Control of late maturity alpha-amylase in wheat by the dwarfing gene Rht-D1b and genes on the 1B/1R translocation

The occurrence of late maturity alpha-amylase (LMA) was investigated using two doubled haploid wheat populations segregating for the dwarfing gene Rht-D1b and the 1B/1R translocation. Genotypes were assessed in the field and in controlled environments where a cold-shock treatment was used to induce LMA. Results from field-grown genotypes from the cross Spark × Rialto suggest that the absence of Rht-D1b or the presence of the 1B/1R translocation increases the expression of LMA.These two genetic factors were found to act independently and to have a positive interaction (complementary epistasis). In Option × Potent genotypes fixed for Rht-D1b, the 1B/1R effect was similar to that seen in the equivalent Spark × Rialto genotypes. Under controlled environment conditions, genotypes with the 1B/1R translocation showed a higher occurrence of LMA under both control and cold-shock conditions. 1B/1R was present in the majority of genotypes expressing LMA under control and cold-shock conditions. The results point to the novel finding that the 1B/1R translocation increases the expression of alpha-amylase in LMA-prone germplasm independently of effects of Rht-D1b, whereas previously it had been thought to act by a modification of the Rht-D1b effect.     

Using Real-Time PCR to Assess Changes in the Hydrocarbon-Degrading Microbial Community in Antarctic Soil During Bioremediation

A real-time polymerase chain reaction (PCR) method to quantify the proportion of microorganisms containing alkane monooxygenase was developed and used to follow changes in the microbial community in hydrocarbon-contaminated Antarctic soil during a bioremediation field trial. Assays for the alkB and rpoB genes were validated and found to be both sensitive and reproducible (less than 2% intrarun variation and 25–38% interrun variation). Results from the real-time PCR analysis were compared to analysis of the microbial population by a culture-based technique [most probable number (MPN) counts]. Both types of analysis indicated that fertilizer addition to hydrocarbon-contaminated soil stimulated the indigenous bacterial population within 1 year. The proportion of alkB containing microorganisms was positively correlated to the concentration of n-alkanes in the soil. After the concentration of n-alkanes in the soil decreased, the proportion of alkane-degrading microorganisms decreased, but the proportion of total hydrocarbon-degrading microorganisms increased, indicating another shift in the microbial community structure and ongoing biodegradation.     

Location of a gene regulating cold-induced carbohydrate production on chromosome 5A of wheat

 Major changes in osmotic potential during cold acclimation are due to changes in sugar concentration, and there is a good correlation between sugar content and frost tolerance. The objective of the present study was to localize a gene(s) responsible for carbohydrate accumulation during cold acclimation on chromosome 5A of wheat using recombinant lines developed from the cross between the substitution lines Chinese Spring (Cheyenne 5A) and CS(Triticum spelta 5A). Previously, major genes influencing frost resistance (Fr1) and vernalization requirement (Vrn1) had been localized on the long arm of that chromosome. The T. spelta 5A chromosome carrying the Fr1 (frost-sensitive) allele for frost tolerance and the Vrn1 (spring-habit) allele for vernalization requirement did not have a major effect on the sucrose and fructan contents in the Chinese Spring background. On the other hand, the presence of Cheyenne alleles for vernalization requirement, vrn1, and frost tolerance, fr1, significantly increased sugar concentrations. A recombinant line thought to exhibit recombination between the Vrn1 and Fr1 loci suggested that the gene regulating sucrose accumulation was closely associated with, or else represented a pleiotropic effect of, Vrn1, but was separable from the Fr1 locus. Received: 3 March 1997 / Accepted: 7 March 1997     

Evaluation of the Induction of Immune Memory following Infant Immunisation with Serogroup C Neisseria meningitidis Conjugate Vaccines – Exploratory Analyses within a Randomised Controlled Trial

Aim

We measured meningococcal serogroup C (MenC)-specific memory B-cell responses in infants by Enzyme-Linked Immunospot (ELISpot) following different MenC conjugate vaccine schedules to investigate the impact of priming on immune memory.

Methods

Infants aged 2 months were randomised to receive 1 or 2 doses of MenC-CRM₁₉₇ at 3 or 3 and 4 months, 1 dose of MenC-TT at 3 months, or no primary MenC doses. All children received a Haemophilus influenzae type b (Hib)-MenC booster at 12 months. Blood was drawn at 5, 12, 12 months +6 days and 13 months of age.

Results

Results were available for 110, 103, 76 and 44 children from each group respectively. Following primary immunisations, and prior to the 12-month booster, there were no significant differences between 1- or 2-dose primed children in the number of MenC memory B-cells detected. One month following the booster, children primed with 1 dose MenC-TT had more memory B-cells than children primed with either 1-dose (p = 0.001) or 2-dose (p<0.0001) MenC-CRM₁₉₇. There were no differences in MenC memory B-cells detected in children who received 1 or 2 doses of MenC-CRM₁₉₇ in infancy and un-primed children.

Conclusions

MenC-specific memory B-cell production may be more dependent on the type of primary vaccine used than the number of doses administered. Although the mechanistic differences between MenC-CRM₁₉₇ and MenC-TT priming are unclear, it is possible that structural differences, including the carrier proteins, may underlie differential interactions with B- and T-cell populations, and thus different effects on various memory B-cell subsets. A MenC-TT/Hib-MenC-TT combination for priming/boosting may offer an advantage in inducing more persistent antibody.

Trial Registration

EU Clinical Trials Register 2009-016579-31 ClinicalTrials.gov {"type":"clinical-trial","attrs":{"text":"NCT01129518","term_id":"NCT01129518"}}NCT01129518     

Persistence of specific bactericidal antibodies at 5 years of age after vaccination against serogroup B meningococcus in infancy and at 40 months

Background:

The multicomponent serogroup B meningococcal (4CMenB) vaccine induces antibodies against indicator strains of serogroup B meningococcus under various schedules. We investigated the persistence of antibodies in 5-year-old children 18–20 months after their last dose (at about 3.5 years of age).

Methods:

We assessed 5-year-old children who received the 4CMenB vaccine or a recombinant protein vaccine in a previous randomized trial. We also recruited 50 vaccine-naive 5-year-olds and administered 2 doses of 4CMenB to each child. We measured serum bactericidal antibody titres against 4 indicator strains of serogroup B meningococcus matched to each individual vaccine component and against 4 mismatched strains.

Results:

Of those who received the 4CMenB vaccine at 2, 4, 6, 12 and 40 months (n = 16), the percentage with protective antibody titres (≥ 1:4) at 60 months ranged from 44% to 88% against matched strains and from 13% to 81% against mismatched strains. Loss of protective titres was also observed for those who received the 4CMenB vaccine at 12, 40 and 42 months (n = 5) (80%–100% against matched strains, 60%–100% against mismatched strains) or at 40 and 42 months (n = 29) (31%–100% against matched strains, 41%–81% against mismatched strains). Administering the 4CMenB vaccine to 5-year-old children yielded protective titres against matched strains in 92%–100% and against mismatched strains in 59%–100%. The majority of these children reported injection-site pain (40/50 [80%] after dose 1, 39/46 [85%] after dose 2) and erythema (47/50 [94%] and 40/46 [87%], respectively); rates of fever were low (5/50 [10%] and 2/46 [4%], respectively).

Interpretation:

Waning of immunity by 5 years of age occurred after receipt of the 4CMenB vaccine in infancy, even with an additional booster at 40 months. The 4CMenB vaccine is immunogenic and was fairly well tolerated by 5-year-old children, although injection-site pain was noteworthy. Trial registration: ClinicalTrials.gov, no. {"type":"clinical-trial","attrs":{"text":"NCT01027351","term_id":"NCT01027351"}}NCT01027351The multicomponent serogroup B meningococcal (4CMenB) vaccine is licensed in the European Union, Australia and Canada to prevent serogroup B meningococcal disease. It was developed using “reverse vaccinology,” in which candidate antigens were identified by interrogating the whole meningococcal genome.¹ The 4CMenB vaccine consists of 3 surface proteins (factor H binding protein [fHbp], Neisseria adhesin A [NadA] and Neisseria heparin-binding antigen [NHBA]), along with a fourth component, the outer membrane vesicle, which acts as both antigen and adjuvant.¹Group B meningococcal disease is a potentially devastating condition, with an average case fatality rate of 5.2% (data for England and Wales²), and over a third of survivors are left with measurable functional deficits.³ The incidence of laboratory-confirmed cases is about 1 per 100 000 population in England⁴ and 0.33 per 100 000 population in Canada.⁵ The recommendation of the United Kingdom Joint Committee on Vaccination and Immunisation that the 4CMenB vaccine be introduced into the routine UK immunization schedule should, if implemented, lead to a reduction in this morbidity and mortality.⁶ Data on the persistence of antibody responses following infant or toddler immunization, and after subsequent boosting, remain limited yet will be important for guiding implementation of this recommendation.We present here the results of a follow-on study investigating the persistence of antibodies 18–20 months after the last dose in 5-year-old children previously immunized under a variety of schedules with 4CMenB vaccine or another investigational vaccine (recombinant protein serogroup B meningococcal [rMenB] vaccine), which lacks the outer membrane vesicle component of the 4CMenB vaccine. Since the original infant study,⁷ 4CMenB vaccine has emerged as the preferred vaccine, because addition of the outer membrane vesicle component improves the breadth of strain coverage;⁸ however, the extension study continued follow-up for all of the original children, and all results are therefore presented here.     

Neuronal Antibodies in Children with or without Narcolepsy following H1N1-AS03 Vaccination

Type 1 narcolepsy is caused by deficiency of hypothalamic orexin/hypocretin. An autoimmune basis is suspected, but no specific antibodies, either causative or as biomarkers, have been identified. However, the AS03 adjuvanted split virion H1N1 (H1N1-AS03) vaccine, created to protect against the 2009 Pandemic, has been implicated as a trigger of narcolepsy particularly in children. Sera and CSFs from 13 H1N1-AS03-vaccinated patients (12 children, 1 young adult) with type 1 narcolepsy were tested for autoantibodies to known neuronal antigens including the N-methyl-D-aspartate receptor (NMDAR) and contactin-associated protein 2 (CASPR2), both associated with encephalopathies that include disordered sleep, to rodent brain tissue including the lateral hypothalamus, and to live hippocampal neurons in culture. When sufficient sample was available, CSF levels of melanin-concentrating hormone (MCH) were measured. Sera from 44 H1N1-ASO3-vaccinated children without narcolepsy were also examined. None of these patients’ CSFs or sera was positive for NMDAR or CASPR2 antibodies or binding to neurons; 4/13 sera bound to orexin-neurons in rat brain tissue, but also to other neurons. MCH levels were a marginally raised (n = 8; p = 0.054) in orexin-deficient narcolepsy patients compared with orexin-normal children (n = 6). In the 44 H1N1-AS03-vaccinated healthy children, there was no rise in total IgG levels or in CASPR2 or NMDAR antibodies three weeks following vaccination. In conclusion, there were no narcolepsy-specific autoantibodies identified in type 1 narcolepsy sera or CSFs, and no evidence for a general increase in immune reactivity following H1N1-AS03 vaccination in the healthy children. Antibodies to other neuronal specific membrane targets, with their potential for directing use of immunotherapies, are still an important goal for future research.     

Determination of the aromatic lignin content in oak wood by quantitative solid state 13C-NMR

The problems concerning quantification with cross polarization (CP) and high-field ¹³C-nmr measurements has meant that, for ligno-cellulosic plant materials, aromatic carbons in lignins are often discriminated against. In this study, the aromatic lignin content of an American red oak sample has been determined at the relatively low field strength of 25 MHz to obviate problems with spinning side bands using both CP and Bloch decay or single pulse excitation (SPE), a more time-consuming acquisition technique but that is, in many cases, considerably more quantitative than CP. The value of 14 mole % carbon from SPE is in close agreement with that of 15% derived from elemental analysis and the Klason lignin content. Although virtually all of the carbon was observed by both SPE and CP, the latter significantly underestimated the aromatic content at contact times less than 1.5 ms and thus longer times should be used for reliable quantification. The quaternary carbon content was estimated as 11 mole % carbon by combining dipolar dephasing with SPE and CP. © 1992 John Wiley & Sons, Inc.