Grasses form the basis for the meat, dairy, wool and deer industries, which contribute to nearly 50% of New Zealand exports, and are also an important component of natural ecosystems. Worldwide >100 plant viruses infect grass species and even mild and symptomless infections can adversely effect plant populations through reduced reproductive rates and greater susceptibility to environmental extremes. The only previously published study on viruses in New Zealand’s natural grasslands found that cereal viruses have invaded the native grass flora of the South Island. This research provided an extensive survey of New Zealand native grasses, showing that barley yellow dwarf virus diseases (BYDV, Luteoviridae) and Cocksfoot mottle virus (CoMV, Sobemovirus) are widespread in the North and South islands of New Zealand. Significant findings include seven new virus hosts amongst the New Zealand native flora, the first report of BYDV-PAS in New Zealand, detection in Hierochloe redolens of a novel virus in the Luteoviridae family (proposed name BYDV-To), and in Festuca novae-zelandiae a novel dsRNA virus possibly belonging to the Partitiviridae family. New virus host reports in New Zealand include CoMV in Poa anceps, P. cita, F. novae-zelandiae, and Chionochloa rubra; BYDV-PAV and BYDV-PAS in Microlaena stipoides and Dichelacne crinita; BYDV-MAV in P. cita, F. novae-zelandiae and H. redolens; and CYDV-RPV in P. cita and M. stipoides. Molecular techniques for virus detection and identification were developed or improved during this study. Phylogenetic analyses of viral coat protein sequences from native and exotic grass species indicate either frequent or recent virus movement into native ecosystems, and multiple virus introduction events in New Zealand. The likely origins of the virus species are discussed. Two CoMV variants were identified, one of which caused severe necrosis in susceptible cocksfoot cultivars. Reciprocal aphid transmission of BYDV-PAV using cereals and native grasses showed that although transmission to natives was low, the efficiency of transmission from natives to cereals was comparable to that between cereal species, suggesting virus adaptation to the cereal host species. The findings from this study are discussed in respect to disease management and bio-security in New Zealand, and recommendations are made for future research. / Whole document restricted, but available by request, use the feedback form to request access.

Lolium perenne cv. Nui is a cultivar of ryegrass grown throughout New Zealand in pastures due to favourable traits such as high palatability for livestock and its ability to withstand intensive grazing. However, the productivity of pastures is reduced when levels of aluminium and other metals accumulate in soils to toxic levels, a phenomenon referred to as the ‘acid soil syndrome’. In response to this toxicity, plants activate a series of antioxidant reactions, with one catalysed by the superoxide dismutase (SOD) enzymatic system. The enzyme system comprises three isoenzymes, a Cu/ZnSOD, FeSOD and a MnSOD which catalyse the same reaction but differ in amino acid sequence, molecular mass and the metal ion co-factor (hence Cu/ZnSOD, FeSOD and MnSOD). Together these isoenzymes combat the damaging effect of superoxide radicals which accumulate due to metal toxicity. In this thesis, the isolation of genes encoding isoenzymes of the SOD enzyme from L. perenne cv. Nui is described. As well, the growth of L. perenne cv. Nui and changes in expression of the SOD genes encoding each isoenzyme in response to aluminium treatment (0.2mM AlCl3) is investigated. A 1072 bp FeSOD gene sequence and a 705 bp MnSOD gene sequence were isolated from shoot tissue of L. perenne cv. Nui using a combination of RT-PCR with degenerate primers and 3'-RACE. The FeSOD gene comprised 572 bp of the coding sequence and 500 bp of 3'-UTR while the MnSOD gene comprised 508 bp of coding sequence and a 197 bp 3'-UTR. By alignment of each sequence with the gene from the database with highest identity it was predicted that the translation start codon (ATG) is located a further 196 bp upstream for the FeSOD gene (aligned with an Oryza sativa FeSOD sequence as a reference) and a further 152 bp upstream for the MnSOD sequence (aligned with a Triticum aestivum MnSOD sequence as a reference). Using RT-PCR with degenerate primers, a 313 bp CuSOD sequence was predominantly cloned from shoot tissue of L. perenne cv. Nui, but it was not possible to generate the 3'-UTR using 3'-RACE. For growth analysis, seedlings of L. perenne cv. Nui were germinated and acclimatised in Hoagland’s solution, and then subjected to either aluminium treatment (0.2mM AlCl3) or no treatment to act as a control over a designated time course of 0, 4, 8, or 24 hours. Two growth trials were conducted that differed in the age of seedlings used and plant tissues were separated into root and shoot tissues. Similar growth trends were observed in both trials, but the sampling regime in the second growth trial meant that statistical analysis could be carried out. In this trial, analysis revealed that over a time course of 24 hours exposure to 0.2mM aluminium, both root and shoot tissue fresh weight did not significantly differ when compared to the control (no aluminium). A general trend of an increase in root and shoot fresh weight was observed in plants treated with aluminium, but this trend was not significant at P=0.05. No significant change in fresh weight partitioning from shoot to root, or root to shoot in response to aluminium was also observed. Using semi-quantitative Reverse Transcriptase-Polymerase Chain Reaction (sqRTPCR) and primers based around the 3'-UTR with RNA isolated from plants grown in the second hydroponic trial, it was determined that under the conditions used, expression of the FeSOD and MnSOD genes isolated in this study were neither up-regulated or downregulated in response to aluminium treatment in both shoot and root tissue. Further, using degenerate primers to detect expression of one or more genes encoding the Cu/ZnSOD isoenzyme, total expression of the Cu/ZnSOD isoenzyme was also unresponsive to aluminium treatment.

Lolium perenne cv. Nui is a cultivar of ryegrass grown throughout New Zealand in pastures due to favourable traits such as high palatability for livestock and its ability to withstand intensive grazing. However, the productivity of pastures is reduced when levels of aluminium and other metals accumulate in soils to toxic levels, a phenomenon referred to as the ‘acid soil syndrome’. In response to this toxicity, plants activate a series of antioxidant reactions, with one catalysed by the superoxide dismutase (SOD) enzymatic system. The enzyme system comprises three isoenzymes, a Cu/ZnSOD, FeSOD and a MnSOD which catalyse the same reaction but differ in amino acid sequence, molecular mass and the metal ion co-factor (hence Cu/ZnSOD, FeSOD and MnSOD). Together these isoenzymes combat the damaging effect of superoxide radicals which accumulate due to metal toxicity. In this thesis, the isolation of genes encoding isoenzymes of the SOD enzyme from L. perenne cv. Nui is described. As well, the growth of L. perenne cv. Nui and changes in expression of the SOD genes encoding each isoenzyme in response to aluminium treatment (0.2mM AlCl3) is investigated. A 1072 bp FeSOD gene sequence and a 705 bp MnSOD gene sequence were isolated from shoot tissue of L. perenne cv. Nui using a combination of RT-PCR with degenerate primers and 3'-RACE. The FeSOD gene comprised 572 bp of the coding sequence and 500 bp of 3'-UTR while the MnSOD gene comprised 508 bp of coding sequence and a 197 bp 3'-UTR. By alignment of each sequence with the gene from the database with highest identity it was predicted that the translation start codon (ATG) is located a further 196 bp upstream for the FeSOD gene (aligned with an Oryza sativa FeSOD sequence as a reference) and a further 152 bp upstream for the MnSOD sequence (aligned with a Triticum aestivum MnSOD sequence as a reference). Using RT-PCR with degenerate primers, a 313 bp CuSOD sequence was predominantly cloned from shoot tissue of L. perenne cv. Nui, but it was not possible to generate the 3'-UTR using 3'-RACE. For growth analysis, seedlings of L. perenne cv. Nui were germinated and acclimatised in Hoagland’s solution, and then subjected to either aluminium treatment (0.2mM AlCl3) or no treatment to act as a control over a designated time course of 0, 4, 8, or 24 hours. Two growth trials were conducted that differed in the age of seedlings used and plant tissues were separated into root and shoot tissues. Similar growth trends were observed in both trials, but the sampling regime in the second growth trial meant that statistical analysis could be carried out. In this trial, analysis revealed that over a time course of 24 hours exposure to 0.2mM aluminium, both root and shoot tissue fresh weight did not significantly differ when compared to the control (no aluminium). A general trend of an increase in root and shoot fresh weight was observed in plants treated with aluminium, but this trend was not significant at P=0.05. No significant change in fresh weight partitioning from shoot to root, or root to shoot in response to aluminium was also observed. Using semi-quantitative Reverse Transcriptase-Polymerase Chain Reaction (sqRTPCR) and primers based around the 3'-UTR with RNA isolated from plants grown in the second hydroponic trial, it was determined that under the conditions used, expression of the FeSOD and MnSOD genes isolated in this study were neither up-regulated or downregulated in response to aluminium treatment in both shoot and root tissue. Further, using degenerate primers to detect expression of one or more genes encoding the Cu/ZnSOD isoenzyme, total expression of the Cu/ZnSOD isoenzyme was also unresponsive to aluminium treatment.

Festuca novae-zelandiae (Hack.) Cockayne is an endemic New Zealand perennial tussock forming grass of the family Poaceae. Morphologically F. novae-zelandiae exhibits a number of leaf adaptations associated with dehydration postponement as reflected in the climatic distribution of this species and its occurrence as a physiognomic dominant grass in semi arid short-tussock grasslands. Biochemical studies into the drought tolerance of this species have indicated the occurrence of tissue specific responses with respect to abscisic acid (ABA) and proline accumulation and protein turnover suggestive of a preferential protection of the tiller base and associated meristematic zones at the expense of lamina tissues. Further tissue specific biochemical responses to water-deficit stress in F. novae-zelandiae have been investigated. Changes in water-soluble carbohydrates (WSC) were monitoured over a 49-day dry-down period (decline in soil water content from 30% to 4%) in consecutive leaf segments comprising the leaf base (meristem region), elongation zone, the enclosed and exposed lamina, as well as basal sheath segments from the two next oldest leaves. In fully hydrated leaf tissues polymers of fructose (fructans) were the main WSC present and were mainly low molecular weight fructans of the inulin and neokestose series with the average degree of polymerization (DP) of fructan pools from 6 to 9. The highest fructan concentrations were present towards the leaf base. Fructan concentrations decreased over the course of the dry-down, although remained significantly higher in the meristem region of the tiller base with respect to any other tissue, until tissue water content fell below 45%. By day 49 of the dry-down period, the average DP of the fructan pool in tissues was from 3 to 5. Sucrose content increased in each tissue during the course of the dry-down, and was highest at the leaf base, where a concentration of 200 µmol g-1 dry weight was measured after 49 days of dry-down. The negative correlation between fructan and sucrose content, which indicates an inter-conversion dependent on tissue water content, suggests that, in this specises, fructans serve as a carbohydrate storage pool, while sucrose stabilises the meristem during extreme water deficit.

Repeat purchase of kiwifruit is primarily driven by consumer judgement of internal fruit quality attributes, including those affected by dry matter concentration (DMC) and mineral composition in fruit. This research investigated mechanisms affecting carbohydrate, mineral and water accumulation in 'Hayward' kiwifruit (Actinidia deliciosa), and related these to specific management practices. Canopy manipulation through pruning and treatments such as artificial pollination, defoliation, girdling, thinning and application of the auxin transport inhibitor TIBA, may affect fruit DMC and mineral composition. Leaf photosynthesis and fruit dry matter concentrations (DMC) started to decline as leaf area index values increased above 3-4. In addition to reducing competition for carbohydrates between vegetative and reproductive growth, leader pruning probably increased DMCs of fruit in the leader zone by improving light interception. Photosynthesis was not affected by crop loads between 20- 60 fruit m-2, but was consistently higher on non-terminating (long) shoots than on terminating (short) shoots, as were fruit DMCs. Differences in photosynthetic rate of leaves on these two shoot types were attributed to differences in shoot exposure to the sun, and also to the greater demand for carbohydrate within long shoots. Leaves subtending fruit may increase Ca, and to a lesser extent Mg, flow into fruit, however their accumulation was not affected by leaves outside the fruiting shoot. Xylem sap Ca and Mg concentrations were higher in shoots with a high rather than a low leaf: fruit (L:F) ratio and this may, at least partially, relate to the increase in shoot transpiration that occurs as shoot L:F ratios increase. Within vine variation in fruit Ca concentrations may reflect variations in xylem sap flow rates and Ca concentrations of xylem sap reaching fruit. Calcium translocation may occur independently of ion movement in the transpiration stream. Timing and extent of vascular differentiation in flower and fruitlet pedicels, possibly regulated by auxin, may influence fruit Ca accumulation. It is likely that early differentiation of vascular tissue in flower and fruitlet pedicels influenced cell division and subsequent (carbohydrate) sink strength of fruit by determining availability of carbohydrate for partitioning into cell walls. While growers have the potential to induce minor changes in fruit DMC, further increases will depend on the separation of carbohydrate and water accumulation. Further research is required to elucidate the mechanisms regulating phloem transport and unloading of sucrose in kiwifruit.

This research investigated the rhizobia that are associated with New Zealand legume plants. Rhizobia are a diverse group of bacteria that live in symbiosis with legumes in root nodules. Rhizobia fix Nitrogen from the atmosphere and provide this nutrient to the plant. The objectives of this research were to: 1) Determine the identity of the rhizobial species nodulating the native legumes of New Zealand: Sophora (kowhai), Carmichaelia (NZ broom), and Clianthus (kakabeak); and the identity and origin of rhizobial species nodulating invasive exotic legumes in New Zealand: Ulex (gorse), Cytisus (broom), and Acacia (wattles). 2) Determine the specificity and nitrogen fixing capacity of both groups of rhizobia. 3) Investigate the possible exchange of transmissible symbiotic genetic elements. A polyphasic strategy was used to determine the identity of bacterial isolates. The 16S rRNA, atpD, recA, and glnII genes were PCR amplified and sequenced, then analysed by maximum likelihood and Bayesian methods. Phenotypic characters were also assessed by use of the Biolog and FAME techniques. Nodulation and fixation ability was assessed by inoculating legume seedlings with rhizobial strains, then determining nitrogenase activity after ten weeks by gas chromatography, and examining roots for nodules. A gene involved in symbiosis, nodA, was sequenced from rhizobial strains to determine if transmission between strains had occurred. The results of the experiments showed that the native legumes were predominately nodulated by diverse Mesorhizobium spp. that contain three different nodA genotypes (two of which are novel) that have transferred between rhizobial strains. The Mesorhizobium spp. showed little nodulation specificity and could nodulate an exotic legume Astragalus (milk vetch), but not the invasive weed legumes. Rhizobium leguminosarum was also found to nodulate native legumes, albeit ineffectively. The exotic invasive woody legumes of this study were nodulated by diverse Bradyrhizobium spp. that had nodA genotypes typical of Australian and European species. The origins of these bacteria can not be categorically determined. However the evidence is presented to suggest that nodulating Mesorhizobium spp. arrived with the ancestors of the native legumes, while Bradyrhizobium spp. nodulating Ulex and Cytisus arrived recently from Europe. Bradyrhizobium spp. nodulating Acacia may be recently introduced, possibly from Australia, although further work is required to confirm these hypotheses. / This study was supported by a grant from the Marsden Fund of the Royal Society of New Zealand, under contract 97-LAN-LFS-002, and a grant from the Non-Specific Output Fund of Landcare Research.

The analysis of presence/absence data of plant species by regression analysis is the subject of this thesis. A nonparametric approach is emphasized, and methods which take into account correlations between species are also considered. In particular, generalized additive models (GAMs) are used, and these are applied to species’ responses to greenhouse scenarios and to examine multispecies interactions. Parametric models are used to estimate optimal conditions for the presence of species and to test several niche theory hypotheses. An extension of GAMs called vector GAMs is proposed, and they provide a means for proposing nonparametric versions of the following models: multivariate regression, the proportional and nonproportional odds model, the multiple logistic regression model, and bivariate binary regression models such as bivariate probit model and the bivariate logistic model. Some theoretical properties of vector GAMs are deduced from those pertaining to ordinary GAMs, and its relationship with the generalized estimating equations (GEE) approach elucidated. / Whole document restricted, but available by request, use the feedback form to request access.

Whole document restricted, see Access Instructions file below for details of how to access the print copy. / This work constitutes a detailed study of the spores, of the ferns and fern allies, both native and adventive, which grow in New Zealand, from the Kermadec Islands in the North, to the Chatham Islands in the East and the Subantarctic Islands in the South. Twenty live families with sixty five genera, two hundred and eleven species (of which c.20 are introduced) and three subspecies are included. Seven species are heterosporous the remainder are homosporous. Trilete spores are found in c.104 species representing c.30 genera. Perine is present in most taxa (with the exception of Gleichenia) and ranges from the large and sac-like form as seen in Cyathea smithii, to the thin and reduced form seen in Adiantum. Monolete spores are found in c.109 species representing c.37 genera (two genera are included twice, Isoetes which has trilete megaspores and monolete microspores and Lindsaea which has both monolete and trilete species). Perine is present in most taxa (except Sticherus spp.) and ranges from a fine deposit as seen in the Psilotaceae, to an enlarged form, heavily winged, as seen in the Aspleniaceae. Sculpture in all taxa (with the exception of the Lycopodiaceae and some members of the Ophioglossaceae, which may have ornamentation distributed distally), is present on both distal and proximal faces. One adventive taxon Equisetum arvense L. has an unusual circular laesura and elaborate elaters. Light micrographs of acetolysed and fresh spores along with scanning electron micrographs, are included for each taxon. Keys presented, are based on gross spore morphology and are applicable to fresh and acetolysed material. Descriptions include a list of synonymous species, details of spore shape, laesura/ae details, perine/exine sculpture and thickness (where sections allow), size (measured from n=50 spores per population), for samples treated with cotton blue (lactophenol aniline blue formula), mounted in glycerine jelly and acetolysed samples, mounted in silicone oil. Percentage size differences for acetolysed material mounted in glycerine jelly are also noted, along with previous spore dimensions recorded in the literature. Descriptions also contain chromosome number, where known, a list of previously recorded descriptions, a brief indication of geographic location and a list of vouchers for each sample. Experiments on the size and morphology of fern spores in reaction to different preparation techniques are discussed. Fresh spores of seven species, four trilete (Adiantum fulvum, Cyathea smithii, Hymenophyllum flabellatum and Lycopodium volubile) and three monolete (Blechnum chambersii, Paesia scaberula and Tmesipteris elongata), were assessed wth regard to the effect of three standard pretreatments (lactophenol aniline blue, 10% KOH, acetolysis) and two mounting media (glycerine jelly and silicone oil). Changes in morphology and size of the various wall layers were noted in comparison to spores observed fresh in water. Results indicate that variation includes shrinkage e.g. silicone oil and expansion of both exine and perine in glycerine jelly. Both effects are modified by previous treatments.

Pterocladia lucida (R. Br.) J. Ag. is the prinipal raw material for agar production in New Zealand. Since collection commenced in 1943 the annual harvest has never stabilised and over recent years there has been a general decline, which has inhibited expansion of the indigenous industry. Field and laboratory studies have been made in order to obtain information on the biology of P. lucida. Form variability is considered in detail and a relationship between external morphology and wave exposure of the habitat is proposed. The culture of P. lucida from spores in a running seawater system is described in detail. Results on carpospore and tetraspore germination, and sporeling growth under light and temperature regimes are presented. Germination of both spore types was inhibited at 10°C while at higher temperatures germination was greatest under 4 klux. Spore germination and sporeling development in culture is described. Only sterile prostrate thalli possessing rhizoidal haptera were obtained and a hypothesis relating the appearance of surface haptera to the direction of irradiation is proposed. The development of an outdoor wave tank for culturing is described and the growth rate and development of sporelings is discussed with reference to water movement. Sporelings were maintained in culture for up to 22 months. Cultured sporelings have also been transplanted into different habitats. Creeping axes established from carpospores produced erect fronds, which produced tetraspores after seven to ten months in situ. This confirmed part of a Polysiphonia-type life history for the first time in a Gelidioid species. Axis elongation rate in situ was estimated to be less than 10cm year-1 at Leigh. It was concluded that the collection of attached weed could not be sustained annually, but controlled harvesting in specific areas every two years is advocated. Seasonal variation in agar yield and gel strength from wave exposed and sheltered populations of P. lucida was studied. Agar levels varied throughout the year, being greatest in spring and summer, while gel strengths showed a similar trend. The feasibility of utilising indigenous Gracilaria secundata Harvey forma pseudoflagellifera May has been studied. The yield and gel properties of agar were determined from different alkali pretreatments. There was a marked increase in gel strength when weed was pretreated in alkali, the maximum gel strength being obtained from a weed treatment of 3 percent NaOH for three hours at 80°C. The gel properties of agar from G. secundata f. pseudoflagellifera are compared with agar from other sources. The agar was suitable for commercial use, but exhibited a greater resilience than that from Pterocladia lucida. The vegetative regeneration of G. secundata f. pseudoflagellifera in the Manukau Harbour has been studied and an experimental harvest of 0.68t is described. During spring 100 percent regeneration from hand cleared and raked areas occurred after eight to thirteen weeks. Based on regeneration results, mechanical harvesting four times a year of Gracilaria mats in the Manukau Harbour is advocated. The predicted sustainable yield is 1084-1304t year-1. The extraction and gel properties of agar from Melanthalia abscissa (Turn.) Hook. et Harv. are reported for the-first time, but the commercial collection of this species is not advocated.

The role of, and response to, disturbance, in the dynamics of the mountain beech forest at western Mt Ruapehu, was examined, using palynology, dendrochronology, and vegetation survey. The pollen record indicates that, two thousand years ago, the west Ruapehu forest was dominated by Nothofagus solandri. About 1800 years B.P., the eruption of Lake Taupo devastated the forest. Though Halocarpus spp. and Phyllocladus asplenifolius were important early colonizers, Nothofagus solandri re-established itself close to its present limits after the eruption. Fire, from about 650 years ago onwards, did not affect the beech forest, but did affect the coniferous vegetation on the ring plain near the mountain. The age structure of the forest shows that there was a large disturbance event shortly before 1740. A dieback occurring about 1969 appears to have been the largest episode of mortality since 1740, larger than the dieback episode described by Cockayne at the beginning of this century. Dieback occurred as a short peak-period of Nothofagus solandri death about 1969. It was the large trees of the mature cohort which tended to be killed by dieback rather than the smaller individuals. Tree-ring analysis, showed that narrow tree-rings occurred in beech in the 1960's. Extreme rainfall years in the early 1960's may have put the beech trees under stress, making them susceptible to dieback. The drought in 1969 may have precipitated extensive mortality. The characteristics of N. solandri rings from Ruapehu are similar to ring characteristics of that species from South Island studies. The 1982 cyclone had a noticeable impact on the forest, affecting some areas severely. However, the impact was relatively minor compared to the influence of dieback. The mean density of trees >= 10cm dbh was 520 stems/ha., of which 220 stems/ha. were Nothofagus solandri. The mean basal area was 22m2 /ha., of which 11m2 /ha. was N. solandri. The basal area is very low compared to that in other N.Z. forests. N. solandri, Griselinia littoralis, Phyllocladus asplenifolius, Podocarpus hallii, and Libocedrus bidwillii, made up 35, 22, 16, 9, and 8 percent, respectively, of the total density. Basal area, density, and species composition varied as much within sites, as between them. The shrub layer formed a large part of the vegetation at west Ruapehu. Coprosma species were particularly abundant. There is sufficient regeneration to indicate that a new cohort of N. solandri is becoming established in most areas. Some areas will probably remain in shrubland for many decades. The result of dieback and windthrow has been to virtually eliminate the canopy of large old beech trees, and to increase the heterogeneity of stand structure. N. solandri seedling densities are not as high as those found in many South Island beech forests. Nothofagus solandri is growing faster than are most of the co-existing tree species, and it appears likely that it will continue as the dominant tree species. Implications for forest management are discussed.