Characterization of the complete sequences and stability of plasmids carrying the genes aac(6')-Ib-cr or qnrS in Shigella flexneri in the Hangzhou area of China.

The aim of this study was to explore the fluoroquinolone resistance mechanism of aac (6')-Ib-cr and qnrS gene by comparing complete sequences and stability of the aac(6')-Ib-cr- and qnrS-positive plasmids from Shigella isolates in the Hangzhou area of China. The complete sequences of four newly acquired plasmids carrying aac(6')-Ib-cr or qnrS were compared with those of two plasmids obtained previously and two similar reference Escherichia coli plasmids. The results showed that the length, antibiotic resistance genes and genetic environment were different among the plasmids. Moreover, the plasmid stability of three wild-type isolates and five plasmid transformants carrying aac(6')-Ib-cr and/or qnrS was measured in vitro, and all eight isolates were found to have lost their aac(6')-Ib-cr- or qnrS-positive plasmids to a different extent at different stages. When the plasmids were electroporated into Shigella flexneri or they lost positive plasmids, the MICs of ciprofloxacin increased or decreased two- to eightfold for aac(6')-Ib-cr-positive plasmids and 16- to 32-fold for qnrS-positive plasmids. To our knowledge, this is the first report comparing the complete sequences and describing stability for the aac(6')-Ib-cr- and qnrS-positive plasmids from Shigella isolates.

Identification and functional analysis of two toxin-antitoxin systems in Campylobacter jejuni.

Toxin-antitoxin (TA) systems are widely distributed in bacteria and play an important role in maintaining plasmid stability. The leading foodborne pathogen, Campylobacter jejuni, can carry multiple plasmids associated with antibiotic resistance or virulence. Previously a virulence plasmid named pVir was identified in C. jejuni 81-176 and IA3902, but determining the role of pVir in pathogenesis has been hampered because the plasmid cannot be cured. In this study, we report the identification of two TA systems that are located on the pVir plasmid in 81-176 and IA3902, respectively. The virA (proteic antitoxin)/virT (proteic toxin) pair in IA3902 belongs to a Type II TA system, while the cjrA (RNA antitoxin)/cjpT (proteic toxin) pair in 81-176 belongs to a Type I TA system. Notably, cjrA (antitoxin) represents the first noncoding small RNA demonstrated to play a functional role in Campylobacter physiology to date. By inactivating the TA systems, pVir was readily cured from Campylobacter, indicating their functionality in Campylobacter. Using pVir-cured IA3902, we demonstrated that pVir is not required for abortion induction in the guinea pig model. These findings establish the key role of the TA systems in maintaining plasmid stability and provide a means to evaluate the function of pVir in Campylobacter pathobiology.

Epidemiologic characterization of 30 confirmed cases of human infection with avian influenza A(H7N9) virus in Hangzhou, China.

BACKGROUND: We examined the clinical and epidemiological characteristics of 30 cases of human infection with avian influenza A(H7N9) virus in Hangzhou and investigated their external environments to provide evidence for contact tracing and disease prevention and control. METHODS: The cases confirmed from April 1 through May 1, 2013 were studied. Field epidemiologic surveys were conducted to collect the clinical and epidemiologic data. Case-related and environmental specimens were collected for etiologic detection. RESULTS: Thirty cases of human infection with avian influenza A(H7N9) virus were confirmed in Hangzhou from April 1 through May 1, 2013, including one pregnant woman and three deaths. The median age of the patients was 62 years (range: 38-86 years). Twenty-three of the patients were men (76.67%). The median duration between disease onset and occurrence of respiratory failure and confirmed diagnosis was 5 and 6 days, respectively. Maximum medical observation of 666 close contacts of the patients revealed no irregularity. Of 314 external environmental specimens, the overall positive detection rate of H7N9 nucleic acid was 28.98%. Eight districts of Hangzhou city had positive detections in the external environments, the highest rate being in Yuhang District (78.13%). Statistical analysis of the specimen collection locations indicates a significant difference between the case-linked locations and the non-case locations (χ2 = 16.563, p < 0.05) in terms of H7N9 viral nucleic acid detection rate. No epidemiologic link has been found among the 30 cases. CONCLUSIONS: Most of the infected were retired individuals aged 60 years or older. Men made the majority. The cases are sporadic at present, with no evidence of human-to-human transmission. Exposures to poultry and live poultry markets may be important sources of infection.

Association of common SNP rs1136410 in PARP1 gene with the susceptibility to male infertility with oligospermia.

PURPOSE: This study aims to explore possible associations between polymorphisms of common SNP rs1136410 and rS1805405 in PARP1 gene and male infertility with spermatogenesis impairment. METHODS: The polymorphic distributions of SNP rs1136410 and rS1805405 were investigated by polymerase chain reaction and restriction fragment length polymorphism analysis in a Chinese cohort including 371 infertile patients with idiopathic azoospermia or oligospermia and 231 controls. RESULTS: Significant differences in the frequencies of allele and genotype of SNP rs1136410 were observed between patients with oligospermia and controls. The allele C (46.3 % vs. 36.4 %, P = 0.003) and genotype CC (22.6 % vs. 13.4 %, P = 0.014) significantly increased, whereas genotype TT (30 % vs. 40.7 %, P = 0.021) significantly decreased in patients with oligospermia compared with controls at this SNP locus. CONCLUSIONS: These results indicated that genotype CC of SNP rs1136410 may increase the risk of oligosoermia and genotype TT of rs1136410 may have some protective effect from oligospermia, suggesting that the polymorphism of SNP rs1136410 in PARP1 gene may modify the susceptibility to male infertility with oligospermia.

Genetic variants of eNOS gene may modify the susceptibility to idiopathic male infertility.

In testis, eNOS is responsible for synthesis of nitric oxide (NO) which is an essential gas message regulator in spermatogenesis, suggesting that eNOS gene plays a role in normal spermatogenesis and the genetic variants of eNOS gene may be potential genetic risk factors of spermatogenesis impairment. In this study, the polymorphic distributions of three common polymorphism loci including T-786C, 4A4B and G894T in eNOS gene were investigated in 355 Chinese infertile patients with azoospermia or oligozoospermia and 246 healthy fertile men and a meta-analysis was carried in order to explore the possible relationship between the three loci of eNOS gene and male infertility with spermatogenesis impairment. As a result, allele -786C of T-786C (11.4% versus 6.5%, p = 0.004) and 4A of 4A4B (11.0% versus 6.3%, p = 0.005) as well as genotype TC of T-786C (22.8% versus 13.0%, p = 0.002) and AB of 4A4B (18% versus 11%, p = 0.015) were significantly associated with idiopathic male infertility. The haplotypes T-4A-G (7.4% versus 4.1%, p = 0.015) and C-4B-G (7.6% versus 4.4%, p = 0.028) could increase the susceptibility to male infertility, whereas haplotype T-4B-G (67.0% versus 75.2%, p = 0.002) might be a protective factor for male infertility. The results of meta-analysis revealed that the polymorphism of T-786C was associated with male infertility. These findings suggested that the variants of eNOS gene may modify the susceptibility to male infertility with impaired spermatogenesis.

SNPs in KIT and KITLG genes may be associated with oligospermia in Chinese population.

KIT/KITLG signaling system is crucial for spermatogenesis, which suggests that KIT and KITLG genes may be involved in spermatogenesis impairment and male infertility. To explore the possible association of KIT and KITLG genes with male infertility having spermatogenesis impairment, polymorphism distributions of SNP rs3819392 in KIT gene as well as rs995030 and rs4474514 in KITLG gene were investigated in 372 patients with idiopathic azoospermia or oligospermia and 205 fertile controls. As a result, the significant differences in polymorphism distributions of SNP rs3819392 in KIT gene and rs4474514 in KITLG gene were observed between the patients with oligospermia and controls. The frequencies of allele G (94.2% versus 90.0% p = 0.022) and genotype GG (89.2% versus 82.0% p = 0.042) in patients with oligospermia were significantly higher than those in controls at rs3819392 locus in KIT gene. In addition, the genotype CC of rs4474514 in KITLG (8.2% versus 3.4%, p = 0.034) also significantly increased in oligospermic patients in comparison to controls. These findings indicated that SNP rs3819392 in KIT gene and rs4474514 in KITLG gene may be associated with oligospermia, suggesting that polymorphism of KIT and KITLG genes may play a role in oligospermia.

Spontaneous mutation frequency and molecular mechanisms of Shigella flexneri fluoroquinolone resistance under antibiotic selective stress.

The incidence of fluoroquinolone-resistant Shigella strains has risen rapidly, presumably in response to ciprofloxacin antibiotic stress. Understanding the molecular mechanisms underlying this resistance phenotype is critical to developing novel and effective therapeutic strategies. In this study, the frequency of ciprofloxacin-induced mutation was measured in antibiotic resistance genes (gyrA, gyrB, parC, parE, marOR, and marA) of Shigella flexneri. The S. flexneri 2a strain 301 was cultured on Luria-Bertani agar plates containing one of seven different ciprofloxacin concentrations (range: 0.03125-2 µg mL(-1)). Resistant colonies were selected for gene-targeted sequencing analysis; the identified point mutations were subsequently confirmed by insertion into antibiotic cassette plasmids and growth under ciprofloxacin stress. The results demonstrated that the seven different ciprofloxacin concentrations produced dose-dependent frequencies of spontaneous mutations: 10(-8) (0.03125 and 0.0625 µg mL(-1)), 10(-9) (0.125 µg mL(-1)), and <10(-9) (0.25, 0.5, 1, 2 µg mL(-1)). PCR sequencing of the ten randomly selected resistant colonies (minimum inhibitory concentrations (MICs) of 0.125 µg mL(-1), n = 5 and 0.25 µg mL(-1), n = 5) revealed that all colonies had mutations in the gyrA gene at either codon 83 (Ser83 → Leu) or 87 (Asp87 → Tyr or → Gly), both of which were confirmed at MIC of 0.125 µg mL(-1). None of the spontaneous mutation colonies exhibited gyrB, parC, parE, marOR, or marA mutations. In conclusion, S. flexneri is normomutable under ciprofloxacin antibiotic stress and fluoroquinolone resistance by spontaneous mutation occurs at a low rate. Codon mutations gyrA 83 and/or gyrA 87 cause a 4-fold increase in the ciprofloxacin MIC, and may represent the natural mechanism of fluoroquinolone resistance.

Association between single-nucleotide polymorphisms of DNMT3L and infertility with azoospermia in Chinese men.

The gene for DNA methyltransferase 3-like protein (DNMT3L) is essential for normal spermatogenesis and may be involved with spermatogenetic impairment and male infertility. To explore the possible association between the DNMT3L gene and male infertility, this study investigated allele, genotype and haplotype frequencies of three single nucleotide polymorphism (SNP) loci, rs2070565, rs2276248 and rs7354779, of DNMT3L in 233 infertile patients with azoospermia and 249 fertile controls from a population of Chinese men using polymerase chain reaction/restriction fragment length polymorphism. Results showed that the frequencies of allele A (20.6% versus 14.9%; P = 0.022) and the allele A carrier (GA + AA; 37.8% versus 28.1%; P = 0.027) in azoospermic patients were significantly higher than those in controls at the rs2070565 locus. The haplotype AAA frequency was significantly higher (18.1% versus 12.4%; P = 0.02) while the haplotype GAA frequency was significantly lower (53.2% versus 62.1%; P = 0.007) in infertile patients compared with fertile controls. These results indicated that SNP rs2070565, as well as haplotypes AAA and GAA, may be associated with male infertility and suggest that DNMT3L may contribute to azoospermia susceptibility in humans.

Salicylate functions as an efflux pump inducer and promotes the emergence of fluoroquinolone-resistant Campylobacter jejuni mutants.

Salicylate, a nonsteroidal anti-inflammatory compound, has been shown to increase the resistance of Campylobacter to antimicrobials. However, the molecular mechanism underlying salicylate-induced resistance has not yet been established. In this study, we determined how salicylate increases antibiotic resistance and evaluated its impact on the development of fluoroquinolone-resistant Campylobacter mutants. Transcriptional fusion assays, real-time quantitative reverse transcription-PCR (RT-PCR), and immunoblotting assays consistently demonstrated the induction of the CmeABC multidrug efflux pump by salicylate. Electrophoretic mobility shift assays further showed that salicylate inhibits the binding of CmeR (a transcriptional repressor of the TetR family) to the promoter DNA of cmeABC, suggesting that salicylate inhibits the function of CmeR. The presence of salicylate in the culture medium not only decreased the susceptibility of Campylobacter to ciprofloxacin but also resulted in an approximately 70-fold increase in the observed frequency of emergence of fluoroquinolone-resistant mutants under selection with ciprofloxacin. Together, these results indicate that in Campylobacter, salicylate inhibits the binding of CmeR to the promoter DNA and induces expression of cmeABC, resulting in decreased susceptibility to antibiotics and in increased emergence of fluoroquinolone-resistant mutants under selection pressure.

NILs of Cold Tolerant Japonica Cultivar Exhibited New QTLs for Mineral Elements in Rice.

Chilling stress at booting stage can cause floret deterioration and sterility by limiting the supply of food chain and the accumulation of essential mineral elements resulting in reduction of yield and grain quality attributes in rice. Genomic selection of chilling tolerant rice with reference to the accumulation of mineral elements will have great potential to cope with malnutrition and food security in times of climate change. Therefore, a study was conducted to explore the genomic determinants of cold tolerance and mineral elements content in near-isogenic lines (NILs) of japonica rice subjected to chilling stress at flowering stage. Detailed morphological analysis followed by quantitative analysis of 17 mineral elements revealed that the content of phosphorus (P, 3,253 mg/kg) and potassium (K, 2,485 mg/kg) were highest while strontium (Sr, 0.26 mg/kg) and boron (B, 0.34 mg/kg) were lowest among the mineral elements. The correlation analysis revealed extremely positive correlation of phosphorus (P) and copper (Cu) with most of the cold tolerance traits. Among all the effective ear and the second leaf length correlation was significant with half of the mineral elements. As a result of comparative analysis, some QTLs (qBRCC-1, qBRCIC-2, qBRZC-6, qBRCHC-6, qBRMC-6, qBRCIC-6a, qBRCIC-6b, qBRCHC-6, and qBRMC-6) identified for calcium (Ca), zinc (Zn), chromium (Cr) and magnesium (Mg) on chromosome number 1, 2, and 6 while, a novel QTL (qBCPC-1) was identified on chromosome number 1 for P element only. These findings provided bases for the identification of candidate genes involved in mineral accumulation and cold tolerance in rice at booting stage.

[The zonal characteristics and cultivated types difference of functional components in brown rice for core collection of Yunnan rice].

The zonal characteristics and cultivated types difference of functional components in brown rice of 907 accessions for primary core collection for rice landrace from 16 prefectures of five rice regions in Yunnan Province were determined by ultraviolet spectrophotometry of DU640 type from BECKMAN. The analytical results showed that the contents mg x (100 g)(-1) of total flavone in brown rice is 306.98 +/- 192.75, the content of upland (341.74 +/- 185.11) is the most significantly higher than that of lowland (290.41 +/- 193.72), that of the glutinous (315.54 +/- 197.64) is significantly higher than non-glutinous (171.68 +/- 11.76), that of early-mid (318.25 +/- 197.93) is the most significantly higher than late (282.12 +/- 178.11), that of red rice (379.22 +/- 197.70) and purple rice (365.61 +/- 195.44) are the most significantly higher than white rice (216.96 +/- 142.11), and that of nuda (332.68 +/- 196.22) is significantly higher than non-nuda (300. 48 +/- 191.14). Among five rice regions, in turn III (327.13) > LL (324.23) > IV (273.11) > V (270.16) > I (258.26), namely it is the most significantly highest (p < 0.01) for contents of total flavone from South Yunnan (II, III) than that of middle Yunnan; It is the most significantly highest from Simao prefecture than that of 8 prefectures, but it is the most significantly low from Baoshan prefecture than that of 7 prefectures. The contents mg x (100 g)(-1) of g-amino butyric acid (GABA) in brown rice is 7.43 +/- 2.53, the content of lowland (7.59 +/- 2. 56) is the most significantly higher than that of upland (7.09 +/- 2.45), that of non-glutinous (8.55 +/- 2.88) is the most significantly higher than glutinous (7.10 +/- 2.32), that of late (7.88 +/- 2.64) is the most significantly higher than early-mid (7.23 +/- 2.45), that of white rice (8.38 +/- 2.66) is the most significantly higher than red rice (6.63 +/- 2.14) and purple rice (7.34 +/- 2.18); Among five rice regions in turn II (7.69) > I (7.40) > IV (7.39) > III (7.33) > V (6.64), viz. the content of GABA from South Yunnan single/double cropping rice region is clearly higher than that of Northwest Yunnan cold highland japonica rice region; It is significant difference of GABA content in brown rice for that Simao prefecture South Yunnan and Yuxi as well as Baoshan prefecture, at least than that of 5 prefectures. The contents (%) of resistant starch in brown rice is 0.75 +/- 0.29, the contents of indica (0.78 +/- 0.35) is significantly higher than japonica (0.74 +/- 0.24), that of glutinous (0.78 +/- 0.31) is the most significantly higher than non-glutinous (0. 67 +/- 0.22), that of late (0.77 +/- 0. 35) is the most significantly higher than early-mid (0.75 +/- 0.26), that of red rice (0.81 +/- 0.30) and purple rice (0.70 +/- 0.30) is the most significantly higher than white rice (0.69 +/- 0.27); Among five rice regions in turn I (0.83) > II (0.79) > III (0.76) > V (0. 55) > IV (0.50), namely it is the highest (p < 0.01) for contents of resistant starch from the middle and South Yunnan province (I, II, III) than that of northern rice regions (V, IV), and the lowest for contents of resistant starch of Lijiang prefecture of northwestern and Zaotong of northeastern in this province compared to that of 13 prefectures except Diqing prefectures. These results not only reveal the most significant difference between lowland versus upland, glutinous versus nonglutinous, early/mid rice and red/purple versus white rice based on the contents of total flavone and resistant starch as well as g-amino butyric acid (GABA) in brown rice (p < 0.01), and no significant difference between indica and japonica, awn and no-awn, and common rice and aromatic/soft rice, but also elucidate obvious zonal characteristics of three functional components in brown rice of Yunnan rice. The above results provided theory bases for the genetic breeding and production of functional rice and solve the problems of chronic for human being.

Characterization of fluoroquinolone-resistant Shigella flexneri in Hangzhou area of China.

OBJECTIVES: The aim of this study was to characterize fluoroquinolone-resistant Shigella and determine whether the qnr and aac(6')-Ib-cr genes could contribute to sporadic shigellosis at the clinic in the Hangzhou area of China. METHODS: A total of 202 strains of Shigella (79 Shigella sonnei and 123 Shigella flexneri ) isolated from sporadic cases of shigellosis from 1998 to 2007 were analysed for their antimicrobial susceptibility. The gyrA, gyrB, parC, parE, qnr and aac(6')-Ib-cr genes and the profiles and incompatibility of plasmids were characterized. Chromosomal DNA fingerprinting was determined by XbaI-based digestion and PFGE. RESULTS: All strains of S. sonnei were susceptible to fluoroquinolones (ciprofloxacin and levofloxacin) while 15 out of 123 strains of S. flexneri were resistant. All of the 15 resistant strains displayed common mutations in the gyrA and parC genes and formed eight distinct groups with unique molecular characteristics. Notably, 10 isolates showed mutations at codon 87 of gyrA, and the other 5 were qnrS-positive. Two strains were positive for the aac(6')-Ib-cr gene. Importantly, this is the first report of qnrS- and aac(6')-Ib-cr-positive Shigella in China, the qnrS-positive S. flexneri serotypes 1a, 2a and 4c and the aac(6')-Ib-cr-positive S. flexneri serotypes 2a and 4c worldwide. CONCLUSIONS: The common mutations at position 83 of gyrA and position 80 of parC were crucial for resistance to nalidixic acid in S. flexneri. The mutation at position 87 of gyrA or the presence of the qnrS gene is necessary for high-level resistance to fluoroquinolones in Shigella isolates from China.

[Multiplex PCR assay for dissemination and diversity of extended-spectrum beta-lactamase genes in Shigella isolates].

OBJECTIVE: To develop a rapid and simple multiplex polymerase chain reaction (PCR) method which discriminates extended-spectrum beta-lactamases (ESBLs) genes in sporadic Shigella isolates from 1998 to 2007 in Hangzhou city, China. METHODS: After ESBLs screening according to the Clinical and Laboratory Standards Institute (CLSI) method, CTX-M, TEM, SHV and OXA-1 encoding genes were detected by using a multiplex PCR method, and the results were verified by 8 single gene PCR amplification. RESULTS: Seventeen isolates harbored ESBLs genes among 195 Shigella isolates (8.72%). Genes encoding CTX-M (17 strains), TEM (2 strains), OXA-1 (10 strains) and SHV (0 strains) were discriminated with multiplex PCR analysis, which coincided with eight single gene PCR analysis at 94.12%. CONCLUSION: Multiplex PCR should be a suitable tool for initial rapid screening and discriminating ESBLs genes in Shigella isolates. With similar trend of national surveillance data, the proportion of sporadic Shigella isolates harbouring ESBLs genes might probably be on increase.

[The zonal characterization of elemental concentrations in brown rice of core collection for rice landrace in Yunnan Province by ICP-AES].

In the present paper, the contents of 8 elements in brown rice of 789 accessions core collection for rice landrace from 16 prefectures of five rice regions in Yunnan province were determined by ICP-AES technique. The method proves to be simple, rapid, highly sensitive and accurate, and can be used to determine many elements at the same time. Its recovery ratio obtained by standard addition method ranged from 97.1% to 110.2%, and its RSD was from 0.7% to 4.4%. The analytical results showed that the elemental concentrations (mg x kg(-1) ) in brown rice are in turn of P(3 834.83 +/- 486.49) > K(2 567.72 +/- 336.74) > Mg (2 567.72 +/- 336.74) > Ca (153.67 +/- 55.90) > Zn( 33.35 +/- 13.65) > Fe(32.08 +/- 25.51) > Cu (14.22 +/- 11.85) > Mn(13.58 +/- 3.22). The highest P content is in brown rice from the northwest Yunnan with the rich nonferrous metals and most abundance of biodiversity in the world, the highest Ca, Mg, Fe and Zn concentrations are in brown rice from the middle Yunnan with early cambrian fauna and phosphorite enrichment, and the highest Cu and Mn contents are in brown rice from the southwest Yunnan with the prominent crop diversity. The distributing zones with the highest P and K, middle Ca, Mg and Mn and lowest Fe and Zn in Yunnan are the enrichment zone of minal resources and largest biodiversity. As far as we know, this is the first report that the zonal characterstics of mineral elemental concentractions in brown rice are associated with a lot of factors, such as biodiversity center, enrichment zone of minal resources, origin of life, mountain ranges and rivers and so on, and further it was deduced that the asymmetry of distribution for mineral elements and its reciprocity of mountain ranges and rivers are the key of origin of life. The above results provided reliable data and theory bases for the malnourished Fe and Zn and Ca for 4 billion people in the world, origin of life, the genetic breeding and production of functional rice.

[Correlation of mineral elements between milled and brown rice and soils in Yunnan studied by ICP-AES].

In the present paper, the contents of 18 mineral elements in milled and brown rice of 55 accessions elite cultivars as well as corresponding soils were determined by ICP-AES technique. The method proves to be simple, rapid, highly sensitive and accurate, and can be used to determine many elements at the same time, its recovery ratio obtained by standard addition method ranged between 93.1% and 110.2%, and its RSD was from 0.8% to 5.1%. The analytical results showed that 18 mineral elements (S, Mo, Ba, Ni, Fe, Cr, Na, Al, Cu, P, Sn, Zn, B, Mn, Mg, Ca, Sr and K) are the important active compositions of functional rice, and their mean contents in milled rice are in the order of P>K>S>Mg>Ca>Zn>Na>Al>Mn>Fe>Cu>B >Mo>Ni>Sn>Cr>Ba>Sr, in brown rice in the order of P>K>Mg>S>Ca>Zn>Mn>Al>Na>Fe>Cu>B>Mo>Sn> Ni>Cr>Ba>Sr, but in soil in the order of Fe>Al>Ca>K>Mg>P>S>Mn>B>Na>Ba>Zn>Cr>Cu>Ni>Sn>Mo> Sr; 16 mineral elements in milled and brown rice (except for S and P) are clearly lower than that in soils. The correlation of 8 microelements (Mo, Ni, Cr, Sr, Mn, Zn, Cu and Na) in milled and brown rice is closer than that of 6 macroelements (P, K, Mg, Ca, S and Al). There are rich Fe, Al and Ca in Yunnan soils, but 4 elements (P, K, Mg and S) are in high priority in milled and brown rice; The milled rice used for the staple is easier to place a premium on chronics than brown rice. The above results provided reliable data and theory bases for genetic breeding and production of functional rice, and for further solving the chronics and the malnourished problems with insufficient Fe, Zn and Ca for 4 billion people in the world.

[The correlation of myxovirus resistance 1 and 2'5'-ollgoadenylate synthetase 1 with clinical features of systemic lupus erythematosus].

OBJECTIVE: To investigate the expression of myxovirus resistance 1(MX1) and 2'5'-oligoadenylate synthetase 1(OAS1) genes in the peripheral blood leukocyte of patients with systemic lupus erythematosus (SLE), and to evaluated the relations between these genes expression levels and disease activity. METHODS: In this study, there were 50 SLE petients, 20 non-SLE patients with rheumatic dieases, and 25 normal controls. The peripheral blood samples of these patients were collected, and the expression levels (indicated as delta Ct value) of MX1 and OAS1 were measured by Sybr green dye based real-time quantitative PCR method. RESULTS: The deltaCt value of MX1 expression in SLE patients was 3.55 +/- 1.39, which was significantly higher than those of non-SLE patients (2.31 +/- 0.52, P = 0.000) and normal controls (2.23 +/- 1.05, P = 0.000). (2) The deltaCt value of OAS1 expression in SLE patients was 4.45 +/- 1.56, which also was significantly higher than those of non-SLE patients (3.03 +/- 0.76, P = 0.000) and normal controls (2.75 +/- 0.64, P = 0.000). (3) The deltaCt value of OAS1 was correlated with the SLEDAI scores (r = 0.338, P = 0.019) and serum IgA level. (4) The ACt value of MX1 was not correlated with the SLEDAI scores (r = 0.064, P = 0.661), but correlated with the serum levels of triglyceride (TG), cholesterol (TC), low density lipoprotein (LDL), albuminuria of 24 hours (r = 0.428, P = 0.003 r = 0.383, P = 0.009 r = 0.394, P = 0.007; r = 0.316, P = 0.025); (5) The deltaCt values of MX1 and OAS1 in the SLE patients with arthritis were significantly higher than those in non-arthritis SLE patients (3.04 +/- 1.42, P = 0.004; 3.89 +/- 1.49, P = 0.006). CONCLUSION: The expression levels of both MX1 and OAS1 in SLE patients are up-regulated, the expression levels of OAS1 genes are associated with SLE disease activity. As IFN-induced genes, MX1 and OAS1 play their respective role in SLE.

[Determination of mineral elements in brown rice of near-isogenic lines population for Japonica rice by ICP-AES].

In the present paper, the contents of 17 mineral elements (S, Mo, Ni, Fe, Cr, Na, Al, Cu, P, Sn, Zn, B, Mn, Mg, Ca, Sr and K) in brown rice of 264 plant lines in near-isogenic lines (NILs) population and parents for Japonica rice were determined by ICP-AES technique. The method proved to be simple, rapid, highly sensitive and accurate, and can be used to determine many elements at the same time. Its recovery ratio obtained by standard addition method ranged between 92.3% and 110.27%, and its RSD was lower than 6%. The analytical results showed that NILs for various mineral elements based on the BC5F5 population consisting of 261 lines were developed from a cross between NIL (BC4F5) and Towada. To our knowledge, this is the first report on that near-isogenic lines NIL(BC4F5) with the important value for high mineral elements associated with cold tolerance at booting stage have been bred. The results of the determination show that the brown rice is rich in eleven elements such as P, K, Mg, S, Ca, Zn, Mn, Na, Fe, Cu and Al necessary to human health. There is smaller variation of 5 macroelements (11.15%-16.45%) but larger variation of 12 microelements (16.57%-94.96%) in brown rice from populations. There is significant correlation among 95 from 136 pairs in 17 elements, moreover, especially the correlation (0.531**-0.921**) among 15 pairs of six micro-elements (Mo, Ni, Al, Sn, Cr and Sr) associated with the active components of functional rice is relatively higher than that of 10 pairs (0.175**-0.867**) of 5 macroelements (P, K, Ca, Mg and S), revealing the importance of microelements to catalysing and synthesizing the active components of functional rice. The above results provided reliable data and theory bases for gene location and cloning of controlling elements contents in brown rice, and further revealed the molecular and physiological mechanism of the relation between mineral elements in brown rice and cold tolerance at booting stage in japonica rice from Yunnan Province of China.

[Evaluation of morphological traits on near-isogenic lines of cold tolerance and molecular validation at booting stage in Japonica rice].

Five parents and their near-isogenic lines (NILs) were grown in field experiments in Kunming (elevation 1916 m, two kinds of water temperatures at booting stage were 19.5 +/- 0.7 degrees C and 17.8 +/- 1.1 degrees C) and Aziying (elevation 2150 m, water temperature at booting stage was 18.2 +/- 0.22 degrees C), Yunnan, China, in 2004. Seventeen agronomic traits and SSR molecular markers were clustered by Statistica. The results showed that: (1) The morphological traits of NILs of cold tolerance and recurrent parents (Towada) at booting stage were similar, but the traits related cold tolerance, such as spike neck length, full grains, seed setting rate, anther length, anther volume, were different obviously; (2) using 7 markers had polymorphism between Towada and NILs were chosen from 78 SSR markers, the cold tolerance of NILs proved to be different to Towada. This demonstrates that these NILs were promising population for fine mapping and cloning genes for cold tolerance.

Complete Sequences and Characterization of Two Novel Plasmids Carrying aac(6')-Ib-cr and qnrS Gene in Shigella flexneri.

The complete sequences of two previously reported plasmids carrying plasmid-mediated quinolone resistance genes from Shigella flexneri in China have not been available. The present study using the p5-C3 assembly method revealed that (1) the plasmid pSF07201 with aac(6')-Ib-cr had 75,335 bp with antibiotic resistance genes CTX-M-3, TEM-1, and FosA3; (2) seven fragments of pSF07201 had more than 99% homology with the seven corresponding plasmids; (3) the other plasmid pSF07202 with qnrS had 47,669 bp with antibiotic resistance gene TEM-1 and 99.95% homology with a segment of pKF362122, which has the qnrS gene from location 162,490 to 163,146. A conjugation and electrotransformation experiment suggested that these two plasmids might horizontally transfer between and coexist in Escherichia coli J53 and S. flexneri 2a 301. Either the aac(6')-Ib-cr or qnrS gene contributed to, but only the coexistence of the two genes conferred to the resistance to ciprofloxacin in these two strains. To the best of our knowledge, this is the first report of the complete sequences of the aac(6')-Ib-cr- and qnrS-positive plasmids in Shigella isolates. Our findings indicate that two genes probably evolve through horizontal plasmid transfer between the different bacterial types.

Time-concentration-mortality modeling of the synergistic interaction of Beauveria bassiana and imidacloprid against Nilaparvata lugens.

Interactions between the entomopathogenic fungus, Beauveria bassiana SG8702, and imidacloprid on Nilaparvata lugens (Homoptera: Delphacidae) were studied in laboratory bioassays by spraying suspensions of unformulated conidia (assay 1) and aqueous dilutions of emulsifiable conidia formulation alone (assay 2) or together with imidacloprid at the sub-lethal rates of 0.5, 1.0 and 2.0 microg ml(-1) (assays 3-5). Each assay consisted of five conidia concentrations plus an appropriate control and three replicates, each including 30-40 third-instar nymphs, so as to generate time-concentration-mortality data for modeling analysis. A mineral oil-based emulsion used to formulate B bassiana slightly enhanced fungal activity but had no significant impact on the background mortality of N lugens. On the basis of LC50 estimates and associated variances on days 4-12 after spraying, synergistic interactions of both agents or formulations were determined by estimating relative potencies of assay 2 over assay 1 (1.2-9.0), assay 3 over assay 2 (1.3-1.7), assay 4 over assay 2 (7.5-9.6), assay 5 over assay 2 (22.7-101), assay 4 over assay 3 (3.8-5.8), assay 5 over assay 3 (16.1-61.0), and assay 5 over assay 4 (3.0-10.5). The time-concentration-mortality modeling method was not only mathematically but also biologically robust to evaluate the interactions of B bassiana and imidacloprid on N lugens. Compared with their counterparts, enhanced fungal formulations displayed consistently earlier or greater activities against the pest species based on LC50 and LT50 estimates determined from their time-concentration-mortality relationships. The results highlight a potential for pest control by combined formulation or application of B bassiana and imidacloprid.