Development of reverse-transcription PCR techniques to analyse the density and sex ratio of gametocytes in genetically diverse Plasmodium chabaudi infections

We have developed cross-genotype and genotype-specific quantitative reverse-transcription PCR (qRT-PCR) assays to detect and quantify the number of parasites, transmission stages (gametocytes) and male gametocytes in blood stage Plasmodium chabaudi infections. Our cross-genotype assays are reliable, repeatable and generate counts that correlate strongly (R(2)s > 90%) with counts expected from blood smears. Our genotype-specific assays can distinguish and quantify different stages of genetically distinct parasite clones (genotypes) in mixed infections and are as sensitive as our cross-genotype assays. Using these assays we show that gametocyte density and gametocyte sex ratios vary during infections for two genetically distinct parasite lines (genotypes) and present the first data to reveal how sex ratio is affected when each genotype experiences competition in mixed-genotype infections. Successful infection of mosquito vectors depends on both gametocyte density and their sex ratio and we discuss the implications of competition in genetically diverse infections for transmission success

Predictability of IL-28B-polymorphism on protease-inhibitor-based triple-therapy in chronic HCV-genotype-1 patients: A meta-analysis

AIM: To investigate the predictability of interleukin-28B single nucleotide polymorphism rs12979860 with respect to sustained virological response (SVR) in chronically hepatitis C virus (HCV) genotype-1 patients treated with a protease-inhibitor and pegylated interferon-$\alpha$ (Peg-INF-$\alpha$) based triple-therapy. METHODS: We searched PubMed, the Cochrane Library and Web of Knowledge for studies regarding the interleukin 28B (IL-28B)-genotype and protease-inhibitor based triple-therapy. Ten studies with 2707 patients were included into this meta-analysis. We used regression methods in order to investigate determinants of SVR. RESULTS: IL-28B-CC-genotype patients achieved higher SVR rates (odds 5.34, CI: 3.81-7.49) than IL-28B-non-CC-genotype patients (1.88, CI: 1.43-2.48) receiving triple-therapy. The line of therapy (treatment-na\"ive or -experienced for Peg-INF-$\alpha$) did not affect the predictive value of IL-28B (p=0.1). IL-28B-CC-genotype patients treated with protease inhibitor-based triple-therapy consisting of Boceprevir, Simeprevir, Telaprevir or Vaniprevir showed odds of 1.86, 9.77, 4.51 and 0.89, respectively. The odds for CC genotype patients treated with Faldaprevir cannot be quantified, as only a single study with a 100% SVR rate was available. CONCLUSION: IL-28B-SNP predicts the outcome for chronic HCV genotype-1 patients receiving protease inhibitor-based triple-therapy. The predictive value varies between the different protease inhibitors.Comment: 21 pages, 3 figures, 1 tabl

Associations between CXCR1 polymorphisms and pathogen-specific incidence rate of clinical mastitis, test-day somatic cell count, and test-day milk yield

The CXCR1 gene plays an important role in the innate immunity of the bovine mammary gland. Associations between single nucleotide polymorphisms (SNP) CXCR1c.735C>G and c.980A>G and udder health have been identified before in small populations. A fluorescent multiprobe PCR assay was designed specifically and validated to genotype both SNP simultaneously in a reliable and cost-effective manner. In total, 3,106 cows from 50 commercial Flemish dairy herds were genotyped using this assay. Associations between genotype and detailed phenotypic data, including pathogen-specific incidence rate of clinical mastitis (IRCM), test-day somatic cell count, and test-day milk yield (MY) were analyzed. Staphylococcus aureus IRCM tended to associate with SNP c.735C>G. Cows with genotype c.735GG had lower Staph. aureus IRCM compared with cows with genotype c.735CC (rate ratio = 0.35, 95% confidence interval = 0.14-0.90). Additionally, a parity-specific association between Staph. aureus IRCM and SNP c.980A>G was detected. Heifers with genotype c.980GG had a lower Staph. aureus IRCM compared with heifers with genotype c.980AG (rate ratio = 0.15, 95% confidence interval = 0.04-0.56). Differences were less pronounced in multiparous cows. Associations between CXCR1 genotype and somatic cell count were not detected. However, MY was associated with SNP c.735C>G. Cows with genotype c.735GG out-produced cows with genotype c.735CC by 0.8 kg of milk/d. Results provide a basis for further research on the relation between CXCR1 polymorphism and pathogen-specific mastitis resistance and MY

PON1 status does not influence cholinesterase activity in Egyptian agricultural workers exposed to chlorpyrifos.

Animal studies have shown that paraoxonase 1 (PON1) genotype can influence susceptibility to the organophosphorus pesticide chlorpyrifos (CPF). However, Monte Carlo analysis suggests that PON1 genotype may not affect CPF-related toxicity at low exposure conditions in humans. The current study sought to determine the influence of PON1 genotype on the activity of blood cholinesterase as well as the effect of CPF exposure on serum PON1 in workers occupationally exposed to CPF. Saliva, blood and urine were collected from agricultural workers (n=120) from Egypt's Menoufia Governorate to determine PON1 genotype, blood cholinesterase activity, serum PON1 activity towards chlorpyrifos-oxon (CPOase) and paraoxon (POase), and urinary levels of the CPF metabolite 3,5,6-trichloro-2-pyridinol (TCPy). The PON1 55 (P≤0.05) but not the PON1 192 genotype had a significant effect on CPOase activity. However, both the PON1 55 (P≤0.05) and PON1 192 (P≤0.001) genotypes had a significant effect on POase activity. Workers had significantly inhibited AChE and BuChE after CPF application; however, neither CPOase activity nor POase activity was associated with ChE depression when adjusted for CPF exposure (as determined by urinary TCPy levels) and stratified by PON1 genotype. CPOase and POase activity were also generally unaffected by CPF exposure although there were alterations in activity within specific genotype groups. Together, these results suggest that workers retained the capacity to detoxify chlorpyrifos-oxon under the exposure conditions experienced by this study population regardless of PON1 genotype and activity and that effects of CPF exposure on PON1 activity are minimal

Association between MAPT polymorphism but not APOE promoter and elite rugby athlete status

INTRODUCTION: Incidence and outcomes of concussions have been hypothesised to be genetically influenced. The APOE Promoter G219T (rs405509) polymorphism has been associated with differential promoter activity and unfavourable outcomes after traumatic brain injury. The TT genotype is associated with a 3-fold greater risk of multiple concussions. The TT genotype of MAPT (rs10445337) has also been associated with poorer outcomes after concussion. Rugby has one of the highest incidences of concussion in sport, so it was hypothesised that APOE Promoter TT and MAPT TT genotypes would be less prevalent in elite rugby athletes because those genotypes, previously associated with increased risk, would be less compatible with achieving elite athlete status. METHODS: Participants were from the RugbyGene project, comprising elite Caucasian male rugby athletes (n = 528; mean (standard deviation) height 1.85 (0.07) m, mass 101 (14) kg, age 29 (7) yr), including 420 rugby union (RU) athletes that for some analyses were divided into forwards and backs and 108 rugby league (RL) athletes. Non-athletes were 592 Caucasian men and women (57% male, height 1.72 (0.10) m, mass 74 (14) kg, age 31 (7) yr). PCR of genomic DNA was used to determine genotypes using TaqMan probes, then groups were compared using χ2 and odds ratio (OR) statistics. RESULTS: All genotype data were in Hardy-Weinberg equilibrium. For MAPT (rs10445337), the risk genotype (TT) was underrepresented in rugby athletes (60%) compared to non-athletes (66%), CT more common in rugby athletes (34%) than non-athletes (29%) and little difference in CC genotype frequencies (χ2 = 7.092, P = 0.029; TT genotype frequency OR = 0.80, 95% confidence intervals (CI) = 0.62-1.02). There were no differences in MAPT (rs10445337) genotype frequencies between RU forwards and backs. For APOE Promoter G219T (rs405509), there were no differences in genotype frequencies between all athletes (RU and RL) and non-athletes (27% TT genotype in players and non-athletes), nor between RU forwards and backs. CONCLUSION: The MAPT (rs10445337) TT genotype is 6% less common in elite rugby athletes than non-athletes. Therefore, carrying at least one rs10445337 C allele appears to increase the probability of sustained career success in the high-risk concussion environment of elite rugby, perhaps via a greater ability to recover from concussions.Peer reviewe

Effects of plant density on seed yield and quality in different common beans (Phaseolus vulgaris L.) : a thesis presented in partial fulfilment of the requirements for the degree of Master of Agricultural Science in Plant Science in Seed Technology at Massey University, New Zealand

Common beans (Phaseolus vulgaris L.) are an annual legume used for human consumption. While many cultivars/genotypes have long been a feature of New Zealand home gardens and the frozen food market, there has recently been an interest in the production of new genotypes of this crop legume suitable for use particularly in fresh and canned salads, as well as for other commercial purposes. In New Zealand, little is known of the growth and performance of many genotypes of this plant as the agro-climatological conditions are different from the original native South American habitat. Therefore this study covered an evaluation of five unnamed but different seed coloured bean genotypes obtained originally from the CIAT collection by New Zealand Seed Bank Ltd. The objectives of this research were to: determine plant growth habit of the genotypes; describe plant growth habit of the genotypes; and assess the effect of plant density on vegetative and reproductive growth, seed yield and cooking quality. To facilitate the recognition of each genotype, they were named white, mottled brown, mottled black, black and brown according to their seed colour, after a visual selection of seeds at the beginning of this study. Plant morphological characteristics were assessed in a trial conducted in a glasshouse at the Seed Technology Centre, Massey University, from September to November 1994. A field trial from November 1994 to March 1995 was aimed at determining the effects of plant density and genotype on seed production and quality for sowing and eating purposes. The minimum and maximum temperatures in the glasshouse were 16°C and 25°C respectively. The daylength in September was around 11 h and gradually increased to about 14.5 h at the end of November. No supplementary illumination and no pesticides and insecticides were used in this trial. For this study, five plants of each colour group were used to determine plant morphological characteristics which included: leaf length and width for the 1st, 3rd and 8th trifoliolate leaves, recorded from the terminal leaf; length of pedicellate bracts; flower (standard and wing) colour; pod colour, length and width; plant height and branch number; main stem internode number and internode length; pods per plant; and seeds per pod. Trifoliolate leaf length was around 22 cm for all genotypes irrespective of leaf position, but leaf width increased from the 1st to the 8th trifoliolate leaf and differed with genotype. For example the 8th trifoliolate leaf width ranged from 11.0 cm in the mottled brown genotype to 14.6 cm in the brown genotype. Pedicellate bract length, main stem internode number and maximum internode length all varied with genotype, with the result that average plant height ranged from 166 cm for the brown genotype to 362 cm for the white genotype. None of the genotypes produced branches in the glasshouse. Flower colour was assessed using the Dictionary of Colour Standards and the Horticultural Colour Chart from the British Colour Council. The standard and wing were white in the white, mottled brown and brown genotypes, mauve in the mottled black genotype, and were either white or mauve to rose purple in the black genotype. The colour of the wing was mauve in the mottled black genotype and was either white or mauve in the black genotype. Pod colour for the white genotype was mimosa yellow to naples yellow, or mottled with either aster violet or hyacinth blue, while in the mottled brown genotype pod colour was predominantly naples yellow, mottled with china rose or also with chrysanthemum crimson. Pods from the mottled black genotype were mimosa yellow to amber yellow in colour, and sometimes mottled with purple brown. Pods from the black genotype were mimosa yellow or naples yellow and were either slightly mottled with lilac purple or with pansy violet, while pod colour from the brown genotype was erythrite red. Dried pod length varied from 9.3 to 12.1 cm in the brown and white genotypes respectively, while dried pod width ranged from 11.8 mm in the mottled black to 12.8 mm in the white genotype. The number of pods per plant varied from 13 in the mottled brown to 16 in the brown genotype, while seeds per pod varied from 4.4 in the brown genotype to 5.8 in the while genotype. Daylength for the field trial ranged from 14.5 h (November) to around 12.3 h (March), with a maximum daylength of about 15 h in December. Seeds from the same seed colour groups used for the glasshouse studies were used in the field trial which was located at the Frewen's block, Massey University. Seeds were sown at three different rates (2.8, 5.6 and 8.4 g/m2) by cone seeder on 28 November 1994 to obtain densities of 6.6, 13.3 and 20.0 plants/m2 at row spacings of 60, 30 and 20 cm respectively. Within the rows a uniform space of 25 cm was maintained. Each treatment (plant density x genotype) was replicated four times in a split plot design. For seed development studies, a total of 450 - 460 flowers per genotype (from the 13.3 plants/m2 density) were randomly selected and labelled at anthesis, and 60 pods per individual genotype were harvested manually at 14, 20, 26, 32, 40 and 50 days after labelling for the determination of seed moisture content, fresh weight, dry weight and percentage seed germination. Seed yield and seed yield components (number of pods per plant and seeds per pod) were recorded after hand harvesting of 10 sample plants/plot. The quality of seed for sowing purposes was assessed by germination, conductivity and accelerated ageing (AA) tests, while for cooking quality, seeds were assessed for their imbibition rate, seed texture and seed integrity after cooking. All the data acquired from this study were analyzed with the statistical analysis system of SAS with least significant differences at the 5% level. The white and black bean genotypes produced 11 and 17% plants with indeterminate climbing characteristics respectively, while the other genotypes each produced 1 - 3% of plants with indeterminate climbing characteristics. All other plants were bush-indeterminate. Plant height in all bean genotypes at all the densities measured between 50 - 60 cm with a min./max. height of 40/85 cm. The onset, peak and duration of flowering in all genotypes were not affected by plant density. The typical three phase sequence of seed development was recorded and physiological maturity, or the attainment of maximum dry weight, occurred at around 40 days after anthesis (d.a.a.) at more or less the same time for all genotypes. Seed germination started around 20 d.a.a. and reached a maximum (of 100%) about the same time as the attainment of maximum seed dry weight at 40 d.a.a. However differences in seed coat permeability influenced the rate of seed desiccation and caused differences in seed moisture content (smc) among genotypes. The number of branches per plant differed significantly from 4.6 in the brown genotype to 5.2 - 5.8 in other genotypes. At the 6.6 plants/m2 density the number of branches per plant was 7.0 and decreased to 3.8 at the 20.0 plants/m2 density. Flowers per plant varied from 46.9 to 63.9 in the brown and mottled brown genotypes respectively but did not differ with density. Pods per plant were similar for all genotypes, and reached 32.2 at the 6.6 plants/m2 density but decreased to 19.0 at the 20.0 plants/m2 density. Seeds per pod varied slightly from 4.1 in the brown genotype to 4.6 in the mottled black genotype, but did not differ with density. Seed weight/100 seeds varied from 35.7 g in the mottled black genotype to 45.2 g in the black genotype, and was similar for all densities. The black genotype produced an average seed yield of 5,705 kg/ha (the highest), while the brown genotype had an average of 4,723 kg/ha (the lowest) at 10% smc. The average seed yield from the white, mottled brown and mottled black genotypes did not differ from that of the black genotype. There was no genotype x density interaction and the average seed yield for all genotypes was 3,800 kg/ha at the 6.6 plants/m2 density, 5,366 kg/ha at the 13.3 plams/m2 density and 6,715 kg/ha at the 20.0 plants/m2 density. The conductivity test result varied from 4.7 μS cm-1 g-1 in the brown genotype to 15.5 μS cm-1 g-1 in the white genotype, which demonstrated the differences in seed coat characteristics among genotypes. The germination before AA was 100% for all genotypes, and did not differ after AA (between 97 and 99%). The conductivity test result, as well as the percentage germination before and after AA did not differ with density. The brown genotype produced an average of 53.5% of seeds with 'delayed permeable' seed coats, while this property varied from 3.5 to 10.0% in the white and mottled brown genotypes respectively. Seeds from all genotypes became soft after cooking for 20 min. However the force required to cut the seeds after cooking varied from 6.83 Newton in the white genotype to 15.23 Newton in the brown genotype. The high seed coat permeability in the white genotype caused 19.3% seed coat/cotyledonary damage, while the brown genotype had only 8.9%. The white, mottled brown, mottled black and black genotypes produced a high yield (between 5,136 and 5,705 kg/ha) of good quality seed for both sowing and eating purposes. The brown genotype had a lower seed yield (4,723 kg/ha), but the seed was also of good quality for sowing and eating purposes. Differences in the seed coat characteristic of different bean genotypes may mean a requirement for different lengths of cooking time to attain the same level of seed softness without seed coat splitting as required for human consumption

TGFβ1 single-nucleotide polymorphism C-509T alters mucosal cell function in pediatric eosinophilic esophagitis.

Eosinophilic esophagitis (EoE) is a chronic Th2 antigen-driven disorder associated with tissue remodeling. Inflammation and remodeling lead to esophageal rigidity, strictures, and dysphagia. TGFβ1 drives esophageal remodeling including epithelial barrier dysfunction and subepithelial fibrosis. A functional SNP in the TGFβ1 gene that increases its transcription (C-509T) is associated with elevated numbers of esophageal TGFβ1-expressing cells. We utilized esophageal biopsies and fibroblasts from TT-genotype EoE children to understand if TGFβ1 influenced fibroblast and epithelial cell function in vivo. Genotype TT EoE esophageal fibroblasts had higher baseline TGFβ1, collagen1α1, periostin, and MMP2 (p < 0.05) gene expression and distinct contractile properties compared with CC genotype (n = 6 subjects per genotype). In vitro TGFβ1 exposure caused greater induction of target gene expression in genotype CC fibroblasts (p < 0.05). Esophageal biopsies from TT-genotype subjects had significantly less epithelial membrane-bound E-cadherin (p < 0.01) and wider cluster distribution at nanometer resolution. TGFβ1 treatment of stratified primary human esophageal epithelial cells and spheroids disrupted transepithelial resistance (p < 0.001) and E-cadherin localization (p < 0.0001). A TGFβ1-receptor-I inhibitor improved TGFβ1-mediated E-cadherin mislocalization. These data suggest that EoE severity can depend on genotypic differences that increase in vivo exposure to TGFβ1. TGFβ1 inhibition may be a useful therapy in subsets of EoE patients

Noise in Genotype Selection Model

We study the steady state properties of a genotype selection model in presence of correlated Gaussian white noise. The effect of the noise on the genotype selection model is discussed. It is found that correlated noise can break the balance of gene selection and induce the phase transition which can makes us select one type gene haploid from a gene group.Comment: 8 pages, 4 figure

Nosocomial hepatitis C virus infection in a renal transplantation center

AbstractNosocomial hepatitis C virus (HCV) infections were recorded in the renal transplantation unit of the university hospital. There were cases of acute HCV infection with aggressive clinical courses diagnosed from a positive HCV RNA test in the early post-transplantation period and which remained anti-HCV negative. Their anti-HCV seronegativity was attributed to them having acquired HCV under intense immunosuppressive therapy and suggested that the aggressive clinical course could be due to the deficient immune response resulting in an inability to limit viral replication. There were also donors diagnosed as having acute HCV infection in the early post-operative period. Genotyping and sequence analysis for HCV were performed on the isolates of eight of these patients who were consecutively transplanted and of three donors whose recipients were infected with HCV prior to transplantation, and who acquired acute HCV infection after transplantation. Of the eight recipients in the first group three were genotype 1a, three were genotype 1b, one wasgenotype 3a, and the last one was genotype 4 according to Simmond's classification. Of the three donor-recipient couples both the HCV isolates from one couple were genotyped as 1b and the phylogenetic analysis indicated that the patients were infected with a common variant of HCV, but the genotypesof HCV isolates from the other couples were different. Recipients were genotype 1b and the donors were genotype 1a in these couples. Genotype results of the first group and donor-recipient couples, and sequence analysis of genotype 1b and 1a isolates, showed that the source of infection was not a unique strain and there were multiple breaks in universal precautions while managing these patients