Comparison of the efficacy and safety of pollen allergen extracts using skin prick testing and serum specific IgE as references.

BACKGROUND: Allergen extracts may be different due to the difference in dissemination of allergen-containing species in various geographical areas. Therefore, we wish to develop our own extracts to ensure the precision and quality of diagnosis. OBJECTIVES: To compare the efficacy and safety of our locally prepared pollen allergen extracts to imported ones, using skin prick testing (SPT) and serum specific IgE (sIgE) as references. METHODS: This prospective, randomized, double-blinded, self-controlled study was performed in respiratory allergic adult volunteers who are sensitized to at least one kind of pollen. Each subject was pricked with our Bermuda grass, Johnson grass and careless weed pollen allergen extracts, and also with the imported ones. sIgE levels were measured by using ImmunoCAP?. RESULTS: In 68 volunteers, our Bermuda, Johnson and careless weed extracts showed 91.2%, 45.6% and 54.4% positive SPTs, respectively, while for the imported ones 73.5%, 45.6% and 54.4% SPTs were positive, respectively. No adverse reaction was found in all procedures. The concentration of 10,000 BAU/mL of Bermuda grass, 1 : 20 w/v or 10,000 PNU/mL of Johnson grass and 1 : 40 w/v or 10,000 PNU/mL of careless weed yielded the most positive SPT results. There was no significant difference in mean wheal diameter (MWD) yielded from using local and imported extracts. Significant correlation was found between MWDs of imported pollen extracts and serum sIgE levels (p < 0.01). CONCLUSIONS: No significant difference between SPT results of local and imported pollen allergen extracts was found. Significant correlation was found between MWDs of imported pollen extract SPT and serum sIgE levels.

Molecular characterization and In silico analysis of Sorghum Panallergens: Profilin and Polcalin.

In India, 20-30% of the human population suffer from allergic rhinitis and 15% of them develop asthma. Plant pollens are one of the causative aeroallergens and are mixture of a number of molecules including major and minor allergens (Panallergens). Profilin and polcalcin are the known pollen specific panallergens. Allergenicity of the Sorghum plant in Andhra Pradesh was found to be 54.9%. But the allergens responsible have not been characterized well. This study highlights identification and molecular characterization of Sorghum bicolor profilin (Sorb PF) and S. bicolor polcalcin (Sorb PC) allergen genes based on homology. The coding sequences of the two genes were PCR amplified from the cDNA constructed from Sorghum pollen total RNA. The gene sequences were deposited in NCBI, KC427126 and KC427125. Recombinantly expressed histidine tag (His-tag) purified Sorghum polcalcin and profilin confirmed 9 and 14 kDa proteins, respectively. Based on multiple sequence alignment and phylogenetic analysis, Sorghum polcalcin and profilin were found to be closely related with Cynodon dactylon, Phleum pratense and Oryza sativa grass species. In silico Algpred based screening of SorbPF and SorbPC showed an allergenicity score of 1.149 and 0.879, respectively. The structure of two Ef-hand sequences (DTNGDGKISLSEL and DTDGDGFIDFNEF) of SorbPC showed an exact match with Phlp7. It is concluded that Sorghum recombinant profilin and polcalcin proteins can be of potential use in developing diagnostic kits for allergenicity to Sorghum pollen grains.

Specific IgE recognition of pollen allergens from subtropic grasses in patients from the subtropics.

BACKGROUND: Pollens of subtropical grasses, Bahia (Paspalum notatum), Johnson (Sorghum halepense), and Bermuda (Cynodon dactylon), are common causes of respiratory allergies in subtropical regions worldwide. OBJECTIVE: To evaluate IgE cross-reactivity of grass pollen (GP) found in subtropical and temperate areas. METHODS: Case and control serum samples from 83 individuals from the subtropical region of Queensland were tested for IgE reactivity with GP extracts by enzyme-linked immunosorbent assay. A randomly sampled subset of 21 serum samples from patients with subtropical GP allergy were examined by ImmunoCAP and cross-inhibition assays. RESULTS: Fifty-four patients with allergic rhinitis and GP allergy had higher IgE reactivity with P notatum and C dactylon than with a mixture of 5 temperate GPs. For 90% of 21 GP allergic serum samples, P notatum, S halepense, or C dactylon specific IgE concentrations were higher than temperate GP specific IgE, and GP specific IgE had higher correlations of subtropical GP (r = 0.771-0.950) than temperate GP (r = 0.317-0.677). In most patients (71%-100%), IgE with P notatum, S halepense, or C dactylon GPs was inhibited better by subtropical GP than temperate GP. When the temperate GP mixture achieved 50% inhibition of IgE with subtropical GP, there was a 39- to 67-fold difference in concentrations giving 50% inhibition and significant differences in maximum inhibition for S halepense and P notatum GP relative to temperate GP. CONCLUSION: Patients living in a subtropical region had species specific IgE recognition of subtropical GP. Most GP allergic patients in Queensland would benefit from allergen specific immunotherapy with a standardized content of subtropical GP allergens.

Total transcriptome, proteome, and allergome of Johnson grass pollen, which is important for allergic rhinitis in subtropical regions.

BACKGROUND: Genomic data are lacking for many allergen sources. To circumvent this limitation, we implemented a strategy to reveal the repertoire of pollen allergens of a grass with clinical importance in subtropical regions, where an increasing proportion of the world's population resides. OBJECTIVE: We sought to identify and immunologically characterize the allergenic components of the Panicoideae Johnson grass pollen (JGP; Sorghum halepense). METHODS: The total pollen transcriptome, proteome, and allergome of JGP were documented. Serum IgE reactivities with pollen and purified allergens were assessed in 64 patients with grass pollen allergy from a subtropical region. RESULTS: Purified Sor h 1 and Sor h 13 were identified as clinically important allergen components of JGP with serum IgE reactivity in 49 (76%) and 28 (43.8%), respectively, of patients with grass pollen allergy. Within whole JGP, multiple cDNA transcripts and peptide spectra belonging to grass pollen allergen families 1, 2, 4, 7, 11, 12, 13, and 25 were identified. Pollen allergens restricted to subtropical grasses (groups 22-24) were also present within the JGP transcriptome and proteome. Mass spectrometry confirmed the IgE-reactive components of JGP included isoforms of Sor h 1, Sor h 2, Sor h 13, and Sor h 23. CONCLUSION: Our integrated molecular approach revealed qualitative differences between the allergenic components of JGP and temperate grass pollens. Knowledge of these newly identified allergens has the potential to improve specific diagnosis and allergen immunotherapy treatment for patients with grass pollen allergy in subtropical regions and reduce the burden of allergic respiratory disease globally.

Age versus stage: does ontogeny modify the effect of phosphorus and arbuscular mycorrhizas on above- and below-ground defence in forage sorghum?

Arbuscular mycorrhizas (AM) can increase plant acquisition of P and N. No published studies have investigated the impact of P and AM on the allocation of N to the plant defence, cyanogenic glucosides. We investigated the effects of soil P and AM on cyanogenic glucoside (dhurrin) concentration in roots and shoots of two forage sorghum lines differing in cyanogenic potential (HCNp). Two harvest times allowed plants grown at high and low P to be compared at the same age and the same size, to take account of known ontogenetic changes in shoot HCNp. P responses were dependent on ontogeny and tissue type. At the same age, P-limited plants were smaller and had higher shoot HCNp but lower root HCNp. Ontogenetically controlled comparisons showed a P effect of lesser magnitude, and that there was also an increase in the allocation of N to dhurrin in shoots of P-limited plants. Colonization by AM had little effect on shoot HCNp, but increased root HCNp and the allocation of N to dhurrin in roots. Divergent responses of roots and shoots to P, AM and with ontogeny demonstrate the importance of broadening the predominantly foliar focus of plant defence studies/theory, and of ontogenetically controlled comparisons.

QTL analysis of ergot resistance in sorghum.

Sorghum ergot, caused predominantly by Claviceps africana Frederickson, Mantle, de Milliano, is a significant threat to the sorghum industry worldwide. The objectives of this study were firstly, to identify molecular markers linked to ergot resistance and to two pollen traits, pollen quantity (PQ) and pollen viability (PV), and secondly, to assess the relationship between the two pollen traits and ergot resistance in sorghum. A genetic linkage map of sorghum RIL population R931945-2-2 x IS 8525 (resistance source) was constructed using 303 markers including 36 SSR, 117 AFLP , 148 DArT and two morphological trait loci. Composite interval mapping identified nine, five, and four QTL linked to molecular markers for percentage ergot infection (PCERGOT), PQ and PV, respectively, at a LOD >2.0. Co-location/linkage of QTL were identified on four chromosomes while other QTL for the three traits mapped independently, indicating that both pollen and non pollen-based mechanisms of ergot resistance were operating in this sorghum population. Of the nine QTL identified for PCERGOT, five were identified using the overall data set while four were specific to the group data sets defined by temperature and humidity. QTL identified on SBI-02 and SBI-06 were further validated in additional populations. This is the first report of QTL associated with ergot resistance in sorghum. The markers reported herein could be used for marker-assisted selection for this important disease of sorghum.