATA20 were identified in early stages (nauplius-postlarvae) of shrimp development to provide more information involving testes formation and development. The transcript data from this study could differentiate a group of genes required at the early and late stages of testis development and both sets of testis development. Therefore, this information would help in manipulating each stage of testicular development.

Biocompatibility and mineralization activity of modified glass ionomer cement in human dental pulp stem cells.

Background/purpose: Fortilin is a multi-functional protein involved in several cellular processes. It has been shown promising potential to be a bioactive molecule that can be incorporated in the dental materials. This study aimed to compare the biocompatibility and mineralization activities of modified glass ionomer cement (Bio-GIC) and Biodentine by direct and indirect method on human dental pulp stem cells (hDPSCs). Materials and methods: Conventional glass ionomer cement (GIC), Bio-GIC (GIC supplemented with chitosan, tricalcium phosphate, and recombinant fortilin from Fenneropenaeus merguiensis), and Biodentine were examined in this study. Recombinant fortilin was purified and tested for its cytotoxicity by 3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyltetrazolium bromide (MTT) assay. Human DPSCs were treated with different material eluate for particular time intervals. At given time points, viability of hDPSCs was examined using MTT assay and calcium deposition was assessed by Alizarin red staining assay. Comparisons of the data among groups were analyzed by analysis of variance and Tukey's multiple comparisons. Results: All test materials demonstrated no cytotoxicity. In addition, Bio-GIC promoted cell proliferation at 72 h. For direct and indirect method, cells treated with Bio-GIC demonstrated significantly higher calcium deposition than other groups (P < 0.05). Conclusion: Bio-GIC and Biodentine are not cytotoxic to hDPSCs. Bio-GIC demonstrates enhanced calcium deposition comparable to Biodentine. Bio-GIC may be further developed as a bioactive material for dentin regeneration.

Biological Activities of Glass Ionomer Cement Supplemented with Fortilin on Human Dental Pulp Stem Cells.

This study aimed to determine the most suitable recombinant fortilin and evaluate the biological activities of glass ionomer cement (GIC) incorporated with fortilin on human dental pulp stem cells (hDPSCs). Full-length and three fragments of Penaeus merguiensis fortilin were cloned and examined for their proliferative and cytoprotective effects on hDPSCs by MTT (3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide) assay. Human DPSCs were cultured with GIC supplemented with fortilin, tricalcium phosphate, or a combination of tricalcium phosphate and fortilin, designated as GIC + FL, GIC + TCP, and GIC + TCP + FL, respectively (n = 4 for each group). At given time points, hDPSCs were harvested and analyzed by MTT, quantitative reverse transcription polymerase chain reaction, alkaline phosphatase activity, and Alizarin Red assays. The full-length fortilin promoted cell proliferation and significantly increased cell survival. This protein was subsequently added into the GIC along with tricalcium phosphate to investigate the biological activities. All experimental groups showed reduced cell viability after treatment with modified GICs on days 1 and 3. The GIC + TCP + FL group significantly promoted odontoblastic differentiation at particular time points. In addition, alkaline phosphatase activity and calcium phosphate deposit were markedly increased in the GIC + TCP + FL group. Among all experimental groups, the GIC incorporated with fortilin and tricalcium phosphate demonstrated the best results on odontogenic differentiation and mineral deposition in hDPSCs.

Long-Term Effect of Modified Glass Ionomer Cement with Mimicked Biological Property of Recombinant Translationally Controlled Protein.

This study modified glass ionomer cement (GIC) by adding mimicked biological molecules to reduce cell death. GIC was modified to BIOGIC by adding chitosan and bovine serum albumin for enhancing protein release. The BIOGIC was supplemented with tricalcium phosphate (TCP) and recombinant translationally controlled tumor protein (TCTP) to improve its biological properties. Four groups of materials, GIC, BIOGIC, BIOGIC+TCP, and BIOGIC + TCP + TCTP, were examined by XRD and SEM-EDX. TCTP released from the specimens was determined by an ELISA method. Human dental pulp stem cells (hDPSCs) were harvested and analyzed by MTT assay, apoptosis, gene expression, and cell differentiation. All groups had the same crystallization characteristic peaks of La2O3. The elemental compositions composed of La, Si, and Al are the main inorganic components. The results show that BIOGIC + TCP + TCTP presented significantly higher percentages of cell viability than other groups on day 1 to day 23 (p < 0.05), but were not different after day 24 to day 41 and had reduced cell apoptosis including BAX, TPT1, BCL-2, and Caspase-3. The BIOGIC + TCP + TCTP demonstrated higher odontoblast mineralization and differentiation markers including ALP activity, DSPP, DMP-1, ALP, BMP-2, and OPN. It enhanced cell proliferation and differentiation as well as mineralization with down-regulation of genes related to apoptosis compared with other groups.

Cryopreservation of Germ Cells of Banana Shrimp (Fenneropenaeus merguiensis) and Black Tiger Shrimp (Penaeus monodon).

Germ cell cryopreservation has been used to preserve many fish species. However, this method has not been established for crustaceans; thus, we attempted to do this herein. The efficiency of slow freezing was compared to vitrification methods for germ cell cryopreservation in two types of marine shrimp, Fenneropenaeus merguiensis and Penaeus monodon. In situ hybridization with a vasa probe was used to identify germ cells. The effects of three cryoprotectants, dimethyl sulfoxide (DMSO), glycerol (GLY), and magnesium chloride (MgCl2), on germ cell viability and recovery rate were compared at three concentrations (5%, 10%, and 15%). The effects of thawing temperature, including 10 and 27 °C, were also investigated. We discovered that 10% DMSO with the vitrification is suitable for preserving the germ cells of F. merguiensis for a long time, whereas 10% GLY with vitrification is suitable for P. monodon. Moreover, the most suitable thawing temperature was 10 °C for both species. This is the first report of germ cell cryopreservation in crustaceans. Thus, we provide evidence that crustacean germ cells can be preserved long-term in liquid nitrogen; this is the first step in the sustainable preservation of crustaceans, especially shrimp.

Effect of the interaction between ribosomal protein L10a and insulin receptor on carbohydrate metabolism.

The number of patients with insulin-resistant diabetes has significantly increased. Thus, alternative insulin mimetics are required for such patients. Some evidences indicate that ribosomal protein L10a (RpL10a) is involved in the insulin pathway. In addition, we previously demonstrated that recombinant RpL10a from Fenneropenaeus merguiensis (Fm-RpL10a) could stimulate cell proliferation and trehalose metabolism in RpL10a-over-expressing flies by inducing insulin receptor (InR) expression and some insulin signaling mediators phosphorylation. In this study, we investigated the in silico binding between Fm-RpL10a and InR. The results indicated that Fm-RpL10a bound to InR at residues 635-640 and 697-702 of the FnIII2 domain. This binding was confirmed using a pull-down and immunofluorescence assay. Further analysis indicated that Fm-RpL10a could stimulate glucose utilisation by insulin-resistant cells (IRCs) and healthy cells. Additionally, Fm-RpL10a at a low concentration (1 µg/ml) altered some glucose metabolism-related genes expression in Fm-RpL10a treated IRCs. The qRT-PCR result revealed the up-regulation of Hk1, which encode key enzymes in glycolysis. Conversely, the expression of G6pc3, which participates in gluconeogenesis, was down-regulated. Overall, the results suggest that Fm-RpL10a can alleviate insulin resistance by stimulating insulin signaling via the FnIII2 domain of InR and activate glycolysis. Therefore, Fm-RpL10a may be a candidate insulin mimetic for the treatment of diabetes.

Biological activities of a recombinant fortilin from Fenneropenaeus merguiensis.

Human Fortilin, an antiapoptotic protein, has also been implicated in several diseases; however, several potential uses of fortilin have also been proposed. Bearing the implications of fortilin in mind, fortilin analog, which has no complication with diseases, is required. Since a recombinant full-length fortilin from Fenneropenaeus merguiensis (rFm-Fortilin (FL)) reported only 44% (3e-27) homologous to human fortilin, therefore the biological activities of the Fm-Fortilin (FL) and its fragments (F2, F12, and F23) were investigated for potential use against HEMA toxicity from filling cement to pulp cell. The rFm-Fortilin FL, F2, 12, and F23 were expressed and assayed for proliferation activity. The rFm-Fortilin (FL) showed proliferation activity on human dental pulp cells (HDPCs) and protected the cells from 2-hydroxy-ethyl methacrylate (HEMA) at 1-20 ng/ml. In contrast, none of the rFm-Fortilin fragments promoted HDPC growth that may be due to a lack of three conserved amino acid residues together for binding with the surface of Rab GTPase for proliferative activity. In addition, rFm-Fortilin (FL) activated mineralization and trend to suppressed production of proinflammatory cytokines, including histamine (at 10 ng/ml) and TNF-α (at 100 ng/ml). Besides, the rFm-Fortilin (FL) did not mutate the Chinese hamster ovary (CHO) cell. Therefore, the rFm-Fortilin (FL) has the potential use as a supplementary medical material to promote cell proliferation in patients suffering severe tooth decay and other conditions.

Lovastatin Production by Aspergillus sclerotiorum Using Agricultural Waste.

RESEARCH BACKGROUND: Lovastatin is a well-known drug used to reduce hypercholesterolaemia. However, the cost of lovastatin production is still high. Therefore, alternative low-cost carbon sources for the production of lovastatin are desirable. EXPERIMENTAL APPROACH: Four different agricultural wastes, namely corn trunks, rice husks, wild sugarcane, and soya bean sludge, were tested separately as substrates to produce lovastatin using a new fungal strain, Aspergillus sclerotiorum PSU-RSPG 178, under both submerged and solid-state fermentation (SSF). RESULTS AND CONCLUSIONS: Of these substrates and cultivation systems, soya bean sludge gave the highest lovastatin yield on dry mass basis of 0.04 mg/g after 14 days of SSF at 25 °C. Therefore, the soya bean sludge was separately supplemented with glucose, wheat flour, trace elements, palm oil, urea and molasses. The addition of the palm oil enhanced the lovastatin yield to 0.99 mg/g. In addition, the optimum conditions, which gave a lovastatin yield of (20±2) mg/g after 18 days of SSF, were soya bean sludge containing 80% moisture (dry basis) at a ratio of soya bean sludge (g) to mycelial agar plugs of 1:4, and a ratio of soya bean sludge (g) to palm oil (mL) of 1:2. Besides, the lovastatin yields obtained from SSF using fresh or dry soya bean sludge were not significantly different. NOVELTY AND SCIENTIFIC CONTRIBUTION: We conclude that A. sclerotiorum PSU-RSPG 178 has a good potential as an alternative strain for producing lovastatin using soya bean sludge supplemented with palm oil as a carbon source.

A novel, 4-h DNA extraction method for STR typing of casework bone samples.

Bones are often found in mass grave crime scene. To increase DNA identification success rates, a highly efficient DNA extraction method should be selected. Several DNA extraction methods for human bones have been published yet never been systematically compared, and some are time-consuming or complex. As such, a quick and highly efficient DNA extraction method was developed and compared with three published methods (Hi-Flow silica-based, total demineralization (TD) and PrepFiler BTA) using 70 fresh and 22 casework bones from different body parts. The highest median DNA concentrations were obtained from developed method (135.85 ng/µL and 0.224 ng/µL for fresh and casework bones, respectively). For residual PCR inhibitors, the threshold cycle (Ct) of the internal positive control (IPC) showed that developed method and PrepFiler BTA removed most PCR inhibitors. Similarly, 95.45% of casework STR profiles obtained using the developed protocol meet the standard requirements for Australian National Criminal Investigative DNA Database (NCIDD) entry, followed by 86.35% using TD, 81.82% using PrepFiler BTA, and 45.45% using Hi-Flow. Additionally, DNA extracts from seven different bones revealed that the 1st distal phalange of the hand contained the highest DNA concentration of 338.43 ng/µL, which was three times higher than the tibia and femur. Our findings suggest that developed method was highly efficient for casework bone analysis. It significantly reduced the extraction processing time down to 4 h and is two to four times cheaper compared with other methods. In practice, both the extraction method and the bone sampling must be considered by a forensic DNA analyst to increase the chances of successful identification.

Abnormal development of zebrafish after knockout and knockdown of ribosomal protein L10a.

In this study, to investigate the secondary function of Rpl10a in zebrafish development, morpholino antisense oligonucleotides (MOs) were used to knock down the zebrafish ribosomal protein L10a (rpl10a). At 25 hpf (hours post-fertilization), embryos injected with the rpl10a MO showed an abnormal morphology, including short bodies, curved tails, and small yolk sac extensions. We observed pigment reductions, edema, larger yolk sacs, smaller eyes and smaller yolk sac extensions at 50 hpf. In addition, reductions in the expression of primordial germ cell (PGC) marker genes (nanos1 and vasa) were observed in rpl10a knockdown embryos. A rescue experiment using a rpl10a mRNA co-injection showed the recovery of the morphology and red blood cell production similar to wild-type. Moreover, the CRISPR-Cas9 system was used to edit the sequence of rpl10a exon 5, resulting in a homozygous 5-bp deletion in the zebrafish genome. The mutant embryos displayed a morphology similar to that of the knockdown animals. Furthermore, the loss of rpl10a function led to reduced expression of gata1, hbae3, and hbbe1 (erythroid synthesis) and increased tp53 expression. Overall, the results suggested that Rpl10a deficiency caused delays in embryonic development, as well as apoptosis and anemia, in zebrafish.

A new cost-effective and fast direct PCR protocol for insects based on PBS buffer.

Insect DNA barcoding is a species identification technique used in biodiversity assessment and ecological studies. However, DNA extraction can result in the loss of up to 70% of DNA. Recent research has reported that direct PCR can overcome this issue. However, the success rates could still be improved, and tissues used for direct PCR could not be reused for further genetic studies. Here, we developed a direct PCR workflow that incorporates a 2-min sample preparation in PBS-buffer step for fast and effective universal insect species identification. The developed protocol achieved 100% success rates for amplification in six orders: Mantodea, Phasmatodea, Neuroptera, Odonata, Blattodea and Orthoptera. High and moderate success rates were obtained for five other species: Lepidoptera (97.3%), Coleoptera (93.8%), Diptera (90.5%), Hemiptera (81.8%) and Hymenoptera (75.0%). High-quality sequencing data were also obtained from these amplifiable products, allowing confidence in species identification. The method was sensitive down to 1/4th of a 1-mm fragment of leg or body and its success rates with oven-dried, ethanol-preserved, food, bat guano and museum specimens were 100%, 98.6%, 90.0%, 84.0% and 30.0%, respectively. In addition, the pre-PCR solution (PBS with insect tissues) could be used for further DNA extraction if needed. The workflow will be beneficial in the fields of insect taxonomy and ecological studies due to its low cost, simplicity and applicability to highly degraded specimens.

Induction of vitellogenesis by glass bottom boat in the female banana shrimp, Fenneropenaeus merguiensis de Man.

In shrimp aquaculture, eyestalk ablation is the only technique that is widely used to accelerate ovarian development. Alternative methods for producing improved ovarian development in broodstock are currently being investigated. Several factors involved in the regulation of ovarian development in shrimp have been investigated. Among these factors, growth factors in the transforming growth factor beta (TGF-ß) superfamily have been implicated as playing potential roles in the regulation of gonad development. In this work, a member of the TGF-ß superfamily known as glass bottom boat (GBB), an ortholog of bone morphogenetic protein (BMP), was investigated to uncover its role in ovarian development in the banana shrimp Fenneropenaeus merguiensis. Full-length cDNA of FmGBB was obtained from transcriptome data. Phylogenetic analysis indicated that the sequence of FmGBB from banana shrimp was similar to those of other arthropods and vertebrate BMP 5/6/7, but was different from those of decapentaplegic proteins and vertebrate BMP 2/4. The FmGBB transcript was found to be widely expressed in shrimp tissues, and its expression in the ovary was dramatically increased in early and late vitellogenic stages during ovarian development and decreased in the mature stage, suggesting its role in vitellogenesis. To study the effects of FmGBB, a soluble recombinant mature FmGBB peptide (His-TF-rgbb) containing both monomers and homodimers was successfully expressed in Escherichia coli. The His-TF-rgbb peptide triggered oocyte proliferation in both cultured ovarian explants and in previtellogenic shrimp upon injection. Interestingly, the injection of His-TF-rgbb into previtellogenic female shrimp stimulated an increase in Vg expression in their ovaries while suppressing production of 20-hydroxyecdysone. Our results suggest the potential role of FmGBB in oocyte proliferation and vitellogenesis; this novel finding can be utilized to stimulate ovarian development in cultured shrimp.

Ribosomal protein L10A and signaling pathway.

Ribosome: machinery in control of messenger RNA's (mRNAs) and several ribosomal proteins are in the small and large subunit of the ribosome. Various aspects of ribosomal proteins have related to cell growth, cell cycle, and diseases. Ribosomal protein L10A (RpL10A) in shrimp and fruit fly has been demonstrated to play a role in oogenesis. Interestingly, deletion RpL10A gene (RpL10Ab-/-) germ line clone of the fruit fly showed a loss of follicle cells surrounding the egg chamber, but nurse cells appeared normal. This phenotype is reminiscent of insulin receptor mutants (InR-/-). In contrast, over-expression of RpL10A in the eyes of the fruit flies resulted in abnormal ommatidia with a loss of red pigment in the center of the eyes. In this study, the abnormal rearrangements of nuclei and lack of cell membranes in those eyes were demonstrated. Furthermore, the expression of InR gene and the InR protein were extensively increased as determined by real-time PCR and immunohistochemistry, respectively. In addition, some insulin signaling mediators were also detected. The Akt and FOXO proteins were highly phosphorylated in the RpL10A over-expressed mutant. The results revealed that RpL10A induced over-expression of the insulin receptor and consequently activated in insulin signaling pathway which affects cell proliferation and we suggest RpL10A stimulates cell proliferation via the insulin signaling pathway.

Effect of recombinant vascular endothelial growth factor and translationally controlled tumor protein on 2­hydroxyethyl methacrylate­treated pulp cells.

Vascular endothelial growth factor (VEGF)-A is a potential signaling protein that may promote angiogenesis. VEGF also helps cells survive in stressfull or hazardous conditions. The present study aimed to compare the effect of VEGF with translationally controlled tumor protein (TCTP), an anti­apoptotic protein in human dental pulp cells (HDPCs), following exposure to 2­hydroxyethyl methacrylate (HEMA), which is a major residual monomer from resin restorative dental materials. Cell viability, alkaline phosphatase (ALP) activity, mineralization and gene expressions for odontogenic and osteogenic differentiation markers of HDPCs were investigated, following exposure to HEMA and in combination with TCTP and VEGF. The results revealed that TCTP at 1 ng/ml and VEGF at 10 ng/ml significantly promoted the proliferation of HDPCs (P<0.05). TCTP (1 ng/ml) and VEGF (10 ng/ml) maintained the cell viability of 4 mM HEMA­treated cells at the same percentage as the control. However, cells treated with HEMA+TCTP+VEGF had a lower cell viability than the groups treated with HEMA and TCTP or VEGF alone. TCTP and VEGF promoted cell proliferation, ALP activity and mineralization, and upregulated of DSPP, DMP­1, BMP­2, and ALP mRNA expression compared with the control. Furthermore, the HEMA+TCTP and HEMA+VEGF groups had significantly higher percentages of calcium deposition than HEMA­treated cells (P<0.001). HEMA was cytotoxic to HDPCs, reduced ALP activity and caused the significant downregulation of odontogenic and osteogenic gene expressions (P<0.05). It was concluded that VEGF and TCTP promoted pulp cell growth and the survival of HEMA­treated cells without synergistic effects. TCTP was required in lower concentrations for these effects. VEGF and TCTP enhanced cell differentiation and mineralization.

Semiquantitative dot-blot immunogold assay for specific detection of white spot syndrome virus.

A dot-blot immunogold assay (DBIA) was developed to detect white spot syndrome virus (WSSV) using the polyclonal antibody VP26 (anti-VP26). The anti-VP26 was immobilized on gold nanoparticles (Ab-AuNPs), and a nitrocellulose membrane was used as a detection pad. When the target WSSV bound to the Ab-AuNPs a reddish dot appeared on the surface of the membrane used within 2-5 Min, which could be seen with the naked eye. The test was able to detect WSSV at concentrations as low as 105 copies µL-1 of WSSV. The DBIA developed had good specificity, and the colloidal gold probe can be applied within 2-3 days when stored at 4 °C. For real sample analysis, the DBIA was applied to samples of seawater used for shrimp cultivation without sample preparation. The results indicate that sample 1 showed a positive result, whereas samples 2 and 3 produced negative results. Then, samples 2 and 3 were spiked with WSSV for method validation. To confirm the performance of the DBIA developed, polymerase chain reaction (PCR) was conducted and the PCR results were the same as those found by the DBIA. Therefore, the DBIA developed could be applied for WSSV detection in real water samples.

In silico analysis of protein toxin and bacteriocins from Lactobacillus paracasei SD1 genome and available online databases.

Lactobacillus paracasei SD1 is a potential probiotic strain due to its ability to survive several conditions in human dental cavities. To ascertain its safety for human use, we therefore performed a comprehensive bioinformatics analysis and characterization of the bacterial protein toxins produced by this strain. We report the complete genome of Lactobacillus paracasei SD1 and its comparison to other Lactobacillus genomes. Additionally, we identify and analyze its protein toxins and antimicrobial proteins using reliable online database resources and establish its phylogenetic relationship with other bacterial genomes. Our investigation suggests that this strain is safe for human use and contains several bacteriocins that confer health benefits to the host. An in silico analysis of protein-protein interactions between the target bacteriocins and the microbial proteins gtfB and luxS of Streptococcus mutans was performed and is discussed here.

Mini-SNaPshot multiplex assays authenticate elephant ivory and simultaneously identify the species origin.

Illegal trading of ivory is mainly responsible for the dramatic decline in elephant populations. Thailand is one of the largest laundering hotspots for African ivory, as the domestic Asian elephant ivory can be legally traded. So, to help combat ivory poaching and smuggling, an efficient method is needed to identify the elephant species from its ivory and ivory products. In this study, a mini-SNaPshot® multiplex assay was developed and fully validated for the identification of confiscated ivory and low DNA template ivory products. Elephantid- and elephant species-specific mitochondrial single nucleotide polymorphisms (SNPs) were identified from 207 mammalian and 1705 elephant/mammoth cytochrome b sequence alignments. Seven informative SNPs were used for assay development. The assay unambiguously and accurately identified authentic elephant ivory and its species of origin on the basis of peak size and color observed in the haplotype profile. The assay was highly efficient for analysis of confiscated ivory and low-template ivory products with a 99.29% success rate (N=140). It was highly reproducible, exhibited no cross-reaction with eight other mammalian DNA; and had 100% identification accuracy. In addition, nested and direct PCR amplification were also compatible with the developed assay. This efficient assay should benefit wildlife forensic laboratories and aid in the prosecution of elephant-related crimes.

Ovarian Transcriptome Analysis of Vitellogenic and Non-Vitellogenic Female Banana Shrimp (Fenneropenaeus merguiensis).

The banana shrimp (Fenneropenaeus merguiensis) is one of the most commercially important penaeid species in the world. Its numbers are declining in the wild, leading to a loss of broodstock for farmers of the shrimp and a need for more successful breeding programs. However, the molecular mechanism of the genes involved in this shrimp's ovarian maturation is still unclear. Consequently, we compared transcriptomic profiles of ovarian tissue from females in both the vitellogenic stage and the non-vitellogenic stage. Using RNA-Seq technology to prepare the transcriptome libraries, a total of 12,187,412 and 11,694,326 sequencing reads were acquired from the non-vitellogenic and vitellogenic stages respectively. The analysis of the differentially expressed genes identified 1,025 which were significantly differentially expressed between the two stages, of which 694 were up-regulated and 331 down-regulated. Four genes putatively involved in the ovarian maturation pathway were chosen for validation by quantitative real-time PCR (RT-qPCR). The data from this study provided information about gene expression in ovarian tissue of the banana shrimp which could be useful for a better understanding of the regulation of this species' reproductive cycle.