Genetic uniformity of international isolates of Leifsonia xyli subsp xyli, causal agent of ratoon stunting disease of sugarcane

An international collection of the sugarcane ratoon stunting disease pathogen, Leifsonia xyli subsp. xyli, was analysed to assess genetic diversity. DNA fingerprinting using BOX primers was performed on 105 isolates, comprising 65 Australian isolates and an additional 40 isolates from Indonesia (n = 8), Japan (n = 1), USA (n = 3), Brazil (n = 2), Mali (n = 2), Zimbabwe (n = 13), South Africa (n = 9) and Reunion (n = 2). Sixty-two of these isolates were also screened using ERIC primers. No variation was found among any of the isolates. The intergenic spacer (IGS) region of the ribosomal RNA genes from 54 isolates was screened for sequence variation using single-stranded conformational polymorphism (SSCP), but none was observed. Direct sequencing of the IGS from a subset of nine isolates, representing all of the countries sampled in this study, confirmed the results of the SSCP analysis. Likewise, no sequence variation was found in the 16S ribosomal RNA genes of the same subset. Four Colombian isolates from sugarcane, morphologically similar to L. xyli subsp. xyli, were putatively shown to be an undescribed Agrococcus species of unknown pathogenicity. The lack of genetic variation among L. xyli subsp. xyli isolates, independent of time of sampling, cultivar of isolation, or country of origin, suggests the worldwide spread of a single pathogenic clone, and further suggests that sugarcane cultivars resistant to ratoon stunting disease in one area should retain this property in other regions.

Development of PCR-based markers for detection of Leifsonia xyli subsp. xyli in fibrovascular fluid of infected sugarcane plants

DNA of Leifsonia xyli subsp. xyli (Lxx), the causal agent of ratoon stunting disease of sugarcane, was detected in the fibrovascular fluid of sugarcane plants using random amplified polymorphic DNA PCR-based amplification using two 10-mer oligonucleotide primers. The primers OPC-02 and OPC-11 produced Lxx-specific markers of approximately 800 bp and 1000 bp, respectively. A cloned DNA fragment from the 800 bp PCR product (pSKC2-800) hybridised to a single genomic DNA fragment from Lxx when used as a probe in Southern hybridisation. This cloned fragment did not hybridise to L. xyli subsp. cynodontis (Lxc), or L. xyli-like bacteria isolated from grasses in Australia, indicating the usefulness of this DNA fragment as a specific probe for Lxx. A cloned fragment from the 1000 bp PCR product ( pSKC11-1000) hybridised to three genomic fragments in Lxx isolates, one genomic fragment in two of the four isolates of L. xyli-like bacteria, and in two of the four isolates of Lxc isolated from the USA. These results indicate that L. xyli-like bacteria are more likely to be related to Lxc than Lxx. These probes did not hybridise to the DNA from strains of the species of Clavibacter, Rathayibacter, Acidovorax, Ralstonia, Pseudomonas and Xanthomonas tested. Two oligonucleotide primers (21-mer) designed from the pSKC2-800 sequences specifically amplified template DNA from Lxx and detected as few as 5 x 10(4) cells/mL in fibrovascular fluid from sugarcane plants infected with Lxx.

Fiji disease resistance in sugarcane: Relationship to cultivar preference in field populations of the planthopper vector Perkinsiella saccharicida

Populations of the planthopper vector Perkinsiella saccharicida on sugarcane cultivars resistant (cvs Q110 and Q87), moderately resistant (cvs Q90 and Q124) and susceptible (evs NCo310 and Q 102) to Fiji disease with known field resistance scores were monitored on the plant (2000-2001) and ratoon (2001-2002) crops. In both crops, the vector population remained very low, reaching its peak in the autumn. The vector population was significantly higher on cultivars susceptible to Fiji disease than on cultivars moderately resistant and resistant to Fiji disease. The number of R saccharicida adults, nymphs and oviposition sites per plant increased with the increase in the Fiji disease susceptibility. The results suggest that under low vector density, cultivar preference by the planthopper vector mediates Fiji disease resistance in sugarcane. To obtain resistance ratings in the glasshouse that reflect field resistance, glasshouse-screening trials should be conducted under both low and high vector densities, and the cultivar preference of the planthopper vector recorded along with Fiji disease incidence.

Effects of high temperature on the growth and composition of sugarcane internodes

Sugarcane grown in the Ord River district of Western Australia has lower sucrose content than expected from earlier trials and experience in other irrigated districts. High temperatures have been hypothesised as a possible cause. The effects of high temperature (above 32 degrees C) on growth and carbon partitioning were investigated. A temperature regime of (25-38 degrees C) was compared with (23-33 degrees C). In one experiment, 7-month-old plants of cvv. Q117 and Q158 were subjected to the treatments for 2 months. In another experiment, the plants were allowed to regrow (ratoon) for 6 months. In both experiments, the higher temperature resulted in more, shorter internodes and higher moisture content. Most internodes from plants in the higher temperature treatment had lower sucrose content than internodes from the lower temperature. On a dry mass basis the internodes from the plants in the higher temperature had proportionately more fibre and hexoses but lower sucrose. Combined with an increased number of nodes in a stem of similar or shorter length this would result in higher stalk fibre and lower sucrose content. The data provided evidence that sugarcane partitions less carbon to stored sucrose when grown under high compared with low temperatures. The two cultivars partitioned carbon between soluble (sugars) and insoluble (fibre) fractions to different degrees. These experiments also indicate that the current models describing leaf appearance and perhaps sugarcane growth at temperatures above 32 degrees C, in general, need revision.

Recent advances in the molecular biology of Leifsonia xyli subsp xyli, causal organism of ratoon stunting disease

Twelve years ago our understanding of ratoon stunting disease (RSD) was confined almost exclusively to diagnosis of the disease and control via farm hygiene, with little understanding of the biology of the interaction between the causal agent (Leifsonia xyli subsp. xyli) and the host plant sugarcane (Saccharum spp. hybrids). Since then, research has focused on developing the molecular tools to dissect L. xyli subsp. xyli, so that better control strategies can be developed to prevent losses from RSD. Within this review, we give a brief overview of the progression in research on L. xyli subsp. xyli and highlight future challenges. After a brief historical background on RSD, we discuss the development of molecular tools such as transformation and transposon mutagenesis and discuss the apparent lack of genetic diversity within the L. xyli subsp. xyli world population. We go on to discuss the sequencing of the genome of L. xyli subsp. xyli, describe the key findings and suggest some future research based on known deficiencies that will capitalise on this tremendous knowledge base to which we now have access.

Sugarcane moth borers (Lepidoptera: Noctuidae and Pyraloidea): phylogenetics constructed using COII and 16S mitochondrial partial gene sequences

Sugarcane moth borers are a diverse group of species occurring in several genera, but predominately within the Noctuidae and Pyraloidea. They cause economic loss in sugarcane and other crops through damage to stems and stalks by larval boring. Partial sequence data from two mitochondrial genes, COII and 16S, were used to construct a molecular phylogeny based on 26 species from ten genera and six tribes. The Noctuidae were found to be monophyletic, providing molecular support for the taxonomy within this subfamily. However, the Pyraloidea are paraphyletic, with the noctuids splitting Galleriinae and Schoenobiinae from the Crambinae. This supports the separation of the Pyralidae and Crambinae, but does not support the concept of the incorporation of the Schoenobiinae in the Crambidae. Of the three crambine genera examined, Diatraea was monophyletic, Chilo paraphyletic, and Eoreuma was basal to the other two genera. Within the Noctuidae, Sesamia and Bathytricha were monophyletic, with Busseola basal to Bathytricha. Many species in this study (both noctuids and pyraloids) had different biotypes within collection localities and across their distribution; however the individual biotypes were not phylogenetically informative. These data highlight the need for taxonomic revisions at all taxon levels and provide a basis for the development of DNA-based diagnostics for rapidly identifying many species at any developmental stage. This ability is vital, as the species are an incursion threat to Australia and have the potential to cause significant losses to the sugar industry.

Efficient developmental mis-targeting by the sporamin NTPP vacuolar signal to plastids in young leaves of sugarcane and Arabidopsis

Plant vacuoles are multi-functional, developmentally varied and can occupy up to 90% of plant cells. The N-terminal propeptide (NTPP) of sweet potato sporamin and the C-terminal propeptide (CTPP) of tobacco chitinase have been developed as models to target some heterologous proteins to vacuoles but so far tested on only a few plant species, vacuole types and payload proteins. Most studies have focused on lytic and protein-storage vacuoles, which may differ substantially from the sugar-storage vacuoles in crops like sugarcane. Our results extend the evidence that NTPP of sporamin can direct heterologous proteins to vacuoles in diverse plant species and indicate that sugarcane sucrose-storage vacuoles (like the lytic vacuoles in other plant species) are hostile to heterologous proteins. A low level of cytosolic NTPP-GFP (green fluorescent protein) was detectable in most cell types in sugarcane and Arabidopsis, but only Arabidopsis mature leaf mesophyll cells accumulated NTPP-GFP to detectable levels in vacuoles. Unexpectedly, efficient developmental mis-trafficking of NTPP-GFP to chloroplasts was found in young leaf mesophyll cells of both species. Vacuolar targeting by tobacco chitinase CTPP was inefficient in sugarcane, leaving substantial cytoplasmic activity of rat lysosomal beta-glucuronidase (GUS) [ER (endoplasmic reticulum)-RGUS-CTPP]. Sporamin NTPP is a promising targeting signal for studies of vacuolar function and for metabolic engineering. Such applications must take account of the efficient developmental mis-targeting by the signal and the instability of most introduced proteins, even in storage vacuoles.

Markers associated with stalk number and suckering in sugarcane colocate with tillering and rhizomatousness QTLs in sorghum

Two important factors influencing sugar yield, the primary focus of sugarcane plant breeding programs, are stalk number and suckering. Molecular markers linked to both of these traits are sought to assist in the identification of high sugar yield, high stalk number, low-suckering sugarcane clones. In this preliminary mapping study, 108 progeny from a biparental cross involving two elite Australian sugarcane clones were evaluated at two sites for two years for both stalk number and suckering. A total of 258 DNA markers, including both restriction fragment length polymorphisms (RFLPs) and radio-labelled amplified fragments (RAFs), were scored and evaluated using single-factor analysis. Sixteen (7 RFLPs and 9 RAFs) and 14 (6 RFLPs and 8 RAFs) markers were identified that were significantly associated (P < 0.01) with stalk number and suckering, respectively, across both years and sites. The seven and six RFLP markers associated with stalk number and suckering, respectively, were generated by eight different RFLP probes, of which seven had been mapped in sorghum and (or) sugarcane. Of significant interest was the observation that all seven RFLP probes could be shown to be located within or near QTLs associated with tillering and rhizomatousness in sorghum. This observation highlights the usefulness of comparative mapping between sorghum and sugarcane and suggests that the identification of useful markers for stalk number and suckering in sugarcane would be facilitated by focussing on sorghum QTLs associated with related traits.

Sugarcane water use from shallow water tables: implications for improving irrigation water use efficiency

The resource potential of shallow water tables for cropping systems has been investigated using the Australian sugar industry as a case study. Literature concerning shallow water table contributions to sugarcane crops has been summarised, and an assessment of required irrigation for water tables to depths of 2 m investigated using the SWIMv2.1 soil water balance model for three different soils. The study was undertaken because water availability is a major limitation for sugarcane and other crop production systems in Australia and knowledge on how best to incorporate upflow from water tables in irrigation scheduling is limited. Our results showed that for the three soils studied (representing a range of permeabilities as defined by near-saturated hydraulic conductivities), no irrigation would be required for static water tables within 1 m of the soil surface. Irrigation requirements when static water tables exceeded 1 m depth were dependent on the soil type and rooting characteristics (root depth and density). Our results also show that the near-saturated hydraulic conductivities are a better indicator of the ability of water tables below 1 m to supply sufficient upflow as opposed to soil textural classifications. We conclude that there is potential for reductions in irrigation and hence improvements in irrigation water use efficiency in areas where shallow water tables are a low salinity risk: either fresh, or the local hydrology results in net recharge. (C) 2003 Elsevier B.V. All rights reserved.

The evaluation of the spatial and temporal stability of sugarcane farm performance based on yield and commercial cane sugar

In broader catchment scale investigations, there is a need to understand and ultimately exploit the spatial variation of agricultural crops for an improved economic return. In many instances, this spatial variation is temporally unstable and may be different for various crop attributes and crop species. In the Australian sugar industry, the opportunity arose to evaluate the performance of 231 farms in the Tully Mill area in far north Queensland using production information on cane yield (t/ha) and CCS ( a fresh weight measure of sucrose content in the cane) accumulated over a 12-year period. Such an arrangement of data can be expressed as a 3-way array where a farm x attribute x year matrix can be evaluated and interactions considered. Two multivariate techniques, the 3-way mixture method of clustering and the 3-mode principal component analysis, were employed to identify meaningful relationships between farms that performed similarly for both cane yield and CCS. In this context, farm has a spatial component and the aim of this analysis was to determine if systematic patterns in farm performance expressed by cane yield and CCS persisted over time. There was no spatial relationship between cane yield and CCS. However, the analysis revealed that the relationship between farms was remarkably stable from one year to the next for both attributes and there was some spatial aggregation of farm performance in parts of the mill area. This finding is important, since temporally consistent spatial variation may be exploited to improve regional production. Alternatively, the putative causes of the spatial variation may be explored to enhance the understanding of sugarcane production in the wet tropics of Australia.

A methodology for analysis of sugarcane productivity trends - 2. Comparing variety trials with commercial productivity

Previous research has reported both agreements and serious anomalies in relationships between production attributes of sugarcane varieties in variety trials (VTs) and commercial production (CP). This paper examines VT and CP data for tonnes of cane per hectare (TCH) and sugar content (CCS). Data, analysed by REML, included 107 VTs and 54 CP mill years for 9 varieties from the mill districts of Mulgrave, Babinda, and Tully for harvest years 1982-99. Important consistencies included high TCH of Q152, high CCS of Q117 and Q120, and low CCS of H56-752. Significant anomalies existed with respect to TCH for Q113, Q117, Q120, Q122, Q138, and H56-752 and to CCS for Q113 and Q124. Investigation of these anomalies was assisted by access to independent REML analyses of CP data for 65692 individual Tully cane blocks from 1988 to 1999 and by the knowledge of persons familiar with the preferential uses of varieties by farmers. Minor anomalies were due to limited year or mill area data. Q124 TCH was deemed to be decreased and its CCS increased by severe disease in Babinda CP in the extremely wet 1998 and 1999 seasons. Other serious anomalies have credible but unsubstantiated explanations. The most convincing, for Q113, Q117, Q138, and H56-752, are that these varieties were deployed unevenly with regard to late season harvesting, predominant use or avoidance on high fertility soils, or use confined to low fertility sandy soils, respectively. Uneven deployment results in confounding of these effects in the varietal CP statistics at mill area level. It is concluded that VTs cannot be enhanced to anticipate or evaluate most effects of uneven deployment. They give adequate predictions of relative CP performance for varieties deployed evenly across confounding influences. Routine analyses of individual block CP data would be useful and enhanced by addition of relevant information to the block records.

Design, development, and use of molecular primers and probes for the detection of Gluconacetobacter species in the pink sugarcane mealybug

Molecular tools for the species-specific detection of Gluconacetobacter sacchari, Gluconacetobacter diazotrophicus, and Gluconacetobacter liquefaciens from the pink sugarcane mealybug (PSMB) Saccharicoccus sacchari Cockerell (Homiptera: Pseudococcidae) were developed and used in polymerase chain reactions (PCR) and in fluorescence in situ hybridizations (FISH) to better understand the microbial diversity and the numerical significance of the acetic acid bacteria in the PSMB microenvironment. The presence of these species in the PSMB occurred over a wide range of sites, but not in all sites in sugarcane-growing areas of Queensland, Australia, and was variable over time. Molecular probes for use in FISH were also designed for the three acetic acid bacterial species, and shown to be specific only for the target species. Use of these probes in FISH of squashed whole mealybugs indicated that these acetic acid bacteria species represent only a small proportion of the microbial population of the PSMB. Despite the detection of Glac. sacchari, Glac. diazotrophicus, and Glac. liquefaciens by PCR from different mealybugs isolated at various times and from various sugarcane-growing areas in Queensland, Australia, these bacteria do not appear to be significant commensals in the PSMB environment.

Resistance gene analogues in sugarcane and sorghum and their association with quantitative trait loci for rust resistance

Fifty-four different sugarcane resistance gene analogue (RGA) sequences were isolated, characterized, and used to identify molecular markers linked to major disease-resistance loci in sugarcane. Ten RGAs were identified from a sugarcane stem expressed sequence tag (EST) library; the remaining 44 were isolated from sugarcane stem, leaf, and root tissue using primers designed to conserved RGA motifs. The map location of 31 of the RGAs was determined in sugarcane and compared with the location of quantitative trait loci (QTL) for brown rust resistance. After 2 years of phenotyping, 3 RGAs were shown to generate markers that were significantly associated with resistance to this disease. To assist in the understanding of the complex genetic structure of sugarcane, 17 of the 31 RGAs were also mapped in sorghum. Comparative mapping between sugarcane and sorghum revealed syntenic localization of several RGA clusters. The 3 brown rust associated RGAs were shown to map to the same linkage group (LG) in sorghum with 2 mapping to one region and the third to a region previously shown to contain a major rust-resistance QTL in sorghum. These results illustrate the value of using RGAs for the identification of markers linked to disease resistance loci and the value of simultaneous mapping in sugarcane and sorghum.

Modelling, nitrogen dynamics in sugarcane systems: Recent advances and applications

Substantial amounts of nitrogen (N) fertiliser are necessary for commercial sugarcane production because of the large biomass produced by sugarcane crops. Since this fertiliser is a substantial input cost and has implications if N is lost to the environment, there are pressing needs to optimise the supply of N to the crops' requirements. The complexity of the N cycle and the strong influence of climate, through its moderation of N transformation processes in the soil and its impact on N uptake by crops, make simulation-based approaches to this N management problem attractive. In this paper we describe the processes to be captured in modelling soil and plant N dynamics in sugarcane systems, and review the capability for modelling these processes. We then illustrate insights gained into improved management of N through simulation-based studies for the issues of crop residue management, irrigation management and greenhouse gas emissions. We conclude by identifying processes not currently represented in the models used for simulating N cycling in sugarcane production systems, and illustrate ways in which these can be partially overcome in the short term. (c) 2005 Elsevier B.V. All rights reserved.