Protective role for C3aR in experimental chronic pyelonephritis.

Emerging evidence suggest that C3aR plays important roles in homeostasis, host defense and disease. Although it is known that C3aR is protective in several models of acute bacterial infections, the role for C3aR in chronic infection is largely unknown. Here we show that C3aR is protective in experimental chronic pyelonephritis. Global C3aR deficient (C3ar-/- ) mice had higher renal bacterial load, more pronounced renal histological lesions, increased renal apoptotic cell accumulation, tissue inflammation and extracellular matrix deposition following renal infection with uropathogenic E. coli (UPEC) strain IH11128, compared to WT control mice. Myeloid C3aR deficient (Lyz2-C3ar-/- ) mice exhibited a similar disease phenotype to global C3ar-/- mice. Pharmacological treatment with a C3aR agonist reduced disease severity in experimental chronic pyelonephritis. Furthermore, macrophages of C3ar-/- mice exhibited impaired ability to phagocytose UPEC. Our data clearly demonstrate a protective role for C3aR against experimental chronic pyelonephritis, macrophage C3aR plays a major role in the protection, and C3aR is necessary for phagocytosis of UPEC by macrophages. Our observation that C3aR agonist curtailed the pathology suggests a therapeutic potential for activation of C3aR in chronic infection.

Accurate Relativistic Chiral Nucleon-Nucleon Interaction up to Next-to-Next-to-Leading Order.

We construct a relativistic chiral nucleon-nucleon interaction up to the next-to-next-to-leading order in covariant baryon chiral perturbation theory. We show that a good description of the np phase shifts up to T_{lab}=200 MeV and even higher can be achieved with a χ[over ˜]^{2}/d.o.f. less than 1. Both the next-to-leading-order results and the next-to-next-to-leading-order results describe the phase shifts equally well up to T_{lab}=200 MeV, but for higher energies, the latter behaves better, showing satisfactory convergence. The relativistic chiral potential provides the most essential inputs for relativistic ab initio studies of nuclear structure and reactions, which has been in need for almost two decades.

Development of a highly thermostable immunoassay based on a nanobody-alkaline phosphatase fusion protein for carcinoembryonic antigen detection.

Carcinoembryonic antigen-related cell adhesion molecule 5 (CEACAM-5) assays are employed in routine clinical settings to diagnose tumor. We selected two nanobodies with high-affinity to CEACAM-5, termed Nb11C12 and Nb2D5, using phage-display technology. The Nb2D5 fused with calf intestinal alkaline phosphatase (CAP), human placental alkaline phosphatase (HAP), or Pyrococcus abyssi alkaline phosphatase (PAP) were expressed in human embryonic kidney (HEK293) cells. The enzymatic activity of Nb2D5-HAP fusion protein was the best and remained stable at 60 °C for 7 days. The affinity of Nb2D5-HAP fusion protein to CEACAM-5 reached 42 pM. A chemiluminescent enzyme immunoassay (CLEIA) based on Nb2D5-HAP fusion protein was established for quantitative CEACAM-5 assay in clinical settings. The CLEIA exhibited a wide linear range of 0.31-640 ng/mL toward CEACAM-5, with a limit of detection (LOD) of 0.85 ng/mL. No cross-reactivity occurred with CEACAM-1, CEACAM-3, CEACAM-6, or CEACAM-8, and no interference was observed with rheumatoid factors. The CLEIA based on Nb2D5-HAP fusion protein was stable for 8 weeks at 37 °C and 50% relative humidity. The CLEIA developed from Nb2D5-HAP fusion protein had much better stability and linearity with similar reproducibility compared with the enzyme-linked immunosorbent assay developed from conventional monoclonal antibodies, which have been widely used in clinics over the past several decades. Graphical abstract.

A novel electrochemical DNA biosensor construction based on layered CuS-graphene composite and Au nanoparticles.

A novel CuS-graphene (CuS-Gr) composite was synthesized to achieve excellent electrochemical properties for application as a DNA electrochemical biosensor. CuS-Gr composite was prepared by a hydrothermal method, in which two-dimensional graphene served as a two-dimensional conductive skeleton to support CuS nanoparticles. A sensitive electrochemical DNA biosensor was fabricated by immobilizing single-stranded DNA (ss-DNA) labeled at the 5' end using 6-mercapto-1-hexane (HS-ssDNA) on the surface of Au nanoparticles (AuNPs) to form ssDNA-S-AuNPs/CuS-Gr, and hybridization sensing was done in phosphate buffer. Cyclic voltammetry and electrochemical impedance spectroscopy were performed for the characterization of the modified electrodes. Differential pulse voltammetry was applied to monitor the DNA hybridization using an [Fe(CN)6](3-/4-) solution as a probe. Under optimum conditions, the biosensor developed exhibited a good linear relationship between the current and the logarithm of the target DNA concentration ranging from 0.001 to 1 nM, with a low detection limit of 0.1 pM (3σ/S). The biosensor exhibited high selectivity to differentiate one-base-mismatched DNA and three-base-mismatched DNA. The results indicated that the sensing platform based on CuS-Gr provides a stable and conductive interface for electrochemical detection of DNA hybridization, and could easily be extended to the detection of other nucleic acids.

In silico evidence implicating novel mechanisms of Prunella vulgaris L. as a potential botanical drug against COVID-19-associated acute kidney injury.

COVID-19-associated acute kidney injury (COVID-19 AKI) is an independent risk factor for in-hospital mortality and has the potential to progress to chronic kidney disease. Prunella vulgaris L., a traditional Chinese herb that has been used for the treatment of a variety of kidney diseases for centuries, could have the potential to treat this complication. In this study, we studied the potential protective role of Prunella vulgaris in COVID-19 AKI and explored its specific mechanisms applied by network pharmacology and bioinformatics methods. The combination of the protein-protein interaction network and Gene Ontology and Kyoto Encyclopedia of Genes and Genomes enrichment -target gene network revealed eight key target genes (VEGFA, ICAM1, IL6, CXCL8, IL1B, CCL2, IL10 and RELA). Molecular docking showed that all these eight gene-encoded proteins could be effectively bound to three major active compounds (quercetin, luteolin and kaempferol), thus becoming potential therapeutic targets. Molecular dynamics simulation also supports the binding stability of RELA-encoded protein with quercetin and luteolin. Together, our data suggest that IL6, VEGFA, and RELA could be the potential drug targets by inhibiting the NF-κB signaling pathway. Our in silico studies shed new insights into P. vulgaris and its ingredients, e.g., quercetin, as potential botanical drugs against COVID-19 AKI, and warrant further studies on efficacy and mechanisms.

Higher Sodium Channel Excitability in Cardiac Purkinje Fibers: Implications for Multifocal Ectopic Purkinje-Related Premature Contractions.

BACKGROUND: Multifocal ectopic Purkinje-related premature contractions (MEPPCs) are associated with SCN5A variants. However, it is not well understood why Purkinje fibers, but not ventricular myocardium, play a predominant role in arrhythmogenesis. OBJECTIVES: This study sought to explore the underlying mechanisms of MEPPC. METHODS: Whole-cell patch-clamp and molecular biology techniques were used in the present study. RESULTS: Clinical data from one patient with R814W variant showed MEPPC syndrome, which is well responsive to amiodarone. Compared with canine ventricular myocytes, Purkinje cells (PCs) had significantly larger sodium current (INa), leftward shift of INa activation and inactivation curves, suggesting higher sodium channel excitability in PCs. Real-time polymerase chain reaction and Western blot analysis showed that the mRNA and protein expression of NaVß1 and NaVß3 was higher in canine Purkinje fibers than in ventricular myocardium. INa in heterologous Chinese hamster ovary cell expression system co-expressing NaV1.5 and NaVß1/NaVß3 exhibited similar biophysical properties of INa in PCs. R814W variant shifted INa activation in a hyperdepolarized direction, caused a larger window current, and generated an outward-gating pore current at depolarized voltages. Coexpression of NaVß1/NaVß3 with Nav1.5-R814W further left-shifted INa activation and caused an even larger window current and gating pore current, suggesting higher susceptibility of Purkinje fibers to R814W variant. Amiodarone inhibited INa, shifted its inactivation to more negative voltages, and significantly decreased the window current. CONCLUSIONS: A higher expression of ß1 and ß3 subunits contributes to higher sodium channel excitability in cardiac Purkinje fibers, making them more susceptible to MEPPC.

[Effect of hepatocyte growth factor and transforming growth factor-ß(1) on atrial fibroblasts fibrosis].

OBJECTIVE: To investigate the effect of hepatocyte growth factor (HGF) and transforming growth factor-ß(1) (TGFß(1)) on the expression of α-smooth muscle actin (α-SMA) and collagen I in human atrial fibroblast in vitro, and to explore the possible molecular mechanism of atrial fibrosis in patients with atrial fibrillation (AF). METHODS: Human atrial fibroblast, isolated from aseptic right atrial appendage tissues of 10 sinus rhythm (SR) and 10 chronic atrial fibrillation (CAF) patients, were cultured with HGF and TGFß(1). mRNA expressions of collagen I and α-SMA were detected by semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR), the protein expression of α-SMA was determined by immunofluorescence and Western blot. RESULTS: (1) Compared with SR group, left atrium was significantly dilated in CAF group (t = 2.692, P < 0.05), the mRNA expression of collagen I and α-SMA of atrial fibroblasts were significantly upregulated (all P < 0.01), mRNA expression of collagen I was positively correlated with left atrial dimension (LAD) (r = 0.836, P = 0.014), AF duration (r = 0.739, P = 0.045) and α-SMA mRNA level (r = 0.886, P = 0.012). (2) Compared with SR group, the expression of α-SMA protein in CAF atrial fibroblasts were significantly increased (P < 0.01). (3) TGFß(1) further stimulated while HGF significantly attenuated the expression of collagen I and α-SMA in CAF atrial fibroblasts (all P < 0.01). CONCLUSIONS: Increasing expression of collagen I and α-SMA in human atrial fibroblasts might promote atria remodeling leading to the development and sustaining of AF. HGF is involved in the negative regulation on the expression of α-SMA and collagen I.

The C5a/C5aR1 Axis Contributes to the Pathogenesis of Acute Cystitis Through Enhancement of Adhesion and Colonization of Uropathogenic E. coli.

Our previous work using a murine model of pyelonephritis demonstrated that the C5a/C5aR1 axis plays a pathogenic role in acute kidney infection. In this study, we report that the C5a/C5aR1 axis also plays a pathogenic role in acute bladder infection. C5aR1-deficient mice had reduced bladder bacterial load and attenuated bladder tissue injury, which is associated with reduced expression of terminal α-mannosyl residues (Man) (a potential ligand for type 1 fimbriae of E. coli) at the luminal surface of the bladder epithelium and reduced early bacterial colonization of the bladder. In vitro, C5a stimulation enhanced mannose expression in and facilitated bacterial adhesion/colonization to human bladder epithelial cells. C5a stimulation also upregulated the activation of ERK1/2 and NF-κB signaling and gene expression of proinflammatory cytokines (i.e., Il6, Il1b, Cxcl1, Ccl2) in the epithelial cells, which could drive pro-inflammatory responses leading to tissue injury. Administration of the C5aR1 antagonist effectively reduced bladder bacterial load and tissue injury. Thus, our findings demonstrate a previously unknown pathogenic role for the C5a/C5aR1 axis in bladder infection and suggest that the C5a/C5aR1 axis-mediated upregulation of Man expression, enhancement of bacterial adhesion/colonization, and excessive inflammatory responses contribute to acute bladder infection. These findings improve our understanding of the pathogenesis of bladder infection with therapeutic implications for UTI.

[Relationship between atrial fibroblast proliferation/fibrosis and atrial fibrillation in patients with rheumatic heart disease].

OBJECTIVE: To investigate the association between gene expressions of basic fibroblast growth factor (bFGF), smooth muscle alpha-actin (alpha-SMA) and proliferating cell nuclear antigen (PCNA) and atrial fibrosis in patients with atrial fibrillation (AF). METHODS: The right atrial tissue samples were taken from 75 patients with rheumatic heart disease underwent heart valve replacement surgery (34 patients with sinus rhythm, 11 patients with paroxysmal AF and 30 patients with persistent AF) and stained with picrosirius red for quantitative analysis of collagen accumulation. The mRNA and protein levels of bFGF, alpha-SMA and PCNA were detected by semi-quantitative reverse transcription polymerase chain reaction (RT-PCR) and immunohistochemical technique, respectively. RESULTS: The percent volume fraction of collagen (CVF) was the highest in persistent AF group and the lowest in the sinus rhythm group (all P < 0.01). CVF significantly correlated with AF duration (r = 0.390, P = 0.010) and left atria (LA) dimension (r = 0.320, P = 0.005). The mRNA and protein levels of bFGF, alpha-SMA and PCNA were significantly higher in the persistent AF group than those in the paroxysmal AF group (all P < 0.05) and significantly higher in both AF groups than those in the sinus rhythm group (P < 0.05-0.01). The mRNA and protein levels of bFGF were positively correlated with CVF (r = 0.330, P = 0.004 and r = 0.292, P = 0.013, respectively), AF duration (r = 0.330, P = 0.005 and r = 0.299, P = 0.010, respectively) and left atrial dimension (r = 0.342, P = 0.003 and r = 0.285, P = 0.015, respectively). CONCLUSION: The increased gene expressions of bFGF, alpha-SMA and PCNA in atrium during AF may contribute to atrial fibrosis by promoting fibroblast proliferation in AF patients.

An intron loss of Dfak gene in species of the Drosophila melanogaster subgroup and phylogenetic analysis.

Drosophila focal adhesion kinase (Dfak) gene is a single-copy nuclear gene. Previous study revealed that Drosophila melanogaster and Drosophila simulans had lost an intron precisely within the tyrosine kinase (TyK) domain of this gene. However, this did not happen in several other Drosophila species, including Drosophila elegans, Drosophila ficusphila, Drosophila biarmipes, Drosophila jambulina, Drosophila prostipennis, Drosophila takahashii, and Drosophila pseudoobscura. In the current study, homologous sequences of Drosophila sechellia, Drosophila mauritiana, Drosophila yakuba, Drosophila teissieri, Drosophila santomea, and Drosophila erecta were amplified by polymerase chain reaction, and further sequencing analysis indicated that these species were missing a TyK domain intron, indicating they were closely related. The relationship of the D. melanogaster species group was reconstructed using TyK domain nucleotide sequences. The resulting phylogenetic tree revealed that these 8 species were the most related species in the melanogaster group. These results strongly support previously proposed classifications based on morphological and molecular data.

Ultra-trace determination of S-nitrosothiols in blood samples by spectrofluorimetry with 8-(3',4'-diaminophenyl)-difluoroboradiaza-s-indacene.

Increasing evidence suggests that S-nitrosothiols (RSNO) may represent naturally occurring nitric oxide (NO) surrogates and function as intermediates in NO metabolism. In this work, a simple, sensitive, and selective micromethod is developed and validated for quantification of RSNO. A fluorescent probe 8-(3',4'-diaminophenyl)-difluoroboradiaza-s-indacence (DABODIPY) is firstly used to label RSNO. The derivatization reaction is performed in aqueous medium at 30 degrees C for 15min in the presence of 6x10(-5)molL(-1)Hg2+ and the derivative is detected by fluorescence at lambda(ex)/lambda(em)=500/510nm. A linear function of concentration in the range of (2.0-600.0)x10(-8)molL(-1) is observed with a correlation coefficient of 0.9992 and detection limit of 1.2x10(-9)molL(-1) (S/N=3). This technique has been successfully applied to quantify RSNO in some human blood samples including healthy persons and patients suffering from cardiovascular diseases.

[Collagen type I and interleukin-1 beta gene expression in human atria during atrial fibrillation].

OBJECTIVE: To determine whether gene expression of collagen type I and interleukin-1 beta (IL-1beta) is altered in patients with atrial fibrillation (AF). METHODS: Right atrial tissue samples were taken from 75 patients with rheumatic heart disease who underwent heart valve replacement surgery. 34 patients had no history of AF, 11 patients had paroxysmal AF and 30 patients had persistent AF. The mRNA content of collagen type I and IL-1beta was measured with semi-quantitative RT-PCR. RESULTS: The mRNA content of collagen type I was significantly increased in the persistent AF group (P < 0.001) and increased in the paroxysmal AF group (P < 0.05) as compared with that in the sinus rhythm group. The mRNA content of IL-1beta was up-regulated in the persistent AF group (P < 0.05), but the trend of increase did not reach statistical significance in the paroxysmal AF group (P > 0.05). The mRNA content of IL-1beta was significantly correlated with the mRNA content of collagen type I (r = 0.295, P = 0.011), left atrial dimension (r = 0.385, P = 0.001) and AF duration (r = 0.326, P = 0.004). CONCLUSION: The upregulation of IL-1beta gene expression in atrium may contribute to the atrial fibrosis during AF through influencing collagen metabolism.

[Effects of tetramethylpyrazine on fibrosis of atrial tissue and atrial fibrillation in a canine model of congestive heart failure induced by ventricular tachypacing].

OBJECTIVE: To explore the effects of tetramethylpyrazine (TMP) on fibrosis of atrial tissue and atrial fibrillation in a canine model of congestive heart failure (CHF) induced by ventricular tachypacing. METHODS: Twenty-one healthy mongrel dogs were randomly divided into three groups, which were normal control group, untreated group and TMP-treated group. Atrial fibrillation (AF) was induced by burst of atrial pacing, after the canine model of CHF was established. The atrial tissues were sampled and stained with Mallory's trichromic stains, then the fibrosis in the atrial tissues was analyzed. The left ventricular ejection fraction (LVEF) was evaluated by echocardiography. The levels of angiotensin II (AngII), aldosterone (ALD), amino-terminal peptide of type III procollagen (PIIINP)ìlaminin (LN) and hyaluronic acid (HA) in peripheral blood were examined by radioimmunoassay. RESULTS: The LVEF was significantly decreased in the untreated group as compared with that in the normal control group (P<0.01), while the frequencies of AF and sustaining AF were significantly increased and the AF duration was obviously prolonged in the untreated group as compared with those in the normal control group (P<0.01). The fibrosis degree in the left or right atrial tissue in the untreated group was more serious than that in the normal control group (P<0.01). The AF duration was positively correlated with the fibrosis degree in the left atrial tissue (r=0.84, P=0.018). The levels of AngII, ALD, PIIINP and HA in peripheral blood were significantly higher in the untreated group than those in the normal control group (P<0.05 or P<0.01). The level of AngII was positively correlated with the level of ALD in peripheral blood (r=0.759, P=0.048). The LVEF and the frequency of sustaining AF were both significantly improved in the TMP-treated group as compared with those in the untreated group (P<0.05). The fibrosis in the left or right atrial tissue in the untreated group was more serious than that in the untreated group (P<0.01). The levels of AngII and PIIINP in peripheral blood were also markedly higher in the TMP-treated group than those in the untreated group (P=0.05, P=0.01). CONCLUSION: Tetramethylpyrazine has the effect of reducing the fibrosis degree of atrial tissue in dogs with CHF, and this efficacy may be related to the mechanism of decreasing the frequency of AF and shortening the AF duration.

[Changes in gelatinases expression and activity in human atria during atrial fibrillation].

OBJECTIVE: To determine whether expression and activity of atrial gelatinases are altered in patients with atrial fibrillation (AF). METHODS: The right atrial tissue samples were taken from 75 patients with rheumatic heart disease who underwent heart valve replacement surgery. 34 patients were in sinus rhythm, 11 patients had paroxysmal AF and 30 patients had persistent AF. The mRNA and protein level of MMP-2, MMP-9, TIMP-1, TIMP-2 were measured by semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR) and Western-blotting analysis respectively. The activity of MMP-2 and MMP-9 was measured by zymographic analysis. RESULTS: (1) The mRNA level of MMP-2 increased significantly in the persistent AF group followed by the paroxysmal AF group compared with the sinus rhythm group (P < 0.01, respectively). MMP-9 mRNA expression remained compatible within groups (P > 0.05). MMP-2 and MMP-9 protein expression was prominent in the persistent AF group compared with the sinus rhythm and paroxysmal AF groups (P < 0.01), the significant difference was also observed between the paroxysmal AF and sinus groups (P < 0.05). (2) TIMP-1 and TIMP-2 expression at mRNA and protein level were all down-regulated in the persistent AF group compared with the sinus rhythm group (P < 0.05), however, the trends of reduction did not reach statistical significance in the paroxysmal AF group (P > 0.05) except that of the mRNA level of TIMP-2 (P < 0.05). (3) The activity of MMP-2 and MMP-9 significantly increased in both paroxysmal AF and persistent AF groups compared with the sinus rhythm group (P < 0.05). The significant difference in MMP-9 was also observed between the persistent AF and paroxysmal AF groups (P < 0.01). (4) MMP-2 and MMP-9 expression at mRNA and protein level were positively correlated with left atrial dimension and AF duration (P < 0.05) and were negatively correlated with the mRNA and protein level of TIMP-2 and TIMP-1 respectively (P < 0.01). CONCLUSIONS: The upregulation of MMP-2,9 gene expression and activity, along with the selective downregulation of TIMP-1,2 may have contributed to the atrial structural remodeling during AF through influencing collagen metabolism.

[Clinical on molecular basis of atrial fibrosis in patients with atrial fibrillation investigation].

OBJECTIVE: To determine the molecular mechanisms involved in atrial fibrosis which occurs in patients with atrial fibrillation (AF) and to investigate their effects on the initiation and maintenance of AF. METHODS: The right atrial tissue samples were taken from 73 patients with rheumatic heart disease who underwent heart valve replacement surgery. 34 patients had no history of AF (sinus rhythm group), 9 patients had paroxysmal AF and 30 patients had persistent AF. The mRNA content of collagen type I, collagen type III, MMP-2, TIMP-1, TIMP-2, TIMP-3 and TIMP-4 was measured by semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR) and normalized to beta-actin or GAPDH. RESULTS: Compared to sinus rhythm group, the mRNA of collagen type I and MMP-2 increased significantly in the persistent AF group (all, P < 0.01), followed by the paroxysmal AF group (all, P < 0.05). The mRNA of collagen type III was slightly higher in both AF groups than in the sinus rhythm group, but the differences were not statistically significant (P > 0.05). The mRNA of TIMP-1, TIMP-2 and TIMP-3 was down-regulated in the persistent AF group (all, P < 0.01, respectively), however, the trends of reduction did not reach statistical significance in the paroxysmal AF group (P > 0.05). The mRNA of TIMP-4 remained compatible in each group. The mRNA of collagen type I was significantly correlated with left atrial dimension (r = 0.336, P = 0.004) and AF duration (r = 0.339, P = 0.003). The mRNA of MMP-2 was significantly correlated with the mRNA of TIMP-2 (r = -0.326, P = 0.006), the mRNA of collagen type I (r = 0.322, P = 0.006), left atrial dimension (r = 0.300, P = 0.011) and AF duration (r = 0.300, P = 0.010). CONCLUSION: The increased level of collagen type I associated with selective downregulation of TIMP-2 and upregulation of MMP-2 gene expression in atrium could be one of the molecular mechanisms of atrial fibrosis during atrial fibrillation, which correlates with the initiation and maintenance of AF.

Genetic differentiation of different populations of four locust species in China.

The genetic structure of eight populations of four locust species in three families (Catantopidae, Pamphagidae and Pyrgomorphidae) was examined by means of horizontal starch gel electrophoresis, the locusts were collected from Shanxi, Jiangsu and Hebei Province in China. The allele frequency and allozyme polymorphism of 12 enzymes (18 loci) were analyzed. Low variability with a few alleles was observed in Shirakiacris shirakii and Atractomorpha sinensis, however, Oxya Chinensis and Haplotropis brunneriana showed high polymorphism. In the eight populations of the four locust species, high percentage of polymorphic loci was observed (53.3%-100.0%). Owing to heterozygote deficiency of some species, all the four species demonstrated low overall heterozygosity (Ho = 0.034-0.139), which lead to the genotype frequency deviated significantly from Hardy-Weinberg equilibrium. However, many of the examined loci were fit for or close to H-W equilibrium in Atractomorpha sinensis. A certain differentiation in mean heterozygosity was found among the four species. Due to the difference of migratory capability, reproductive manner and living bound, mean heterozygosity (Ho) is higher in Haplotropis brunneriana (Ho = 0.089-0.139) and Oxya Chinensis (Ho = 0.073-0.090) than in Atractomorpha sinensis (Ho = 0.034-0.050) and Shirakiacris shirakii (Ho = 0.048-0.068). Genetic differentiation from F-statistics was also different at population level among four locust species. It was high in Haplotropis brunneriana (Fst = 0.32) and Atractomorpha sinensis (Fst = 0.31), and low genetic differentiations were observed in Oxya Chinensis (Fst = 0.20) and Shirakiacris shirakii (Fst = 0.18). It was confirmed that migratory capability, adaptability and surrounding factors had an important influence on the genetic differentiation of locust populations. The taxon relationships based on Nei's genetic identity (I) and genetic distance (D) were consistent with results obtained from karyotypic analyses. Oxya Chinensis and Shirakiacris shirakii in the same family have a higher genetic identity (I = 0.576) and a smaller genetic distance (D = 0.559); the species examined in Pampagidae displayed somewhat closer relationship to those in Pyrgomorphidae (D = 0.776, I = 0.464) than to those in Catantopidae (D = 0.908, I = 0.406). It is suggested that the allozyme analysis is an useful molecular marker for phylogenetic reconstruction of locust at both species and population levels.

Comparative allozyme analysis of Oriental migratory locust Locusta migratoria manilensis from two breeding areas in north China.

The allozyme analysis using horizontal starch gel electrophoresis was employed to compare the genetic structure in the population of oriental migratory locust Locusta migratoria manilensis from two breeding areas, Beidagang(Tianjin) and Huanghua(Hebei). The two areas are adjacent but with distinct ecological features, with the recorded locust outbreaks and migration. The zymograms showed that among nineteen loci four (Mdh-1, Pgm, Adk and G3pd) showed extremely low variability level with the frequency of the most common allele higher than 0.95 in the populations from both sites. The rest loci had 2 to 4 alleles but the allele frequencies between the two populations were all similar except Fbp and Got-2 loci. In the 27 chi 2-tests for the genotypes at polymorphic loci only two (Pgi and Got-1) of beidagang population did fit the Hardy-Weinberg's expectations. This is due to high frequencies of the most common homozygotes and the corresponding heterozygote deficiency. The allozyme data demonstrated that the locusts had remarkable genetic variability within each population, but little divergence between the populations. The genetic variability measurements were found similar: Percentage of polymorphic loci (P) was between 73.7% and 78.9%; The mean number of alleles per locus (A) was from 2.9 to 3.1; and the mean heterozygosity (Ho) was nearly identical (about 0.138). The F-statistics (FST = 0.053) also showed the genetic uniformity of the populations, corresponding to the high Nei's genetic identity (I = 0.938). These results of the allozyme analysis suggested that the two populations appeared to be a part of a large population. It is reasoned that the genetic polymorphism and differentiation at certain loci between the two populations may depend on at least two agnostic factors that are all related to migration. First, the unusual dispersal capability of L. m. manilensis tends to make a continuous genetic structure distribution. Second, the frequent migration also results in the individuals to be exposed to drastically various environments. Since the broad adaptability is crucial to survive the changing environments, the genetic variation at population level is necessarily required to offer the population resilience for successful survival and reproduction under those ecologically divergent abiotic and biotic conditions. Thus, the migration contributes to the maintenance of dynamic equilibrium of genetic polymorphism in this highly specialized subspecies.

Genetic studies on eight populations of eight locust species from Shanxi Province, China.

The genetic structure of eight locust species in three families (Catantopidae, Oedipodidae and Arcypteridae) from Shanxi Province in China was compared using allozyme analysis with horizontal starch gel electrophoresis. Among 17 loci identified in zymograms, Ao-1, Est-3, G3pd-1, Idh-2 and Mdh-2 had low variability with a few alleles. High polymorphism was observed at Ldh-1, Me-1 and Gpi-1. Each of the eight species demonstrated high percentage of polymorphic loci (P = 64.7%-94.1%) but low observed heterozygosity (H0 = 0.024-0.087) due to heterozygote deficiency. It was noted that the migratory locusts usually had higher percentage of polymorphic loci (P = 88.2%-94.1%) than non-migratory species (P = 64.7%-94.1%). The only exception is Oxya chinensis(P = 94.1%). It is reasoned that the higher polymorphism is necessary for migratory species to cope with the environments that might be drastically different from the habitats before migration. The taxon relationships using cluster analysis based on Nei's genetic identity (I) and Roger's genetic distance (D) were the same at species and genus levels. The differences were found at family level, possibly due to the alternative algorithms. The cladogram using Roger's genetic distance (D) overlapped the relationship obtained from karyotypic analyses, which demonstrated that the species examined in Catantopidae displayed somewhat closer relationship to those in Oedipodidae than to those in Arcypteridae. It is suggested that the allozyme analysis is useful as molecular marker for locusts in phylogenetic reconstruction at the species and genus level, while additional data from other studies are necessary when used for higher taxa.

Comparative allozyme analysis of two geographic populations of Pararcyptera microptera meridionalis in China.

The genetic structure of the two populations of Pararcyptera microptera meridionalis (Ikonn.) from Hebei and Liaoning in China was analyzed with horizontal starch gel electrophoresis. Among 15 loci of 11 enzymes identified in zymograms, Adk-1, Fbp-1, Mdh-2 and G3pd-1 showed low variability with few alleles. Higher allelic polymorphisms were observed at Fbp-2, Mdh-1 and Me-1. The two populations demonstrated high percentage of polymorphic loci (93.3% and 100.0%) but low observable overall heterozygosity (0.061 and 0.086), that could be attributed to heterozygote deficiencies, which led to the genotype frequency deviating from Hardy-Weinberg expectations. It is reasoned that the strong movement capability of the insect makes the individuals likely to be exposed to drastically varied environments, which tends to maintain dynamic equilibrium of genetic polymorphisms. The F-statistics between the two populations was comparatively smaller ( F(st) = 0.084), but larger when compared with those in migratory locusts like Locusta migratoria manilensis. Nei's genetic identity (I) and Roger's genetic distance (D) also showed close genetic relationship of the two populations by their high genetic identity (I = 0.904) and small genetic distance (D = 0.256). However,considerably qualitative and quantitative differences were noted at loci Acp-1 (F(st) = 0.462) and Pgi-1 (F(st) = 0.182).

Electrochemical behavior and voltammetric determination of tryptophan based on 4-aminobenzoic acid polymer film modified glassy carbon electrode.

Herein, a novel electrochemical method was developed for the determination of tryptophan based on the poly(4-aminobenzoic acid) film modified glassy carbon electrode (GCE). The electrochemical behaviors of tryptophan at the modified electrode were investigated. It was found that the oxidation peak current of tryptophan at the modified GCE was greatly improved compared with that at the bare GCE. The effects of supporting electrolyte, pH value, scan rate, accumulation potential and time were examined. The oxidation peak current of tryptophan was proportional to its concentration over the range from 1.0 x 10(-6) to 1.0 x 10(-4)mol L(-1). The limit of detection was evaluated to be 2.0 x 10(-7)mol L(-1). The proposed method was sensitive and simple. It was successfully employed to determine tryptophan in pharmaceutical samples.