Honeybee - RIRDC Completed Projects in 1999-2000 & Research in Progress as at June 2001

This publication, Honeybee Research Report 2001, provides details of honeybee research from July 2000 until June 2001 and lists projects commencing in the 2001/2002 financial year. It follows the Honeybee Research and Development Council Research Report 1980-1995 and the RIRDC Reports 1995-1997, 1998, 1999 and 2000, which were a collection of final report and progress summaries of levy funded honeybee research until June 2000.

This report provides information to help apiarists and others access research recommendations and research in progress, together with researcher contact details, in a simple, easy to read format.

This report, a new addition to RIRDC’s diverse range of over 600 research publications, forms part of our Honeybee R&D program, which aims to improve the productivity and profitability of the Australian beekeeping industry

Most of our publications are available for viewing, downloading or purchasing online through our website:
downloads at www.rirdc.gov.au/reports/Index.htm
purchases at www.rirdc.gov.au/eshop

Alternatively, there is a RIRDC order form included on the last page of this publication.

Peter Core
Managing Director
Rural Industries Research and Development Corporation

Project Title Improving queen bee production

RIRDC Project No.: CSE-85A

Start Date: 01/07/99

Finish Date: 31/07/02

Researcher: Dr. Denis Anderson

Organisation: CSIRO Entomology
GPO Box 1700
CANBERRA ACT 2601

Phone: (02) 6246 4148

Fax: (02) 6246 4000

Email: Denis.Anderson@ento.csiro.au

Objectives To improve the profitability of commercial queen producers by improving queen performance. This will be achieved by:
1. Determining whether nutritional supplements significantly improve queen and drone quality;
2. Determining the factors which are responsible for causing disappearing disorder.

Current Progress Effects of nutrient supplements on queen cell-building colonies
This sub-project aims to determine whether the feeding of nutrient supplements during queen production improves the quality of adult queen bees. During the 1999/00 season, the following supplements were each fed to 5 queen cell-building colonies of equal strength: (a) no supplement (control), (b) irradiated pollen, (c) irradiated pollen with added vitamins, (d) soya-flour based supplement, (e) soya-flour based supplement with added vitamins and (f) vitamins. These test-supplements were those recommended by queen producers and participants at the ‘Honey Bee Nutrition Workshop’ held in Sydney in May 1998. Sister-queens were subsequently raised in the cell-builders, open mated, and tested for: (1) weight, (2) number of ovarioles, (3) number of spermatozoa in spermathecae, (4) nosema levels and (5) body protein.
No significant differences were found in the quality of adult queen bees raised from cell-builders fed the different diets. It was concluded that this unexpected result was due to the excellent forage conditions that prevailed in the local environment where the trials were conducted. It is planned to repeat these trials in the future when local forage conditions are poor. At the present time (during the latter part of the 2000/01 season), the effects of feeding nutrient supplements to queen banks is being assessed.
Disappearing Disorder
This sub-project aims to determine the cause of disappearing disorder which affects bee colonies at certain times of the year in North-East NSW and South-East Queensland.
During the 1999/2000 season, the disorder was mild and tests conducted on the few samples that were collected showed no evidence for the presence of pathogenic micro-organisms or excessive levels of toxic elements in the nectar and pollen being consumed by the affected larvae. During the 2000/01 season, the disorder was moderately severe at Cunningham’s gap in SE Queensland, but almost absent in the Gympie area to the north-east. Large numbers of samples of affected larvae were collected from an apiary at Cunningham’s Gap. Tests on these larvae again showed no evidence that the disorder resulted from the presence of pathogenic micro-organisms or toxic elements. Further tests are currently being conducted on the samples.

Project Title Introduction and early performance of queen bees – some factors affecting success

RIRDC Project No.: DAN-182A

Start Date: 01/09/99

Finish Date: 31/07/01

Researcher: Mr. John Rhodes

Organisation: NSW Department of Agriculture
RMB 944
Calala Lane
TAMWORTH NSW 2340

Phone: (02) 6763 1206

Fax: (02) 6763 1222

Email: Rhodesj@agric.nsw.gov.au

Objectives · To determine whether the age of the queen bee at introduction influences introduction success.
· To identify factors affecting sperm counts of sister queen bees mated under the same mating conditions and to investigate relationships between low sperm counts and queen failure following introduction.
· To examine the effects of temperature and humidity during transport of queen bees on their introduction success.
· A comparison of sperm counts between sister bees mated in a commercial queen bee breeder`s apiary and between those mated in a commercial honey producer`s queen bee breeding apiary.
· Development of a method for preserving adult drones which would allow accurate sperm counts to be carried out on those drones.

Current Progress Field and laboratory work for objectives 1 to 4 have been completed and relevant data are being analysed. Compared to the 1999-2000 season, seasonal conditions were harder on bees than the 2000-2001 season however higher numbers of queen bees survived for each age group for both the 14 day and 15 week survival counts. The trend of increased numbers of queens surviving corresponding to increased age queens were caught from their mating nucleus continued for both seasons.
Work on objective 5 is continuing, small trials in which whole drones were stored in preserving liquids and under refrigeration have been unsatisfactory .

Project Title Device for finding queen bees in managed beehives

RIRDC Project No.: PFP-1A

Start Date: 01/02/01

Finish Date: 01/11/01

Researcher: Mr. Alan Roberts

Organisation: Podargus Farm Products
Boundary Creek Road
BENTLEY NSW 2480

Phone: (02) 6663 5224

Fax: -

Email: alanrbts@fan.net.au

Objectives To design and prove an electronic device capable of pinpointing the position of queen bees in managed beehives.

Current Progress The project has only recently started. We have completed a literature survey of possible electronic techniques and have been assembling and sourcing equipment for the first experiments to determine the attenuation by the beehive material at the electromagnetic frequencies of interest.

Project Title Molecular taxonomy of Nosema apis

RIRDC Project No.: CSE-3H

Researcher: Robert N. Rice

Organisation: CSIRO Division of Entomology
GPO Box 1700
CANBERRA ACT 2601

Phone: (02) 6249 3069

Fax: (02) 6249 5573

Email: r.rice@mailbox.uq.edu.au

Objectives To study genotypic variation in the large subunit (LSU) ribosomal RNA (rRNA) of Nosema apis, and determine if variation detected was suitable as a genetic fingerprint for strain identification. To investigate if thymol is likely to suppress Nosema disease in honeybees.

Background Nosema apis is an economically important pathogen of honeybees and yet little is known about its genetic diversity. Analysis of genetic diversity requires suitable genetic fingerprints that define strains or isolates. This study attempts to identify useful N. apis genetic fingerprints. Some essential oils are known to control some pathogenic bacteria and fungi. Observations by a beekeeper suggest that thymol may suppress nosema disease in honeybees. N. apis cause Nosema disease.

Research A partial gene sequence from the LSU rRNA of nine N. apis isolates was determined and compared for intraspecific variation. Interspecific comparisons were also made to two other species of microsporidia. The efficacy of thymol as a control for nosema disease in honeybees was investigated in a model system. The closely related species Nosema vespula and host caterpillars of the moth Helicoverpa armigera were used. A dietary regime containing thymol was fed to caterpillars orally inoculated with N. vespula spores. Ten days post-infection, the total number of spores per caterpillar was determined and the data statistically analysed.

Outcomes Genetic variation was determined amongst the nine N. apis isolates and between N. apis and two other species. The level intraspecific variation detected was not sufficient to be useful as a genetic fingerprint. The efficacy of thymol as a possible control for Nosema disease was demonstrated.

Implications Further research is required to identify suitable genetic fingerprints for strain identification amongst N. apis. It is likely that thymol will suppress Nosema disease in honeybees when included in artificial diets. The data presented warrants further research.

Publications Rice. R.N., 1999. An Evolutionary Study of Two Microsporidia. A thesis submitted for the degree of Doctor of Philosophy of the Australian National University, Canberra, Australia. Other papers in preparation.
Nosema Disease in Honeybees – Genetic Variation and Control, RIRDC Publication No. 01/046

Project Title Can the technique of 'shaking bees' and antibiotic therapy be used as a means of controlling American Foulbrood?

RIRDC Project No.: DAN-176A

Researcher: Michael Hornitzky

Organisation: NSW Agriculture
Elizabeth Macarthur Agricultural Institute
PMB 8, CAMDEN NSW 2570

Phone: (02) 4640 6311

Fax: (02) 4640 6400

Email: michael.hornitzky@agric.nsw.gov.au

Objectives To determine whether the technique of shaking bees from hives with American foulbrood onto either foundation or irradiated equipment, followed by treatment with oxytetracycline hydrochloride (OTC) is an effective alternative to current control strategies

Background AFB is a major disease of honeybees in Australia and it attracts more funds for its control than any other bee disease. The current control strategies for AFB on mainland Australia are destruction of diseased colonies or the gamma irradiation of infected hive material once the bees have been destroyed. These measures are costly especially when hives are destroyed. The losses incurred from diseased hives not only include the value of the hives but the loss of production, which occurs as a result of the destruction of bees. An alternative strategy, which does not involve destruction of the bees, may decrease the economic impact of this disease on the beekeeper. At State Brood Disease Forums and the National Workshop on Honeybee Brood Diseases held in recent years a consensus was reached that the shaking and OTC treatment technique be tested as an alternative control strategy.

Research One hundred and one hives in five apiaries under field conditions were used to determine whether the technique of shaking bees is an effective alternative AFB control strategy. Five shaking treatments were used.

Outcomes None of the treatments was 100% successful. The most successful treatment appeared to be shaking bees onto foundation with one dose of OTC. Of the 29 hives treated in this way 19 (65.5%) were successfully treated although the adult bees from two hives still carried Paenibacillus larvae (AFB)spores at the end of the trial. Two hives (6.8%) developed AFB and 8 (27.6) died out. The second most successful treatment appeared to be shaking bees onto foundation without OTC treatment where 10 (41.7%) of 24 hives were successfully treated although the adult bees from two of these hives also carried P. larvae spores. However, 2 (8.3%) developed AFB and 12 (50.0%) died out. Of the 32 hives, which had been prepared by shaking bees onto irradiated equipment and one OTC treatment 14 (43.8%), were successfully treated. Three hives (9.4%) developed AFB and 15 (46.9%) died out. AFB developed at varying times in the test apiaries. In one apiary AFB developed in hives after 3 months, 12 months, 14 months and 17 months after the shaking procedure. No further examinations were carried out after the 17-month examination. This indicates that further breakdowns could have occurred after this date.

Implications There is no doubt that the three treatment methods: (a) shaking bees on to foundation (b) shaking bees onto foundation with one OTC treatment and (c) shaking bees onto irradiated material with one OTC treatment can successfully eliminate AFB in some hives. However, it is not certain how many more hives would have broken down with AFB had hive examinations been continued. This uncertainty coupled with the facts that one or more hives developed AFB in each shaking treatment procedure and 39 (41.1%) of the remaining 95 trial hives (6 had been mistakenly killed by a beekeeper) died, suggests that shaking bees is unlikely to be an effective alternative AFB control strategy.

Publications Controlling American Foulbrood – Assessing Effectiveness of Shaking Bees and Antibiotic Therapy Strategies, RIRDC Publication No. 01/048

Project Title Literature review of chalkbrood

RIRDC Project No.: DAN-190A

Researcher: Michael Hornitzky

Organisation: NSW Agriculture
Elizabeth Macarthur Agricultural Institute
PMB 8
CAMDEN 2570

Phone: 0246406311

Fax: 0246406400

Email: michael.hornitzky@agric.nsw.gov.au

Objectives To prepare a literature review of chalkbrood (a fungal disease of honeybees)

Background Chalkbrood is a major disease of honeybees in Australia. A literature review of all aspects of this disease would be useful to facilitate the identification of productive research areas for the control of this disease.

Research Computer data bases, journals and books were used to compile this literature review

Outcomes A broad range of literature dealing with the cause, occurrence, multiplication and spread, diagnosis and control of the disease was examined. This data was compiled into one document which also includes abstracts of the more important papers.

Implications This document is an up to date review of chalkbrood including an outline and assessment of current control strategies.

Publications -

Project Title Sterilisation of beehive components using hot wax dipping and case studies of honey bee disease barrier systems

RIRDC Project No.: DAV-167A

Researcher: Russell Goodman

Organisation: Institute for Horticultural Development
Agriculture Victoria
Department of Natural Resources and Environment

Phone: (03) 9210 9222

Fax: (03) 9800 3521

Email: russell.goodman@nre.vic.gov.au

Objectives To assist the Australian apiary industry control and reduce the spread of the notifiable honey bee brood disease, American Foulbrood, by identifying and documenting 'Best Practice' used in:
· sterilisation of beehive components using hot wax dipping techniques
· beehive barrier management systems.

Background The honey bee brood disease American Foulbrood is caused by the bacterium Paenibacillus larvae and is endemic in all Australian States. This notifiable disease slowly kills honey bee colonies and if not controlled can cause significant economic losses in apiary enterprises.
The frequent interchange of beehive components from hive to hive, and from apiary to apiary can cause significant spread of the disease. Barrier management systems restrict the interchange of hive material to individual hives, small groups of hives or entire yards of hives. The barrier aims to prevent the spread of disease. Barrier management is widely practised in Western Australia and to a large degree in South Australia but rarely in the eastern States.
Hot wax dipping is used to both preserve wooden beehive components and also to sterilise beehive components contaminated with P. larvae spores.
This project was funded to provide the apiary industry with detailed information about best practice in barrier management and hot wax dipping techniques.

Research A literature search was conducted to identify recent developments in barrier management and hot wax dipping techniques.
Selected apiarists using barrier management systems and/or hot wax dipping techniques were visited and interviewed to determine ‘best practice’ methods currently in use. Occupational health and safety issues relating to the use of hot molten wax for dipping were also identified.

Outcomes The information obtained from these cooperating apiarists was complied in two illustrated documents entitled:
· Hot wax dipping of beehive components for preservation and sterilisation
· Case studies of honey bee disease barrier management systems.

Implications The two documents will provide the apiary industry with detailed information about ‘best practice’ in hot wax dipping and barrier management techniques.
The information will enable apiarists to:
· use hot wax dipping safely and effectively to extend the life of wooden hive components and sterilise American Foulbrood contaminated hive components which would otherwise have been destroyed
· incorporate barrier systems into disease management plans to minimise spread of American Foulbrood disease and reduce loss of hives and honey production in apiary enterprises.

Publications Hot wax dipping of beehive components for preservation and sterilisation, RIRDC Publication No. 01/051
Honey bee disease barrier management systems – case studies, RIRDC Publication No. 01/052

Project Title European Foulbrood - investigating control measures

RIRDC Project No.: DAV-157A

Start Date: 07/12/98

Finish Date: 10/07/02

Researcher: Mr Russell Goodman

Organisation: Department of Natural Resources & Environment (Vic)
Institute for Horticultural Development
Private Bag 15
SOUTH EAST MAIL CENTRE VIC 3176

Phone: (03) 9210 9324

Fax: (03) 9800 3521

Email: russell.goodman@nre.vic.gov.au

Objectives a) To protect the apiary industry's continued access to domestic and export honey markets by reducing or eliminating industry's dependence on oxtetracycline hydrochloride (OTC) for the control of the bacterial honeybee brood disease, European Foulbrood (EFB) (Melissococcus pluton).
b) To determine the efficacy of reduced doses of OTC and use of OTC extender patties for the control of EFB and to determine if these measures reduce or eliminate the occurrence of OTC residues in honey.
c) To identify and develop alternative, non-antibiotic measures for control of EFB by investigating, primarily, the effect of enhanced honeybee colony nutrition and changed pH of honeybee larval guts.
d) To obtain a greater understanding of active and latent infections of M. pluton and Paenibacillus alvei (a common secondary invader) in honeybee larvae and to develop new Polymerase Chain Reaction (PCR) methodologies for detection of M. pluton as a necessary prerequisite and support of the preceding aim c).

Current Progress During the first spring of this two-year sub-trial, 120 honey bee colonies with no previous history of oxytetracycline hydrochloride treatment were allotted to 3 groups of 40 hives and treated with the following:
1. pollen supplement protein cakes during late autumn and spring on a 'as needs basis'
2. oxytetracycline hydrochloride (1g active) in early spring
3. controls - no treatment.
Hives were regularly monitored for incidence of European Foulbrood disease from early spring to early summer.
European Foulbrood disease (EFB) symptoms were detected in only four hives. At first, this lack of infection presented a problem and continuation of this work was questioned. However, hemi-nested PCR assay of larvae sampled from colonies belonging to each treatment group confirmed the presence of the causal organism, M. pluton, in all cases. The low number of hives expressing EFB symptoms was attributed to anti-microbial properties (ie high fatty acid composition including linoleic acid) of the pollens foraged by the bees throughout the spring season. The results therefore provide valuable anecdotal evidence of the value of pollens with anti-microbial properties.
At the time of writing, hives were being prepared for the second season of this work. Hives were moved to an apiary site in northern Victoria to access an Ironbark honey flow and to provide field conditions which might encourage development of EFB and expression of symptoms during the 2001 spring. In late autumn, hives belonging to treatment group 1. will be fed pollen supplement protein cakes. In late winter, all hives will be moved to an almond orchard because of the reputed association of EFB outbreaks with this blossom. For the remainder of spring, the hives will be deliberately located where the bees can only access pollen with low anti-microbial properties.

Project Title: Export package bees - evaluation of a lupin flour based feed for increased live bee production

RIRDC Project No.: DAW-75A

Researcher: Rob Manning

Organisation: Agriculture WA

Phone: 08 9 3683567

Fax: 08 9 4741295

Email: rmanning@agric.wa.gov.au

Objectives To evaluate the use of lupin flour as a substitute for torula yeast or soyflour in patties for supplementing the diets of honey bees.

Background To optimise the amount of honey bees shaken from beehives for packages supplementary feeding of beehives has to become part of the hive management process. The primary supplementary foodstuff fed to honey bees is soyflour, an imported feedstuff into Australia, thus not freely available and relatively expensive. Other ingredients used in the mixes (e.g. torula yeast) are also expensive. Lupin flour, as an ingredient, is very cheap when compared to soyflour or torula yeast as Western Australia is the world’s leading exporter of lupins with production exceeding one million tonnes annually. Lupin flour has not been used in a scientific experiment on feeding honey bees and as lupin is the largest crop grown in Western Australia it makes for a potentially cheap and freely available alternative source of protein for honey bee feedstuffs Australia wide.

Research Two separate series of experiments were carried out to evaluate the methodology of feeding feedstuffs to honey bees to gauge their preference and palatability to the different feeds in tests using lupin flour.
The standard test of preference, where five feeds are placed into one hive (Stace and Hayter 1994) was used in this experiment. Initial concerns (Jane Speijers, Agriculture WA - Biometrician) about this method centred around whether the different feeds placed together would affect the honey bees behaviour towards particular feeds and thus affect their palatability. Based on this concern, another experiment on testing the palatability of feedstuffs was carried where each of the five patties were placed into five hives at the same time.
The preference and palatability tests were carried out simultaneously over 137 days, with each trial (of three replicates) taking about 27 days to complete.

Outcomes 1. A mix of lupin and sucrose fed as a patty was most preferred by honey bees over similar mixes containing soyflour or torula yeast. Consumption of the lupin-sucrose mix was ranked third and behind a soyflour and torula yeast-sucrose mix in palatability tests. Torula yeast was the most palatable feed.
2. Lupin based feeds still need further development with different ingredients to achieve higher consumption rates and increased brood rearing.
3. Supplementary feeding of beehives suppressed pollen intake when pollen traps are used.
4. The rate of consumption of patties by honey bees declined, the longer they are left in the hive for testing.
5. Statistically, measurements of the brood frames and pollen intake showed that the brood frame development and the amount of pollen trapped had no impact on the rate of consumption of the feedstuffs used in this experiment.
6. Experiments on supplementary feedstuffs can be done throughout the year if pollen traps are used to restrict the amount of pollen entering hives.
The experiment was terminated when it was well into its second round of testing when chalkbrood was discovered by the researcher (1^st time in WA) and as a consequence State quarantine orders were initiated and a [premature] decision was made to destroy the apiary, which was subsequently burnt on the 29^th April 1998. With the prolonged quarantine measures, and other projects undertaken, a decision was made to abandon the project to some future date.

Implications 1. Preference testing should be replaced by palatability tests as more qualitative information about each of the feeds can be gathered. The advantage is that the actual effect of the feedstuffs can be seen from hive measurements of burr-comb and live-bee production.
2. Fixed feeding times of short duration (about 5-7 days) should be part of the methodology in any experiment testing supplementary feeds.

Publications Nil

Project Title Pollen analysis of Eucalyptus patens (Blackbutt), E. accedens (Powderbark) and E. wandoo (Winter Wandoo variety) in Western Australia

RIRDC Project No.: DAW-91A

Researcher: Mr Rob Manning

Organisation: Agriculture WA
Locked Bag No 4
BENTLEY DELIVERY CENTRE WA 6983

Phone: (08) 9368 3567

Fax: (08) 9474 1295

Email: rmanning@agric.wa.gov.au

Objectives Provide an analysis of six commercially important species of eucalypt pollens, four of which will be new to science and provide a fatty acid profile of the lipid fraction for six species of eucalypt for the first time in Australia.

Background Western Australia has the largest number of endemic eucalyptus species identified in Australia (Chippendale and Wolf, 1981). Nectar (as honey) and pollen from these eucalypts play an important part in the health and sustainability of the beekeeping industry and both products are extensively used by the community and contribute to Western Australia’s export market.
This project examined six commercially important beekeeper-targeted species: E. accedens (powderbark), E. diversicolor (karri), E. marginata (Jarrah), E. patens (forest blackbutt) and Eucalyptus wandoo (wandoo) for their protein composition, fatty acid and mineral profile and vitamin content. Redgum pollen (Corymbia calophylla) was used as a base to measure differences between the other species because it is considered by beekeepers to be excellent pollen for honey bee nutrition.

Research Pollen from six endemic Western Australian eucalypt species were examined for amino acid, fatty acid, mineral and vitamin composition (though incomplete).

Outcomes 1. The lipid fraction of eucalypt pollen is high in C-18 fatty acids, most notably linoleic acid.
2. Amino acid deficiencies were found in powderbark (iso-leucine); jarrah (iso-leucine, histidine & possibly threonine) which also had total protein below the minimum of 20%; winter, spring and summer wandoo (iso-leucine).
3. Nutrition balance needs to be adjusted for the above species by supplementary feeding of soyflour or yeast (as examples).

Implications 1. The honey flows from jarrah, powderbark and wandoo will reduce the lifespans of honey bees. If there are no pollens from other species being collected by honey bees then the effect on the honey production (for example) should be significant.
2. The high levels of C-18 fatty acids, particularly linoleic and linolenic in pollens assist in inhibiting of bacteria and yeasts within the hive. Of particular importance for beekeepers is the control of the bacteria that causes American foulbrood, European foulbrood and botulism.

Publications Pollen Analysis of Eucalypts from Western Australia, RIRDC Publication No. 01/053

Project Title	Improving queen bee production
RIRDC Project No.:	CSE-85A
Start Date:	01/07/99
Finish Date:	31/07/02
Researcher:	Dr. Denis Anderson
Organisation:	CSIRO Entomology GPO Box 1700 CANBERRA ACT 2601
Phone:	(02) 6246 4148
Fax:	(02) 6246 4000
Email:	Denis.Anderson@ento.csiro.au
Objectives	To improve the profitability of commercial queen producers by improving queen performance. This will be achieved by: 1. Determining whether nutritional supplements significantly improve queen and drone quality; 2. Determining the factors which are responsible for causing disappearing disorder.
Current Progress	Effects of nutrient supplements on queen cell-building colonies This sub-project aims to determine whether the feeding of nutrient supplements during queen production improves the quality of adult queen bees. During the 1999/00 season, the following supplements were each fed to 5 queen cell-building colonies of equal strength: (a) no supplement (control), (b) irradiated pollen, (c) irradiated pollen with added vitamins, (d) soya-flour based supplement, (e) soya-flour based supplement with added vitamins and (f) vitamins. These test-supplements were those recommended by queen producers and participants at the ‘Honey Bee Nutrition Workshop’ held in Sydney in May 1998. Sister-queens were subsequently raised in the cell-builders, open mated, and tested for: (1) weight, (2) number of ovarioles, (3) number of spermatozoa in spermathecae, (4) nosema levels and (5) body protein. No significant differences were found in the quality of adult queen bees raised from cell-builders fed the different diets. It was concluded that this unexpected result was due to the excellent forage conditions that prevailed in the local environment where the trials were conducted. It is planned to repeat these trials in the future when local forage conditions are poor. At the present time (during the latter part of the 2000/01 season), the effects of feeding nutrient supplements to queen banks is being assessed. Disappearing Disorder This sub-project aims to determine the cause of disappearing disorder which affects bee colonies at certain times of the year in North-East NSW and South-East Queensland. During the 1999/2000 season, the disorder was mild and tests conducted on the few samples that were collected showed no evidence for the presence of pathogenic micro-organisms or excessive levels of toxic elements in the nectar and pollen being consumed by the affected larvae. During the 2000/01 season, the disorder was moderately severe at Cunningham’s gap in SE Queensland, but almost absent in the Gympie area to the north-east. Large numbers of samples of affected larvae were collected from an apiary at Cunningham’s Gap. Tests on these larvae again showed no evidence that the disorder resulted from the presence of pathogenic micro-organisms or toxic elements. Further tests are currently being conducted on the samples.

Project Title	Introduction and early performance of queen bees – some factors affecting success
RIRDC Project No.:	DAN-182A
Start Date:	01/09/99
Finish Date:	31/07/01
Researcher:	Mr. John Rhodes
Organisation:	NSW Department of Agriculture RMB 944 Calala Lane TAMWORTH NSW 2340
Phone:	(02) 6763 1206
Fax:	(02) 6763 1222
Email:	Rhodesj@agric.nsw.gov.au
Objectives	· To determine whether the age of the queen bee at introduction influences introduction success. · To identify factors affecting sperm counts of sister queen bees mated under the same mating conditions and to investigate relationships between low sperm counts and queen failure following introduction. · To examine the effects of temperature and humidity during transport of queen bees on their introduction success. · A comparison of sperm counts between sister bees mated in a commercial queen bee breeder`s apiary and between those mated in a commercial honey producer`s queen bee breeding apiary. · Development of a method for preserving adult drones which would allow accurate sperm counts to be carried out on those drones.
Current Progress	Field and laboratory work for objectives 1 to 4 have been completed and relevant data are being analysed. Compared to the 1999-2000 season, seasonal conditions were harder on bees than the 2000-2001 season however higher numbers of queen bees survived for each age group for both the 14 day and 15 week survival counts. The trend of increased numbers of queens surviving corresponding to increased age queens were caught from their mating nucleus continued for both seasons. Work on objective 5 is continuing, small trials in which whole drones were stored in preserving liquids and under refrigeration have been unsatisfactory .

Project Title	Device for finding queen bees in managed beehives
RIRDC Project No.:	PFP-1A
Start Date:	01/02/01
Finish Date:	01/11/01
Researcher:	Mr. Alan Roberts
Organisation:	Podargus Farm Products Boundary Creek Road BENTLEY NSW 2480
Phone:	(02) 6663 5224
Fax:	-
Email:	alanrbts@fan.net.au
Objectives	To design and prove an electronic device capable of pinpointing the position of queen bees in managed beehives.
Current Progress	The project has only recently started. We have completed a literature survey of possible electronic techniques and have been assembling and sourcing equipment for the first experiments to determine the attenuation by the beehive material at the electromagnetic frequencies of interest.

Project Title	Molecular taxonomy of Nosema apis
RIRDC Project No.:	CSE-3H
Researcher:	Robert N. Rice
Organisation:	CSIRO Division of Entomology GPO Box 1700 CANBERRA ACT 2601
Phone:	(02) 6249 3069
Fax:	(02) 6249 5573
Email:	r.rice@mailbox.uq.edu.au
Objectives	To study genotypic variation in the large subunit (LSU) ribosomal RNA (rRNA) of Nosema apis, and determine if variation detected was suitable as a genetic fingerprint for strain identification. To investigate if thymol is likely to suppress Nosema disease in honeybees.
Background	Nosema apis is an economically important pathogen of honeybees and yet little is known about its genetic diversity. Analysis of genetic diversity requires suitable genetic fingerprints that define strains or isolates. This study attempts to identify useful N. apis genetic fingerprints. Some essential oils are known to control some pathogenic bacteria and fungi. Observations by a beekeeper suggest that thymol may suppress nosema disease in honeybees. N. apis cause Nosema disease.
Research	A partial gene sequence from the LSU rRNA of nine N. apis isolates was determined and compared for intraspecific variation. Interspecific comparisons were also made to two other species of microsporidia. The efficacy of thymol as a control for nosema disease in honeybees was investigated in a model system. The closely related species Nosema vespula and host caterpillars of the moth Helicoverpa armigera were used. A dietary regime containing thymol was fed to caterpillars orally inoculated with N. vespula spores. Ten days post-infection, the total number of spores per caterpillar was determined and the data statistically analysed.
Outcomes	Genetic variation was determined amongst the nine N. apis isolates and between N. apis and two other species. The level intraspecific variation detected was not sufficient to be useful as a genetic fingerprint. The efficacy of thymol as a possible control for Nosema disease was demonstrated.
Implications	Further research is required to identify suitable genetic fingerprints for strain identification amongst N. apis. It is likely that thymol will suppress Nosema disease in honeybees when included in artificial diets. The data presented warrants further research.
Publications	Rice. R.N., 1999. An Evolutionary Study of Two Microsporidia. A thesis submitted for the degree of Doctor of Philosophy of the Australian National University, Canberra, Australia. Other papers in preparation. Nosema Disease in Honeybees – Genetic Variation and Control, RIRDC Publication No. 01/046

Project Title	Can the technique of 'shaking bees' and antibiotic therapy be used as a means of controlling American Foulbrood?
RIRDC Project No.:	DAN-176A
Researcher:	Michael Hornitzky
Organisation:	NSW Agriculture Elizabeth Macarthur Agricultural Institute PMB 8, CAMDEN NSW 2570
Phone:	(02) 4640 6311
Fax:	(02) 4640 6400
Email:	michael.hornitzky@agric.nsw.gov.au
Objectives	To determine whether the technique of shaking bees from hives with American foulbrood onto either foundation or irradiated equipment, followed by treatment with oxytetracycline hydrochloride (OTC) is an effective alternative to current control strategies
Background	AFB is a major disease of honeybees in Australia and it attracts more funds for its control than any other bee disease. The current control strategies for AFB on mainland Australia are destruction of diseased colonies or the gamma irradiation of infected hive material once the bees have been destroyed. These measures are costly especially when hives are destroyed. The losses incurred from diseased hives not only include the value of the hives but the loss of production, which occurs as a result of the destruction of bees. An alternative strategy, which does not involve destruction of the bees, may decrease the economic impact of this disease on the beekeeper. At State Brood Disease Forums and the National Workshop on Honeybee Brood Diseases held in recent years a consensus was reached that the shaking and OTC treatment technique be tested as an alternative control strategy.
Research	One hundred and one hives in five apiaries under field conditions were used to determine whether the technique of shaking bees is an effective alternative AFB control strategy. Five shaking treatments were used.
Outcomes	None of the treatments was 100% successful. The most successful treatment appeared to be shaking bees onto foundation with one dose of OTC. Of the 29 hives treated in this way 19 (65.5%) were successfully treated although the adult bees from two hives still carried Paenibacillus larvae (AFB)spores at the end of the trial. Two hives (6.8%) developed AFB and 8 (27.6) died out. The second most successful treatment appeared to be shaking bees onto foundation without OTC treatment where 10 (41.7%) of 24 hives were successfully treated although the adult bees from two of these hives also carried P. larvae spores. However, 2 (8.3%) developed AFB and 12 (50.0%) died out. Of the 32 hives, which had been prepared by shaking bees onto irradiated equipment and one OTC treatment 14 (43.8%), were successfully treated. Three hives (9.4%) developed AFB and 15 (46.9%) died out. AFB developed at varying times in the test apiaries. In one apiary AFB developed in hives after 3 months, 12 months, 14 months and 17 months after the shaking procedure. No further examinations were carried out after the 17-month examination. This indicates that further breakdowns could have occurred after this date.
Implications	There is no doubt that the three treatment methods: (a) shaking bees on to foundation (b) shaking bees onto foundation with one OTC treatment and (c) shaking bees onto irradiated material with one OTC treatment can successfully eliminate AFB in some hives. However, it is not certain how many more hives would have broken down with AFB had hive examinations been continued. This uncertainty coupled with the facts that one or more hives developed AFB in each shaking treatment procedure and 39 (41.1%) of the remaining 95 trial hives (6 had been mistakenly killed by a beekeeper) died, suggests that shaking bees is unlikely to be an effective alternative AFB control strategy.
Publications	Controlling American Foulbrood – Assessing Effectiveness of Shaking Bees and Antibiotic Therapy Strategies, RIRDC Publication No. 01/048

Project Title	Literature review of chalkbrood
RIRDC Project No.:	DAN-190A
Researcher:	Michael Hornitzky
Organisation:	NSW Agriculture Elizabeth Macarthur Agricultural Institute PMB 8 CAMDEN 2570
Phone:	0246406311
Fax:	0246406400
Email:	michael.hornitzky@agric.nsw.gov.au
Objectives	To prepare a literature review of chalkbrood (a fungal disease of honeybees)
Background	Chalkbrood is a major disease of honeybees in Australia. A literature review of all aspects of this disease would be useful to facilitate the identification of productive research areas for the control of this disease.
Research	Computer data bases, journals and books were used to compile this literature review
Outcomes	A broad range of literature dealing with the cause, occurrence, multiplication and spread, diagnosis and control of the disease was examined. This data was compiled into one document which also includes abstracts of the more important papers.
Implications	This document is an up to date review of chalkbrood including an outline and assessment of current control strategies.
Publications	-

Project Title	Sterilisation of beehive components using hot wax dipping and case studies of honey bee disease barrier systems
RIRDC Project No.:	DAV-167A
Researcher:	Russell Goodman
Organisation:	Institute for Horticultural Development Agriculture Victoria Department of Natural Resources and Environment
Phone:	(03) 9210 9222
Fax:	(03) 9800 3521
Email:	russell.goodman@nre.vic.gov.au
Objectives	To assist the Australian apiary industry control and reduce the spread of the notifiable honey bee brood disease, American Foulbrood, by identifying and documenting 'Best Practice' used in: · sterilisation of beehive components using hot wax dipping techniques · beehive barrier management systems.
Background	The honey bee brood disease American Foulbrood is caused by the bacterium Paenibacillus larvae and is endemic in all Australian States. This notifiable disease slowly kills honey bee colonies and if not controlled can cause significant economic losses in apiary enterprises. The frequent interchange of beehive components from hive to hive, and from apiary to apiary can cause significant spread of the disease. Barrier management systems restrict the interchange of hive material to individual hives, small groups of hives or entire yards of hives. The barrier aims to prevent the spread of disease. Barrier management is widely practised in Western Australia and to a large degree in South Australia but rarely in the eastern States. Hot wax dipping is used to both preserve wooden beehive components and also to sterilise beehive components contaminated with P. larvae spores. This project was funded to provide the apiary industry with detailed information about best practice in barrier management and hot wax dipping techniques.
Research	A literature search was conducted to identify recent developments in barrier management and hot wax dipping techniques. Selected apiarists using barrier management systems and/or hot wax dipping techniques were visited and interviewed to determine ‘best practice’ methods currently in use. Occupational health and safety issues relating to the use of hot molten wax for dipping were also identified.
Outcomes	The information obtained from these cooperating apiarists was complied in two illustrated documents entitled: · Hot wax dipping of beehive components for preservation and sterilisation · Case studies of honey bee disease barrier management systems.
Implications	The two documents will provide the apiary industry with detailed information about ‘best practice’ in hot wax dipping and barrier management techniques. The information will enable apiarists to: · use hot wax dipping safely and effectively to extend the life of wooden hive components and sterilise American Foulbrood contaminated hive components which would otherwise have been destroyed · incorporate barrier systems into disease management plans to minimise spread of American Foulbrood disease and reduce loss of hives and honey production in apiary enterprises.
Publications	Hot wax dipping of beehive components for preservation and sterilisation, RIRDC Publication No. 01/051 Honey bee disease barrier management systems – case studies, RIRDC Publication No. 01/052

Project Title	European Foulbrood - investigating control measures
RIRDC Project No.:	DAV-157A
Start Date:	07/12/98
Finish Date:	10/07/02
Researcher:	Mr Russell Goodman
Organisation:	Department of Natural Resources & Environment (Vic) Institute for Horticultural Development Private Bag 15 SOUTH EAST MAIL CENTRE VIC 3176
Phone:	(03) 9210 9324
Fax:	(03) 9800 3521
Email:	russell.goodman@nre.vic.gov.au
Objectives	a) To protect the apiary industry's continued access to domestic and export honey markets by reducing or eliminating industry's dependence on oxtetracycline hydrochloride (OTC) for the control of the bacterial honeybee brood disease, European Foulbrood (EFB) (Melissococcus pluton). b) To determine the efficacy of reduced doses of OTC and use of OTC extender patties for the control of EFB and to determine if these measures reduce or eliminate the occurrence of OTC residues in honey. c) To identify and develop alternative, non-antibiotic measures for control of EFB by investigating, primarily, the effect of enhanced honeybee colony nutrition and changed pH of honeybee larval guts. d) To obtain a greater understanding of active and latent infections of M. pluton and Paenibacillus alvei (a common secondary invader) in honeybee larvae and to develop new Polymerase Chain Reaction (PCR) methodologies for detection of M. pluton as a necessary prerequisite and support of the preceding aim c).
Current Progress	During the first spring of this two-year sub-trial, 120 honey bee colonies with no previous history of oxytetracycline hydrochloride treatment were allotted to 3 groups of 40 hives and treated with the following: 1. pollen supplement protein cakes during late autumn and spring on a 'as needs basis' 2. oxytetracycline hydrochloride (1g active) in early spring 3. controls - no treatment. Hives were regularly monitored for incidence of European Foulbrood disease from early spring to early summer. European Foulbrood disease (EFB) symptoms were detected in only four hives. At first, this lack of infection presented a problem and continuation of this work was questioned. However, hemi-nested PCR assay of larvae sampled from colonies belonging to each treatment group confirmed the presence of the causal organism, M. pluton, in all cases. The low number of hives expressing EFB symptoms was attributed to anti-microbial properties (ie high fatty acid composition including linoleic acid) of the pollens foraged by the bees throughout the spring season. The results therefore provide valuable anecdotal evidence of the value of pollens with anti-microbial properties. At the time of writing, hives were being prepared for the second season of this work. Hives were moved to an apiary site in northern Victoria to access an Ironbark honey flow and to provide field conditions which might encourage development of EFB and expression of symptoms during the 2001 spring. In late autumn, hives belonging to treatment group 1. will be fed pollen supplement protein cakes. In late winter, all hives will be moved to an almond orchard because of the reputed association of EFB outbreaks with this blossom. For the remainder of spring, the hives will be deliberately located where the bees can only access pollen with low anti-microbial properties.

Project Title:	Export package bees - evaluation of a lupin flour based feed for increased live bee production
RIRDC Project No.:	DAW-75A
Researcher:	Rob Manning
Organisation:	Agriculture WA
Phone:	08 9 3683567
Fax:	08 9 4741295
Email:	rmanning@agric.wa.gov.au
Objectives	To evaluate the use of lupin flour as a substitute for torula yeast or soyflour in patties for supplementing the diets of honey bees.
Background	To optimise the amount of honey bees shaken from beehives for packages supplementary feeding of beehives has to become part of the hive management process. The primary supplementary foodstuff fed to honey bees is soyflour, an imported feedstuff into Australia, thus not freely available and relatively expensive. Other ingredients used in the mixes (e.g. torula yeast) are also expensive. Lupin flour, as an ingredient, is very cheap when compared to soyflour or torula yeast as Western Australia is the world’s leading exporter of lupins with production exceeding one million tonnes annually. Lupin flour has not been used in a scientific experiment on feeding honey bees and as lupin is the largest crop grown in Western Australia it makes for a potentially cheap and freely available alternative source of protein for honey bee feedstuffs Australia wide.
Research	Two separate series of experiments were carried out to evaluate the methodology of feeding feedstuffs to honey bees to gauge their preference and palatability to the different feeds in tests using lupin flour. The standard test of preference, where five feeds are placed into one hive (Stace and Hayter 1994) was used in this experiment. Initial concerns (Jane Speijers, Agriculture WA - Biometrician) about this method centred around whether the different feeds placed together would affect the honey bees behaviour towards particular feeds and thus affect their palatability. Based on this concern, another experiment on testing the palatability of feedstuffs was carried where each of the five patties were placed into five hives at the same time. The preference and palatability tests were carried out simultaneously over 137 days, with each trial (of three replicates) taking about 27 days to complete.
Outcomes	1. A mix of lupin and sucrose fed as a patty was most preferred by honey bees over similar mixes containing soyflour or torula yeast. Consumption of the lupin-sucrose mix was ranked third and behind a soyflour and torula yeast-sucrose mix in palatability tests. Torula yeast was the most palatable feed. 2. Lupin based feeds still need further development with different ingredients to achieve higher consumption rates and increased brood rearing. 3. Supplementary feeding of beehives suppressed pollen intake when pollen traps are used. 4. The rate of consumption of patties by honey bees declined, the longer they are left in the hive for testing. 5. Statistically, measurements of the brood frames and pollen intake showed that the brood frame development and the amount of pollen trapped had no impact on the rate of consumption of the feedstuffs used in this experiment. 6. Experiments on supplementary feedstuffs can be done throughout the year if pollen traps are used to restrict the amount of pollen entering hives. The experiment was terminated when it was well into its second round of testing when chalkbrood was discovered by the researcher (1^st time in WA) and as a consequence State quarantine orders were initiated and a [premature] decision was made to destroy the apiary, which was subsequently burnt on the 29^th April 1998. With the prolonged quarantine measures, and other projects undertaken, a decision was made to abandon the project to some future date.
Implications	1. Preference testing should be replaced by palatability tests as more qualitative information about each of the feeds can be gathered. The advantage is that the actual effect of the feedstuffs can be seen from hive measurements of burr-comb and live-bee production. 2. Fixed feeding times of short duration (about 5-7 days) should be part of the methodology in any experiment testing supplementary feeds.
Publications	Nil

Project Title	*Pollen analysis of Eucalyptus patens* (Blackbutt), E. accedens (Powderbark) and E. wandoo (Winter Wandoo variety) in Western Australia**
RIRDC Project No.:	DAW-91A
Researcher:	Mr Rob Manning
Organisation:	Agriculture WA Locked Bag No 4 BENTLEY DELIVERY CENTRE WA 6983
Phone:	(08) 9368 3567
Fax:	(08) 9474 1295
Email:	rmanning@agric.wa.gov.au
Objectives	Provide an analysis of six commercially important species of eucalypt pollens, four of which will be new to science and provide a fatty acid profile of the lipid fraction for six species of eucalypt for the first time in Australia.
Background	Western Australia has the largest number of endemic eucalyptus species identified in Australia (Chippendale and Wolf, 1981). Nectar (as honey) and pollen from these eucalypts play an important part in the health and sustainability of the beekeeping industry and both products are extensively used by the community and contribute to Western Australia’s export market. This project examined six commercially important beekeeper-targeted species: E. accedens (powderbark), E. diversicolor (karri), E. marginata (Jarrah), E. patens (forest blackbutt) and Eucalyptus wandoo (wandoo) for their protein composition, fatty acid and mineral profile and vitamin content. Redgum pollen (Corymbia calophylla) was used as a base to measure differences between the other species because it is considered by beekeepers to be excellent pollen for honey bee nutrition.
Research	Pollen from six endemic Western Australian eucalypt species were examined for amino acid, fatty acid, mineral and vitamin composition (though incomplete).
Outcomes	1. The lipid fraction of eucalypt pollen is high in C-18 fatty acids, most notably linoleic acid. 2. Amino acid deficiencies were found in powderbark (iso-leucine); jarrah (iso-leucine, histidine & possibly threonine) which also had total protein below the minimum of 20%; winter, spring and summer wandoo (iso-leucine). 3. Nutrition balance needs to be adjusted for the above species by supplementary feeding of soyflour or yeast (as examples).
Implications	1. The honey flows from jarrah, powderbark and wandoo will reduce the lifespans of honey bees. If there are no pollens from other species being collected by honey bees then the effect on the honey production (for example) should be significant. 2. The high levels of C-18 fatty acids, particularly linoleic and linolenic in pollens assist in inhibiting of bacteria and yeasts within the hive. Of particular importance for beekeepers is the control of the bacteria that causes American foulbrood, European foulbrood and botulism.
Publications	Pollen Analysis of Eucalypts from Western Australia, RIRDC Publication No. 01/053

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Rural Industries
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