VH gene usage in immunoglobulin E responses of seasonal rhinitis patients allergic to grass pollen is oligoclonal and antigen driven

Background: IgE is the pivotal-specific effector molecule of allergic reactions yet it remains unclear whether the elevated production of IgE in atopic individuals is due to superantigen activation of B cell populations, increased antibody class switching to IgE or oligoclonal allergen-driven IgE responses. Objectives: To increase our understanding of the mechanisms driving IgE responses in allergic disease we examined immunoglobulin variable regions of IgE heavy chain transcripts from three patients with seasonal rhinitis due to grass pollen allergy. Methods: Variable domain of heavy chain-epsilon constant domain 1 cDNAs were amplified from peripheral blood using a two-step semi-nested PCR, cloned and sequenced. Results: The VH gene family usage in subject A was broadly based, but there were two clusters of sequences using genes VH 3-9 and 3-11 with unusually low levels of somatic mutations, 0-3%. Subject B repeatedly used VH 1-69 and subject C repeatedly used VH 1-02, 1-46 and 5a genes. Most clones were highly mutated being only 86-95% homologous to their germline VH gene counterparts and somatic mutations were more abundant at the complementarity determining rather than framework regions. Multiple sequence alignment revealed both repeated use of particular VH genes as well as clonal relatedness among clusters of IgE transcripts. Conclusion: In contrast to previous studies we observed no preferred VH gene common to IgE transcripts of the three subjects allergic to grass pollen. Moreover, most of the VH gene characteristics of the IgE transcripts were consistent with oligoclonal antigen-driven IgE responses.

A novel grass pollen allergen mimotope identified by phage display peptide library inhibits allergen-human IgE antibody interaction

The aim of this study was to investigate the molecular basis of human IgE-allergen interaction by screening a phage-displayed peptide library with an allergen-specific human IgE-mimicking monoclonal antibody (mAb). A mAb that reacted with major grass pollen allergens was successfully identified and shown to inhibit human IgE-allergen interaction. Biopanning of a phage-displayed random peptide library with this mAb yielded a 12 amino acid long mimotope. A synthetic peptide based on this 12-mer mimotope inhibited mAb and human IgE binding to grass pollen extracts. Our results indicate that such synthetic peptide mimotopes of allergens have potential as novel therapeutic agents. © 2001 Published by Elsevier Science B.V. on behalf of the Federation of European Biochemical Societies.

A rapid and efficient two-step gel electrophoresis method for the purification of major rye grass pollen allergens

Purified proteins are mandatory for molecular, immunological and cellular studies. However, purification of proteins from complex mixtures requires specialised chromatography methods (i.e., gel filtration, ion exchange, etc.) using fast protein liquid chromatography (FPLC) or high-performance liquid chromatography (HPLC) systems. Such systems are expensive and certain proteins require two or more different steps for sufficient purity and generally result in low recovery. The aim of this study was to develop a rapid, inexpensive and efficient gel-electrophoresis-based protein purification method using basic and readily available laboratory equipment. We have used crude rye grass pollen extract to purify the major allergens Lol p 1 and Lol p 5 as the model protein candidates. Total proteins were resolved on large primary gel and Coomassie Brilliant Blue (CBB)-stained Lol p 1/5 allergens were excised and purified on a secondary "mini"-gel. Purified proteins were extracted from unstained separating gels and subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and immunoblot analyses. Silver-stained SDS-PAGE gels resolved pure proteins (i.e., 875 μg of Lol p 1 recovered from a 8 mg crude starting material) while immunoblot analysis confirmed immunological reactivity of the purified proteins. Such a purification method is rapid, inexpensive, and efficient in generating proteins of sufficient purity for use in monoclonal antibody (mAb) production, protein sequencing and general molecular, immunological, and cellular studies.

Structural, physiological and molecular characterisation of the Australian native resurrection grass Tripogon loliiformis (F.Meull) C.E.Hubb during dehydration and rehydration

There is an urgent need to develop crops that can withstand future climates. Results from this thesis demonstrated that a native Australian resurrection grass exhibits structural, physiological and metabolic strategies to tolerate drying. These strategies may be utilized for the generation of stress tolerant crops.

Effect of seasonal conditions on fleece weight and its components in merino sheep grazing Mitchell grass pasture in North West Queensland

SUMMARY Seasonal conditions in the pre to post natal period and selected periods before and during wool growth were described using climatic measures and estimates of the quality and quantity of pasture on offer derived from a validated pasture production model (GRASP). The variation in greasy and clean fleece weight, yield, staple length, fibre diameter, neck and side wrinkle score of Merinos grazing Mitchell grass in north west Queensland was explained in terms of these pasture and climatic measures and animal characteristics such as reproductive status, age and skin area. Multiple regression equations predicting clean and greasy fleece weight from the proportion of days in the wool growth period that the green pool in the pasture was less than one kg/ha, the percentage utilisation of the pasture, age, reproductive status and skin area of the ewes explained 87% and 79% of the variation respectively. Equations with similar predictors explained 58-85% of the variation of the other components. The inclusion of pasture conditions in the pre to post natal period did not significantly improve the predictions of the animal’s later performance. 22nd Biennial Conference.

Milk Production Responses to Oversowing a Portion of Tropical Grass Pasture with Summer Crop Legumes on Sub-tropical Dairy Farms

Northern Australian dairy farms have a large area of tropical dryland grass pasture available for use as summer pastures. Late summer-autumn in sub-tropical Australia is traditionally a difficult period in which to produce milk because of the decline in both quality and quantity of tropical grasses (Ehrlich et al. 1994). Options to improve autumn feed on dairy farms include introducing forage crops and conservation, increasing concentrate feeding and introducing legumes. Perennial tropical legumes have not been successful at this time of year because of their inability to sustain stocking rates above one cow/ha. This experiment, conducted on farms, was designed to test if annual crop legumes could be successfully oversown into tropical grass areas using minimal till methods to measure the subsequent impact on milk production on farms. Previous experiments using annual legumes in plots at Mutdapilly Research Station had demonstrated yields up to 10 t/ha can be achieved using annual tropical legumes with protein levels as high as 20% in the whole legume plant. Animal production for a consuming world : proceedings of 9th Congress of the Asian-Australasian Association of Animal Production Societies [AAAP] and 23rd Biennial Conference of the Australian Society of Animal Production [ASAP] and 17th Annual Symposium of the University of Sydney, Dairy Research Foundation, [DRF]. 2-7 July 2000, Sydney, Australia.

Using Alkanes to Predict Voluntary Intake, Faecal Output and Digestibility of Buffel Grass Hay Fed to Steers

The cuticular waxes of forage plants contain long chain n-alkanes with odd carbon chain lengths in the range C25-C37 which are quantitatively recovered in faeces. When these concentrations are used with the concentrations of administered synthetic even chain length alkanes, the voluntary intake (VI), faecal output (FO) and digestibility (DMD) of forages can be estimated (Dove and Mayes 1991, 1996).

Response By Dairy Cows Grazing Tropical Grass Pasture To Barley or Sorghum Grain Based Concentrates and Lucerne Hay

This study investigated the responses by dairy cows grazing Callide Rhodes grass (Chloris gayana cv. Callide) pasture to supplementation with barley or sorghum based concentrates (5 grain:1 cotton seed meal) or barley concentrate plus lucerne (Medicago sativa) hay. It was conducted in summer - autumn 1999 with 20 spring calved cows in 4 treatments in 3 consecutive periods of 4 weeks. Rain grown pastures, heavily stocked at 4.4 cows/ha, provided 22 to 35 kg green DM and 14 to 16 kg green leaf DM/cow.day in periods 1 to 3. Supplements were fed individually twice daily after milking. Cows received 6 kg concentrate/day in period 1, increased by 1 kg/day as barley, sorghum or lucerne chaff in each of periods 2 and 3. The Control treatment received 6 kg barley concentrate in all 3 periods. Milk yields by cows fed sorghum were lower than for cows fed equivalent levels of barley-based concentrate (P<0.05). Faecal starch levels (14, 18 and 17%) for cows fed sorghum concentrate were much higher (P<0.01) than those of cows fed similar levels of barley (2.1, 1.2 and 1.7%) in each period respectively. Additional supplementation as lucerne chaff did not increase milk production (P>0.05). Increased concentrate supplementation did not alleviate the problem of low protein in milk produced by freshly calved Holstein-Friesian cows grazing tropical grass pasture in summer. Animal production for a consuming world : proceedings of 9th Congress of the Asian-Australasian Association of Animal Production Societies [AAAP] and 23rd Biennial Conference of the Australian Society of Animal Production [ASAP] and 17th Annual Symposium of the University of Sydney, Dairy Research Foundation, [DRF]. 2-7 July 2000, Sydney, Australia.

Supply capability of sheep on the Mitchell grass downs of North and Central West Queensland

While, in the past, sheep have been predominantly reared and grazed in western Queensland for wool, interest in the sheep meat industry increased when wool prices became depressed. For north west and central west Queensland producers, opportunities may exist to participate in live sheep and meat export to Asia. The capability of the Mitchell grass downs to provide sufficient numbers of export quality sheep under the variable climatic conditions while sustaining the land resources has been simulated. Sheep numbers were found to be insufficient to maintain a consistent supply for live export. However, raising marking rates and lowering mortalities effectively increased reproductive performance to a level at which a surplus for export could be sustainable. Other practices might be required for the liveweight specifications to be met. 24th Biennial Conference. Adelaide, South Australia.

Simulation of the supply capability of sheep on the Mitchell grass downs of north west Queensland

Sheep in western Queensland have been predominantly reared for wool. When wool prices became depressed interest in the sheep meat industry, increased. For north west Queensland producers, opportunities may exist to participate in live sheep and meat export to Asia. A simulation model was developed to determine whether this sheep producing area has the capability to provide sufficient numbers of sheep under variable climatic conditions while sustaining the land resources. Maximum capacity for sustainability of resources (as described by stock numbers) was derived from an in-depth study of the agricultural and pastoral potential of Queensland. Decades of sheep production and climatic data spanning differing seasonal conditions were collated for analysis. A ruminant biology model adapted from Grazplan was used to simulate pregnancy rate. Empirical equations predict mortalities, marking rates, and weight characteristics of sheep of various ages from simple climatic measures, stocking rate and reproductive status. The initial age structure of flocks was determined by running the model for several years with historical climatic conditions. Drought management strategies such as selling a proportion of wethers progressively down to two-tooth and oldest ewes were incorporated. Management decisions such as time of joining, age at which ewes were cast-for-age, wether turn-off age and turning-off rate of lambs vary with geographical area and can be specified at run time. The model is run for sequences of climatic conditions generated stochastically from distributions based on historical climatic data correlated in some instances. The model highlights the difficulties of sustaining a consistent supply of sheep under variable climatic conditions.

Protein supplementation and growth enhancer strategies in successive years to maximise growth of Brahman cross steers grazing sown grass pastures on brigalow lands

A strategy comprising a winter/spring protein supplement, rumen modifier and hormonal growth promotant (Compudose 400) was used in either the first year (Tl), second year (T2), or in both years (T1+2) following weaning in Brahman cross steers as a means of increasing liveweight gain up to 2.5 years of age. T2 produced the heaviest final liveweight (544.7 kg) and highest overall liveweight gain (366.7 kg), but these were not significantly different from T1 (538.6 kg; 360.9 kg), or T1+2 (528.7 kg; 349.3 kg). However, final liveweight and overall liveweight gains of T1 and T2 but not T1+2 were significantly greater than for untreated (C) steers (504.9 kg; 325.2 kg, both P < 0.05). Regardless of the strategy imposed, liveweight and liveweight gain were enhanced, however final liveweights in each treatment were below the preferred minimum target liveweight (570-580 kg) for premium export markets. Treatment in both years gave no benefit over treatment in 1 year only. 19th Biennial Conference. 5-9 July 1992. LaTrobe University, Melbourne.

Transcriptome analysis of Neotyphodium and Epichloë grass endophytes

Large-scale gene discovery has been performed for the grass fungal endophytes Neotyphodium coenophialum, Neotyphodium lolii, and Epichloë festucae. The resulting sequences have been annotated by comparison with public DNA and protein sequence databases and using intermediate gene ontology annotation tools. Endophyte sequences have also been analysed for the presence of simple sequence repeat and single nucleotide polymorphism molecular genetic markers. Sequences and annotation are maintained within a MySQL database that may be queried using a custom web interface. Two cDNA-based microarrays have been generated from this genome resource. They permit the interrogation of 3806 Neotyphodium genes (NchipTM microarray), and 4195 Neotyphodium and 920 Epichloë genes (EndoChipTM microarray), respectively. These microarrays provide tools for high-throughput transcriptome analysis, including genome-specific gene expression studies, profiling of novel endophyte genes, and investigation of the host grass–symbiont interaction. Comparative transcriptome analysis in Neotyphodium and Epichloë was performed

Biological control of parthenium (Parthenium hysterophorus) in Australian rangeland translates to improved grass production.

Biological control of parthenium, a major weed in grazing areas in Australia, was initiated in the mid 1970s. Since then, nine species of insects and two rust fungi have been introduced. Evaluation using pesticide exclusion at two sites (Mt. Panorama and Plain Creek) in Queensland, Australia, revealed that classical biological control had a significant negative effect on the target weed, but the impact varied between years. In this study, I quantified the effects of biological control of parthenium on grass production. Grass production declined with the increase in parthenium biomass. Significant increase in grass production due to biological control was observed, but only in 1 of 4 yr at Mt. Panorama and 2 of 4 yr at Plain Creek. At Mt. Panorama, there was a 40% increase in grass biomass in 1997 because of defoliation by Zygogramma bicolorata and galling by Epiblema strenuana. At Plain Creek, grass biomass increased by 52% in 1998 because of E. strenuana and by 45% in 2000 because of combined effects of E. strenuana and the summer rust Puccinia melampodii. This study provides evidence on the beneficial effects of biological control of parthenium in areas under limited grazing.

Diet quality and liveweight gain of steers grazing Leucaena-grass pasture estimated with faecal near infrared reflectance spectroscopy (F.NIRS)

Three drafts of Bos indicus cross steers (initially 178-216 kg) grazed Leucaena-grass pasture [Leucaena leucocephala subspecies glabrata cv. Cunningham with green panic (Panicum maximum cv. trichoglume)] from late winter through to autumn during three consecutive years in the Burnett region of south-east Queensland. Measured daily weight gain (DWGActual) of the steers was generally 0.7-1.1 kg/day during the summer months. Estimated intakes of metabolisable energy and dry matter (DM) were calculated from feeding standards as the intakes required by the steers to grow at the DWGActual. Diet attributes were predicted from near infrared reflectance spectroscopy spectra of faeces (F.NIRS) using established calibration equations appropriate for northern Australian forages. Inclusion of some additional reference samples from cattle consuming Leucaena diets into F.NIRS calibrations based on grass and herbaceous legume-grass pastures improved prediction of the proportion of Leucaena in the diet. Mahalanobis distance values supported the hypothesis that the F.NIRS predictions of diet crude protein concentration and DM digestibility (DMD) were acceptable. F.NIRS indicated that the percentage of Leucaena in the diet varied widely (10-99%). Diet crude protein concentration and DMD were usually high, averaging 12.4 and 62%, respectively, and were related asymptotically to the percentage of Leucaena in the diet (R2 = 0.48 and 0.33, respectively). F.NIRS calibrations for DWG were not satisfactory to predict this variable from an individual faecal sample since the s.e. of prediction were 0.33-0.40 kg/day. Cumulative steer liveweight (LW) predicted from F.NIRS DWG calibrations, which had been previously developed with tropical grass and grass-herbaceous legume pastures, greatly overestimated the measured steer LW; therefore, these calibrations were not useful. Cumulative steer LW predicted from a modified F.NIRS DWG calibration, which included data from the present study, was strongly correlated (R2 = 0.95) with steer LW but overestimated LW by 19-31 kg after 8 months. Additional reference data are needed to develop robust F.NIRS calibrations to encompass the diversity of Leucaena pastures of northern Australia. In conclusion, the experiment demonstrated that F.NIRS could improve understanding of diet quality and nutrient intake of cattle grazing Leucaena-grass pasture, and the relationships between nutrient supply and cattle growth.