Year 1 (01/01/2003-31/12/2003)
A survey of farmers throughout all citrus growing regions of Sikkim sought to establish the relative importance of citrus production to the livelihood of local families, and the desire of people to grow this crop. Citrus has the potential to be the most profitable crop Sikkim farmers can grow, and they are eager to do so. However the tree decline problem has dramatically reduced yields and regions formerly dominated by citrus have gone completely out of production. The survey found that farmers were unable to do anything to combat the decline problem once it arrived in their area, and there was no evidence that the dramatic deterioration in tree health was in any way related to mismanagement. Some orchards with normal productivity were identified, and their superior performance might be explained in terms of clean planting material and isolation from infected areas.
A broad genetic base has been established in Sikkim, by the Sikkim Development Foundation, using material supplied from Queensland's citrus germplasm collection. More than 100 scion and rootstock genotypes were supplied as seed and grown in insect proof conditions at Tadong, Sikkim, where they have passed Indian quarantine requirements, and been distributed to different regions of the State. Their performance will be assessed over the coming years, while remaining material will be incorporated into rootstock and disease management experiments.
Molecular techniques were used to confirm the widespread distribution of the devastating citrus disease huanglongbing (formerly known as 'Greening'). Samples were collected from orchards and nurseries throughout the citrus growing belt of Sikkim and processed in the North Australia Quarantine Strategy laboratory in Mareeba, Queensland.
Rootstocks for Eureka lemon are being examined in an experiment planted near Bundaberg, Queensland. Material includes some recently developed hybrids as well as citrus relatives never previously tested as rootstocks. Excellent growth has been achieved with the first crop expected in early 2005. An Imperial mandarin rootstock experiment has also been propagated and is being field tested. It includes some Chinese germplasm introduced in a former ACIAR project, and never previously evaluated under subtropical conditions. In addition, this experiment incorporates new material from rootstock breeding in Australia, South Africa and USA. Graft compatibility experiments are underway in both Sikkim and Australia, aimed at identifying genera that might be useful in future disease management strategies.
Good nurseries are critical to any commercial citrus industry, and are needed to supply vigorous high health status trees to farmers. Sikkim collaborators are developing their expertise in growing citrus with poor quality potting media has been identified as a major constraint. Potential ingredients for improved potting media are readily available and preliminary experiments with these ingredients have been conducted.

The survey of citrus growing regions of Sikkim has been completed. From a technical viewpoint, this was aimed at determining the distribution of Huanglongbing (HLB) disease within commercial citrus orchards. Areas that had previously tested negative during the 2003 survey were revisited and retested, and furthermore all the remaining citrus growing areas were captured in the 2004 survey. Molecular analysis of samples collected during this work has shown that HLB disease is widespread in Sikkim, occurring in all citrus growing areas of the State.

A substantial nursery facility is now well established at Linjok and has been used to propagate trees for experiments outlined in the Project Document. Different species and varieties were budded with Sontala mandarin in July 2004 and these trees should be ready for planting in field experiment in late 2005 or early 2006. Sixteen different rootstock genotypes were propagated during 2004 for inclusion in these experiments.

The project has identified (through extensive field surveys and discussion with Sikkim farmers) six 'superior' individual trees growing in different parts of Sikkim. These show superior tree health and yield compared with neighbouring trees. Budwood was collected from each of these individual trees and propagated onto a standard rootstock to enable a replicated comparison of their performance. Budwood was also collected from a tree known to be infected with HLB (project sampling in 2003) and from material derived from shoot-tip-grafting. All of this propagation was conducted at the Linjok nursery in 2004 and trees are being maintained there (under individual insect proof nets) until they are large enough for field planting. Diagnostic samples were also collected from each of these 'superior' trees at the time of budwood collection and molecular testing performed for HLB. Interestingly, all six of the selections proved to be free of HLB.

Germplasm, sent from Queensland as seed, has been established at seven sites around Sikkim. During 2004, these seedlings grew sufficiently to be ready for field planting in early 2005. Indeed at one site the trees have already been successfully established in the field. Once field-planted, the germplasm with be exposed to the prevailing conditions and can then be assessed for the presence of genetic tolerance to environmental (particularly disease) pressures.

As a results of the projects activities during 2004, a consensus is fast emerging amongst stakeholders as to the cause of citrus decline problems in Sikkim. This represents a major achievement of the project given that the problem has been recognised for at least 50 years but attributed to a host of different, and unsubstantiated, causes.

The lemon rootstock experiment established by the project has grown particularly well and first fruit matured during 2004. Yields and detailed fruit quality assessments were made in early 2005. The project has been interested in developing an objective way to assess skin texture, as this is an important determinant of lemon quality in international trade. Substantial progress was made during 2004 such that a prototype was ready for testing in time for the first harvest on the lemon rootstock experiment.

A large mandarin rootstock experiment (420 trees) was field planted late in 2004, on a commercial collaborators property in Gayndah. This experiment contains 33 rootstock treatments including germplasm introduced from China under a former ACIAR project operated by NSWDPI. Legal agreements were finalised to protect intellectual property rights prior to the trees being planted.

A wide range of citrus species and relatives were screened for their susceptibility to citrus canker disease. Although this had always been as an experiment within the project, it took on increased urgency and relevance with the disastrous establishment of this exotic disease in commercial orchards at Emerald in June 2004. The work was successfully completed utilising the high security PC3 facility at AQIS Brisbane, and leaf samples collected predominantly from the QDPI&F germplasm collection at Bundaberg. It has substantially expanded the knowledge of host range of this disease and demonstrated the need for more screening work to occur.

The completion of disease survey work and propagation of nursery trees for field experiments in Sikkim, and the establishment of two large rootstock experiments and canker germplasm screening in Australia, indicate the success of project activities in 2004 in delivering outcomes of relevance and benefit to both countries.

Sikkims first semi-commercial citrus nursery facility became operational during 2005 as a direct consequence of the project. While primarily intended for propagating trees for the various experiments in the project, the Sikkim collaborators were eager to see the facility providing budded trees to local farmers. This has never happen in Sikkim before as all trees supplied to farmers have only been seedlings, which take more than 7 years to start fruiting where as budded trees fruit within 3 years. Experimental trees budded in 2004 were successfully maintained at the Linjok nursery facility and during 2005 reached sufficient size to be ready for field planting. Trees in the budwood-source-experiment were maintained in near-insect-proof conditions and treated with systemic insecticide to reduce chances of changing disease status during the nursery phase. Diagnostic samples were collected from these trees just prior to field planting to ensure that their disease status had remained the same as the original source material.

Visits to commercial citrus orchards in Sikkim in late 2005 further demonstrated the devastating nature of Huanglongbing disease, its long-term socio-economic impacts, and the need for real scientific solutions rather than workshops and training sessions which raise expectations but can offer no means of addressing the disease impacts. The project team re-visited an orchard that had been inspected in 2003, at which time there had been clear signs of disease, but the orchard was still productive and the farmer was proudly photographed in front of her best tree. Arriving at the property in 2005, all trees were unproductive and the once prosperous farmer now dejected and facing economic ruin. The training course that the farmer had attended at the National Citrus Centre during the intervening period had done nothing to avert the impending disaster. It is only from technology developed in research project like this one that there may be an opportunity for successful citrus production in Huanglongbing endemic areas.

In Australia, a Eureka lemon rootstock experiment established to identify improved genetics for subtropical lemon production has produced substantial quantities of fruit despite the young age of the trees. These fruit were subject to detailed assessment to determine what effect rootstock has on commercially important quality traits. In the absence of diseases like Huanglongbing, yield is seldom a limitation in subtropical citrus production and it is factors like fruit size, juice content and skin texture that affect the marketability and profitability of lemon growing. Australian lemons are often seen as poor quality on the international market. The rootstock experiment established in this project is beginning to reveal opportunities to increase the market attractiveness of Australian lemons.

With more than 30 different varieties, the Imperial rootstock experiment established by this project represents the most diverse range of germplasm ever tested under mandarins in Australia. Imperial is the main domestic mandarin variety and the vast majority of production occurs in the subtropics. Previous work by the project team has demonstrated the potential of rootstock genotype to overcome some of the quality problems associated with Imperial mandarin, and this new experiment made possible by the ACIAR project is building on this experience. Trees are now well established in the field and will produce their first crop of fruit in 2007. However, the work has made some important discoveries even before fruit production commences. The experiment was designed to incorporate germplasm that has performed well in recent overseas research, and as such the citrange variety 'Savage' was included. However our experiment has discovered within 12 months of field planting that 'Savage' is highly susceptible to local strains of Citrus Tristeza Virus and Imperial trees on this rootstock show severe stem pitting, stunting and tree death. Such results demonstrate the commercial importance of testing germplasm under local disease conditions, a philosophy which has been central to project work in both Sikkim and Australia.

Citrus canker disease has resulted in the recent destruction of more than half a million citrus trees in commercial orchards in an attempt to eradicate the disease from Australia. Research work from this ACIAR project has aided this campaign by providing crucial information on the susceptibility of native Rutaceae species to the disease. Without such information native species may unknowingly harbour the disease and act as a source of re-infection. The ability of project members to work effectively as a team and bring together a range of diverse expertise enabled the project to address the interaction between pathogen, host and disease development. This information was then utilised in the eradication campaign to target susceptible species.