Progress on genomics and locus of important agronomic traits in Chenopodium quinoa.

Quinoa (Chenopodium quinoa, Willd.) as a new health food in the 20th century, its comprehensive nutritional composition, stress resistance and other characteristics have been paid much of attention, and enjoys the reputation of "nutritional gold", "vegetarian king" and "food in the future" in the world. In recent years, with the rapid development of genomics and high-throughput sequencing technology, the high-quality whole genome sequence of quinoa has been completed, and the omics analysis and functional research of a series of key genes have been gradually carried out. In this review, we summarize the research progress in quinoa genomics, gene family analysis of important transcription factors, genetic map construction, QTL mapping of important traits, and genes for important agronomic and yield traits. Moreover, according to the current status of quinoa breeding, this paper also put forward five key problems in quinoa breeding, and pointed out four important directions of genetic improvement and breeding of quinoa in the future, so as to provide reference for the realization of directional genetic improvement of quinoa in the future.

The complete mitochondrial genome of Neobenedenia melleni (Platyhelminthes: Monogenea): mitochondrial gene content, arrangement and composition compared with two Benedenia species.

The complete mitochondrial (mt) genome sequences of Neobenedenia melleni were determined and compared with those of Benedenia seriolae and B. hoshinai. This circular genome comprises 13,270 bp and includes all 36 typical mt genes found in flatworms. Total AT content of N. melleni is 75.9 %. ATG is the most common start codon, while nad4L is initiated by GTG. All protein-coding genes are predicted to terminate with TAG and TAA. N. melleni has the trnR with a TCG anticodon, which is the same to B. seriolae but different from B. hoshinai (ACG). The mt gene arrangement of N. melleni is similar to that of B. seriolae and B. hoshinai with the exception of three translocations (trnF, trnT and trnG). The overlapped region between nad4L and nad4 was found in the N. melleni mt genome, which was also reported for the published Gyrodactylus species, but it was not found in those of B. seriolae and B. hoshinai, which are non-coding regions instead. The present study provides useful molecular characters for species or strain identification and systematic studies of this parasite.

Effects of Nitrogen Accumulation, Transportation, and Grain Nutritional Quality and Advances in Fungal Endophyte Research in Quinoa (Chenopodium quinoa Willd.) Plants.

This study aims to understand the influence of nitrogen accumulation, fungal endophyte, yield, nitrogen use efficiency, and grain nutritional quality parameters on the yield of quinoa in some areas of China. The endophytic microbial community in plants plays a crucial role in plant growth, development, and health, especially in quinoa plants under different nitrogen fertilizer levels. The results from the present study indicated that appropriate nitrogen application significantly enhanced the nitrogen accumulation and yield of quinoa grains during maturity, increasing by 34.54-42.18% and 14.59-30.71%, respectively. Concurrently, protein content, amylose, total starch, ash, and fat content also increased, with respective growth rates of 1.15-18.18%, 30.74-42.53%, 6.40-12.40%, 1.94-21.94%, and 5.32-22.22%. Our constructed interaction network of bacterial and fungal communities revealed that bacteria outnumbered fungi significantly, and most of them exhibited synergistic interactions. The moderate increase in N150 was beneficial for increasing quinoa yield, achieving nitrogen use efficiency (NUE) of over 20%. The N210 was increased, and both the yield and NUE significantly decreased. This study provides novel insights into the impact of nitrogen fertilizer on quinoa growth and microbial communities, which are crucial for achieving agricultural sustainable development.

Genome-wide identification, phylogeny and expression analysis of the R2R3-MYB gene family in quinoa (Chenopodium quinoa) under abiotic stress.

The MYB transcription factor (TF) are among the largest gene families of plants being responsible for several biological processes. The R2R3-MYB gene family are integral player regulating plant primary and secondary metabolism, growth and development, and responses to hormones and stresses. The phylogenetic analysis combined with gene structure analysis and motif determination resulted in division of R2R3-MYB gene family into 27 subgroups. Evidence generated from synteny analyses indicated that CqR2R3-MYBs gene family is featured by tandem and segmental duplication events. On the basis of RNA-Seq data, the expression patterns of different tissues under salt treatment were investigated resulting CqR2R3-MYB genes high expression both in roots and stem of quinoa (Chenopodium quinoa ) plants. More than half of CqR2R3-MYB genes showed expression under salt stress. Based on this result, CqR2R3-MYB s may regulate quinoa plant growth development and resistance to abiotic stresses. These findings provided comprehensive insights on role of CqR2R3-MYBs gene family members in quinoa and candidate MYB gene family members can be further studies on their role for abiotic stress tolerance in crop plants.

Experiment Study on the Influence of Density and Confining Pressure on Triaxial Shear Properties of Calcareous Sand.

Calcareous sand is one of the main building materials in the construction of islands and reefs, and its shear property is very important for predicting their strength and deformation. However, the correlation research on the shear properties of calcareous sand is limited. In this paper, a series of the triaxial consolidation drainage shear tests of calcareous sand with relative densities (Dr) of 70% and 90% under confining pressures of 100, 200, 400 and 800 kPa were carried out by a triaxial testing apparatus, and the effects of relative density and confining pressure on the deformation and strength characteristics of calcareous sand were analyzed. The results show that the stress-strain curves of calcareous sand show a strain softening characteristic, and both peak deviatoric stress and failure strain increase with confining pressure, but the increase in failure strain is restrained when the confining pressure is larger than 400 kPa. The initial shear modulus of calcareous sand is positively correlated with confining pressure. Additionally, the molar circular envelope of calcareous sand is linear in the range of 100~400 kPa, but it deviates from linear when confining pressure exceeds 400 kPa. The critical state line (CSL) of calcareous sand is nonlinear, with almost the same exponent for calcareous sand with different relative densities. The research results have important reference value for the foundation construction of islands and reefs.

Discussion on Static Resistance of Granite under Penetration.

A total of 9 tests were carried out with 30 mm and 78 mm caliber scaled projectiles penetrating into granite targets. The penetration depth, crater diameter, and mass loss rate were examined and discussed. The results indicate that the dimensionless penetration depth of large-caliber projectiles is 20% greater than small-caliber projectiles. Based on the description of static resistance Ra in the Forrestal semi-empirical formula, the size effect of dimensionless penetration depth can be attributed to the size effect of static resistance Ra, and it can be seen that the penetration static resistance of projectile A is 40% higher than that of projectile B. Numerical simulations of projectile penetration into granite targets were conducted using the finite element program ANSYS/LS-DYNA. In terms of penetration depth and crater damage, the numerical simulation results agree well with the test data. This suggests that the selection of parameters was reasonable. The influence of compressive strength, projectile striking velocity, mass, diameter, and caliber-radius-head (CRH) ratio on the static resistance Ra were studied by RHT model parameterization. Based on the numerical results from the parametric study, an empirical formula was derived to predict the static resistance Ra.

The complete mitochondrial genome of Pseudochauhanea macrorchis (Monogenea: Chauhaneidae) revealed a highly repetitive region and a gene rearrangement hot spot in Polyopisthocotylea.

The complete mitochondrial genome of Pseudochauhanea macrorchis was determined and compared with other monogenean mitochondrial genomes from GenBank. The circular genome was 15,031 bp in length and encoded 36 genes (12 protein-coding genes, two ribosomal RNAs, and 22 transfer RNAs) typically found in flatworms. Structures of the mitochondrial genome were mostly concordant with that known for Microcotyle sebastis and Polylabris halichoeres, but also contained two noted features-a gene rearrangement hot spot and the highly repetitive region (HRR) in major non-coding region (NCR). The gene rearrangement hot spot located between the cox3 and nad5 genes, including a cluster of tRNA genes, nad6 gene and one major NCR. The HRR seemed to be a unique feature of the polyopisthocotylean mitochondrial genomes. In conclusion, the present study provided new molecular data for future studies of the comparative mitochondrial genomics and also served as a resource of markers for the studies of species populations and monogenean phylogenetics.

Oesophagostomum dentatum and Oesophagostomum quadrispinulatum: characterization of the complete mitochondrial genome sequences of the two pig nodule worms.

In the present study, the complete mitochondrial DNA (mtDNA) sequences of the pig nodule worm Oesophagostomum quadrispinulatum were determined for the first time, and the mt genome of Oesophagostomum dentatum from China was also sequenced for comparative analysis of their gene contents and genome organizations. The mtDNA sequences of O. dentatum China isolate and O. quadrispinulatum were 13,752 and 13,681 bp in size, respectively. Each of the two mt genomes comprises 36 genes, including 12 protein-coding genes, two ribosomal RNA and 22 transfer RNA genes, but lacks the ATP synthetase subunit 8 gene. All genes are transcribed in the same direction and have a nucleotide composition high in A and T. The contents of A+T are 75.79% and 77.52% for the mt genomes of O. dentatum and O. quadrispinulatum, respectively. Phylogenetic analyses using concatenated amino acid sequences of the 12 protein-coding genes, with three different computational algorithms (maximum likelihood, maximum parsimony and Bayesian inference), all revealed that O. dentatum and O. quadrispinulatum represent distinct but closely-related species. These data provide novel and useful markers for studying the systematics, population genetics and molecular diagnosis of the two pig nodule worms.

Blockade of TIM-3 and PD-1 enhances the antitumor effects of MAGE-A11 antigen-specific cytotoxic T lymphocytes.

Cancer immunotherapy is an attractive approach for antigen-specific T cell-mediated antitumor therapy, especially for the induction of cytotoxic T lymphocytes (CTLs). An human leukocyte antigen (HLA)-A2-restricted melanoma-associated antigen-A11 (MAGE-A11) peptide was developed that exhibited a potent capacity to induce cytotoxicity towards MAGE-A11-positive breast cancer cells by activating CTLs. However, this antitumor immune response can be suppressed by inhibitory pathways. Programmed death-1 (PD-1) and T cell immunoglobulin domain and mucin domain 3 (TIM-3) pathways are two important pathways involved in the tumor-mediated immune suppression. The present study aimed to augment the efficacy of MAGE-A11 antigen-specific CTLs via blocking PD-1 and TIM-3. The results showed that the expression levels of PD-1 and TIM-3 were unregulated during T cell induction, expansion and target cell killing. Blockade of PD-1 and TIM-3 modulated T cell proliferation, transformation and survival. In addition, treatment of cells with antibodies against PD-1 and TIM-3 enhanced the cytotoxicity of MAGE-A11 antigen-specific CTLs against breast cancer cells. The aforementioned findings suggested that MAGE-A11 antigen-specific CTLs accompanied by PD-1 and TIM-3 blockade could be considered as a potential therapy approach for breast cancer.

Correlation between promoter methylation of the LDH-C4 gene and DNMT expression in breast cancer and their prognostic significance.

DNA methylation plays an important role in tumorigenesis and development. The potential of aberrant DNA methylation to act as a biomarker for tumor diagnosis and prognostic evaluation is currently being explored. Lactate dehydrogenase C4 (LDH-C4) is a member of the cancer/testis antigen family that is expressed in a broad range of human tumor types, with particularly high expression patterns observed in lung cancer, melanoma and breast cancer. However, whether the methylation of the promoter region of LDH-C4 can be used as a tumor marker and its association with prognosis remain poorly understood. The present study aimed to determine the potential of the methylation status of LDH-C4 and DNA methyltransferase (DNMT) expression to be biological markers for the prognostic evaluation of breast cancer. Methylation-specific PCR was conducted to evaluate alterations in the methylation levels of LDH-C4. Immunohistochemical analysis was performed to assess the expression levels of DNMTs in breast cancer tissues. The association between the methylation status of LDH-C4 or the expression of DNMTs, and clinical pathological parameters was also evaluated. The results of the present study revealed that the demethylation rate of LDH-C4 in breast cancer tissues was significantly increased compared with that of adjacent tissues, and associations were identified between the demethylation of LDH-C4 and the histological grade, estrogen receptor, progesterone receptor and HER-2 status, and lymph node metastasis. The level of LDH-C4 demethylation was negatively correlated with the expression of DNMTs. Demethylation of the LDH-C4 promoter and DNMT expression predicted an unfavorable prognosis of patients with breast cancer. In addition, demethylation of the LDH-C4 promoter, high expression of DNMT3a and DNMT3b, histological grade and lymph node metastasis were all discovered to be independent prognostic factors in patients with breast cancer. In conclusion, results of the present study indicated that the demethylation of LDH-C4 and DNMT expression levels may be closely associated with the occurrence and development of breast cancer, in addition to lymph node metastasis. Thus, both may be used to assist the clinical evaluation of prognosis.

[Advances in liquid chromatography-mass spectrometry analysis of several important secondary metabolites in plant metabolomics].

Metabolomics, which mainly studies the metabolite components of organisms, tissues, cells and their dynamic changes, is an emerging omics technology following genomics and proteomics. Metabolites are the final products of cellular regulation, and the concentration of metabolites is considered to be the ultimate response of a biological system to genetic or environmental changes. Secondary metabolites with chemical diversity are widely present in living organisms, thus accurate quantification of secondary metabolites through appropriate analytical platforms is an important task of metabolomics. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) is the most commonly used method for the detection of metabolites, providing a basis for the wide application of plant secondary metabolites. This review summarizes the advances of using LC-MS/MS techniques for the detection of phytohormone, folic acid, flavonoids and other secondary metabolites.

Epidemiology, Environmental Risks, Virulence, and Resistance Determinants of Klebsiella pneumoniae From Dairy Cows in Hubei, China.

Klebsiella pneumoniae (K. pneumoniae) is an opportunistic pathogen, which causes serious infections in humans and animals. To investigate the antimicrobial resistance pattern and virulence profile of K. pneumoniae, a total of 887 samples were collected from both the healthy and mastitis cows and the bedding, feed, feces, air, drinking water, spraying water, washing water, and milk cup swabs from five dairy farms in Hubei, China, during 2019 and 2020. K. pneumoniae was isolated and identified using PCR of the khe and 16S rDNA sequencing. A genotypic characterization was performed for K. pneumoniae isolates using wzi typing and multilocus sequence typing (MLST). Antimicrobial resistances were confirmed using broth microdilution against 17 antimicrobial agents and resistance and virulence genes were determined by PCR. The prevalence of K. pneumoniae was 26.94% (239/887) distributed in 101 wzi allele types (199/239, 83.26%) and 100 sequence types (STs) (209/239, 87.45%), including 5 new wzi allele type and 25 new STs. Phylogenetic analysis showed that K. pneumoniae isolated from milk, nipple swab, feed, and feces is classified in the same clone complex. By comparing with the PubMLST database, at least 67 STs have the risk of spreading in different species and regions. Interestingly, 60 STs have been isolated from humans. The isolates were highly sensitive to meropenem and colistin, but resistant to ampicillin (100%), sulfisoxazole (94.56%), cephalothin (47.28%), streptomycin (30.13%), and so on. Noteworthy, multidrug-resistant (MDR) rate was found to be 43.93% in this study. By PCR, 30 of 68 antimicrobial resistance (AMR) genes were identified; the prevalence rate of blaTEM, blaSHV, strA, strB, aadA1, and aac(6')-Ib-cr was more than 50%. Eleven CTX-M-producing K. pneumoniae were found. The detection rate of fimH, mrkD, uge, wabG, entB, iutA, iroN, and ureA was over 85%. This study reinforces the epidemiological importance of K. pneumoniae in food-producing animals in Hubei. The emergence and spread of environmental MDR K. pneumoniae may pose a potential threat to food safety and public health.

Electrophoretic detection of genetic variability among Schistosoma japonicum isolates by sequence-related amplified polymorphism.

In the present study, sequence-related amplification polymorphism (SRAP) was utilized to study the genetic variability among Schistosoma japonicum isolates from different provinces in China, using Schistosoma mansoni from Puerto Rico for comparison. Five out of ten tested SRAP primer combinations displayed significant polymorphisms among S. japonicum isolates from China, namely ME2/EM1, ME4/EM1, ME4/EM6, ME5/EM4 and ME5/EM5. Analysis of the 61 S. japonicum samples from China with five SRAP primer combinations identified a total of 83 reproducible polymorphic fragments. The number of fragments using each primer combination ranged from 14 to 19, with an average of 16 polymorphic bands per primer pair, and the size of fragment ranged approximately from 100 to 1000 bp. Representative-specific SRAP fragments were excised from the gels, and confirmed by PCR amplification of genomic DNA using primers designed and based on the sequences of these SRAP fragments. Based on SRAP profiles, unweighted pair-group method with arithmetic averages (UPGMA) dendrogram was constructed. UPGMA clustering algorithm categorized S. japonicum isolates from China into nine clades and two lineages (representing the mountainous and lake/marshland regions). These results indicate the usefulness of the SRAP technique for revealing genetic variability among S. japonicum isolates from China, and the SRAP technique should be applicable to other living organisms.

Cloning and characterization of two ferritin subunit genes from bay scallop, Argopecten irradians (Lamarck 1819).

We have cloned two full-length cDNAs from two ferritin genes (Aifer1 and Aifer2) of the bay scallop, Argopecten irradians (Lamarck 1819). The cDNAs are 1,019 and 827 bp in length and encode proteins of 171 and 173 amino acids, respectively. The 5' UTR of each contains a conserved iron response element (IRE) motif. Sequence analyses reveal that both proteins belong to the H-ferritin family with seven conserved amino acids in the ferroxidase center. Highest expression of Aifer1 is found in the mantle and adductor muscle, while that of Aifer2 is only in the latter tissue. These Aifer genes are differentially expressed following bacterial challenge of the scallop. The expression level of Aifer1 was acutely up-regulated (over 10 fold) at 6 h post-bacteria injection, whereas Aifer2 expression was not significantly changed by bacterial challenge. Both genes were effectively expressed in E. coli BL21 (DE3), producing proteins of similar molecular weight, approximately 23 kDa. Purified Aifer1 and Aifer2 proteins exhibited iron-chelating activity of 33.1% and 30.4%, respectively, at a concentration of 5 mg/ml. Cations, Mg(2+), Zn(2+) and Ca(2+), depressed iron-chelating activity of both proteins. Additionally, the E. coli cells expressing recombinant Aifer1 and Aifer2 showed tolerance to H(2)O(2), providing a direct evidence of the antioxidation function of ferritin. The results presented in this study suggest important roles of Aifer1 and Aifer2 in the regulation of iron homeostasis, immune response, and antioxidative stress in A. irradians.

Expression of LINC01606 in multiple myeloma and its effect on cell invasion and migration.

Given the increasing incidence of multiple myeloma (MM) in recent years, a full understanding of its pathogenesis to find effective molecular markers carries huge implications for future clinical diagnosis and treatment of MM. As the research advances, accumulating studies have pointed out that long non-coding RNAs (LncRNAs) may be the key to the future diagnosis and treatment of neoplastic diseases. OBJECTIVE: This study investigated the clinical implications of LncRNA LINC01606 in MM and its effects on the biological behavior of MM cells. METHODS: In this prospective study, 72 patients with MM (group A) admitted between July 2014 and July 2016 and 68 healthy subjects (group B) who concurrently underwent physical examination in our hospital were included. The expression of LINC01606 in peripheral blood of patients in the two groups was detected to analyze its diagnostic and prognostic value in MM. In addition, MM cells were purchased and transfected with plasmids for mimics, inhibitors and negative control of LINC01606 and miR-579-3p respectively to detect the changes in cell proliferation, invasion and migration. RESULTS: The expression of LINC01606 in group A was higher than that in group B (P<0.050). The sensitivity and specificity of peripheral blood LINC01606 in predicting MM were 85.29% and 72.39%, respectively (P<0.001). Prognostic follow-up analysis revealed higher LINC01606 levels in the dead than those in the survival. The predictive sensitivity of LINC01606 for the 3-year mortality of MM patients was 63.16%, and the specificity was 86.00% (P<0.001). Higher expression of LINC01606 indicated increased risk of 3-year mortality in patients with MM (P<0.001). Compared with LINC01606 overexpression and miR-579-3p inhibition, the proliferation, invasion and migration of cells decreased more significantly by LINC01606 inhibition and miR-579-3p overexpression (P<0.050). Dual luciferase reporter (DLR) assay confirmed the targeting relationship between LINC01606 and miR-579-3p. There was no significant difference in the activity of MM cells co-transfected with LINC01606-inhibitor and miR-579-3p-inhibitor plasmids compared with the blank group (P>0.050). CONCLUSIONS: LINC01606, with a high expression profile in MM, promotes the proliferation, migration and invasion of MM cells through targeted inhibition of miR-579-3p, which may be the key to future diagnosis and treatment of MM.

Transcriptome Analysis Reveals the Molecular Mechanism of GABA Accumulation during Quinoa (Chenopodium quinoa Willd.) Germination.

Quinoa (Chenopodium quinoa Willd.) with a history of 5000 years as food is extremely rich in nutrients and bioactive compounds, including γ-aminobutyric acid (GABA), a natural four-carbon non-protein amino acid with great benefits to human health. In quinoa, GABA generally increases with the germination time, but the underlying molecular mechanism is unclear. Here, we found that the GABA content in quinoa varied significantly among 25 varieties using an automatic amino acid analyzer. Next, six varieties (three low-GABA and three high-GABA varieties) were used for further analyses. The content of GABA in six varieties all showed an increasing trend after germination. In addition, Pearson's correlation analysis showed that the changes in GABA content were closely related to the transcript level or enzyme activity of three key enzymes including glutamate decarboxylase (GAD), GABA transaminase (GABA-T), and succinate-semialdehyde dehydrogenase (SSADH) in the GABA shunt, especially GAD. Based on RNA-sequencing analysis, eight GAD genes, two GABA-T genes, one SSADH gene, nine polyamine oxidase (PAO) genes, five diamine oxidase (DAO) genes, four 4-aminobutyraldehyde dehydrogenase (BADH) genes, and three thermospermine synthase ACAULIS5 (ACL5) genes were identified. Among these, CqGAD8 and CqGABA-T2 may make a greater contribution to GABA accumulation during quinoa germination.

Structure, preparation, modification, and bioactivities of ß-glucan and mannan from yeast cell wall: A review.

In order to solve the antibiotic resistance, the research on antibiotic substitutes has received an extensive attention. Many studies have shown that ß-glucan and mannan from yeast cell wall have the potential to replace antibiotics for the prevention and treatment of animal diseases, thereby reducing the development and spread of antibiotic-resistant bacterial pathogens. ß-Glucan and mannan had a variety of biological functions, including improving the intestinal environment, stimulating innate and acquired immunity, adsorbing mycotoxins, enhancing antioxidant capacity, and so on. The biological activities of ß-glucan and mannan can be improved by chemically modifying its primary structure or reducing molecular weight. In this paper, the structure, preparation, modification, and biological activities of ß-glucan and mannan were reviewed, which provided future perspectives of ß-glucan and mannan.

The mitogenome of Paphia euglypta (Bivalvia: Veneridae) and comparative mitogenomic analyses of three venerids.

Extraordinary variation has been found in mitochondrial (mt) genome inheritance, gene content and arrangement among bivalves. However, only few bivalve mt genomes have been comparatively analyzed to infer their evolutionary scenarios. In this study, the complete mt genome of the venerid Paphia euglypta (Bivalvia: Veneridae) was firstly studied and, secondly, it was comparatively analyzed with other venerids (e.g., Venerupis philippinarum and Meretrix petechialis) to better understand the mt genome evolution within a family. Though several common features such as the AT content, codon usage of protein-coding genes, and AT/GC skew are shared by the three venerids, a high level of variability is observed in genome size, gene content, gene order, arrangements and primary sequence of nucleotides or amino acids. Most of the gene rearrangement can be explained by the "tandem duplication and random loss" model. From the observed rearrangement patterns, we speculate that block interchange between adjacent genes may be common in the evolution of mt genomes in venerids. Furthermore, this study presents several new findings in mt genome annotation of V. philippinarum and M. petechialis, and hence we have reannotated the genome of these two species as: (1) the ORF of the formerly annotated cox2 gene in V. philippinarum is deduced by using a truncated "T" codon and a second cox2 gene is identified; (2) the trnS-AGN gene is identified and marked in the mt genome of both venerids. Thus, this study demonstrated a high variability of mt genomes in the Veneridae, and showed the importance of comparative mt genome analysis to interpret the evolution of the bivalve mt genome.

Genetic characterization of ticks from southwestern Romania by sequences of mitochondrial cox1 and nad5 genes.

In the present study, samples representing three hard tick species and one soft tick species, namely Dermacentor marginatus, Haemaphysalis punctata, Ixodes ricinus and Argas persicus from southwestern Romania, and one hard tick, Haemaphysalis longicornis, from China were characterized genetically by a portion of mitochondrial cytochrome c oxidase subunit 1 gene (pcox1) and a portion of nicotinamide adenine dinucleotide dehydrogenase subunit 5 gene (pnad5). The pcox1 and pnad5 were amplified separately from individual ticks by PCR, sequenced and analyzed. The length of pcox1 and pnad5 sequences of all samples was 732 and 519 bp, respectively. The intra-specific sequence variation in De. marginatus was 0.1-1.0% for pcox1 and 0.2-1.2% for pnad5, whereas in Ha. punctata it was 0.4-1.9% for pcox1 and 0.4-1.0% for pnad5. For the tick species examined in the present study, sequence comparison revealed that the inter-specific sequence differences were higher: 15.9-27.6% for pcox1 and 20.3-42.4% for pnad5. This suggests that the cox1 and nad5 sequences could provide useful genetic markers for the specific identification and genetic characterization of ticks in Romania and elsewhere.

Effects of dietary riboflavin levels on antioxidant defense of the juvenile grouper Epinephelus coioides.

This study was conducted to evaluate the effects of dietary riboflavin on antioxidant defense in the juvenile grouper Epinephelus coioides. Graded levels of riboflavin (0.9, 1.6, 4.4, 6.7, 12.9 and 19.4 mg kg(-1) dry diet) were fed to grouper juveniles (mean weight: 14.90 +/- 0.46 g) for 12 weeks. Higher levels of liver thiobarbituric acid reactive substances (TBARS) content were observed in grouper fed low doses (0.9 and 1.6 mg kg(-1) diet) of riboflavin. Both liver glutathione reductase (GR) activity and its activation coefficient (GR-AC) poorly responded to riboflavin deficiency. In addition, other indices of the glutathione-dependent defense system, including the activities of glutathione peroxidase (GSH-PX) and glutathione-S-transferase (GST), and the content of glutathione (GSH), were also non-significantly affected by dietary riboflavin levels. However, the activities of liver superoxide dismutase (SOD) and catalase (CAT) were significantly lower in fish fed 0.9 mg kg(-1) diet, with a positive correlation between the different groups. In conclusion, the present study indicated that the juvenile grouper fed the riboflavin-unsupplemented diet was susceptible to lipid peroxidation (LPO), with lower SOD and CAT activities in the liver. However, the glutathione-dependent defense system of grouper was not affected by dietary riboflavin levels.