Governance approval for multisite, non-interventional research: what can Harmonisation of Multi-Centre Ethical Review learn from the New South Wales experience?

BACKGROUND: In 2007, New South Wales Health mandated the separation of ethical and scientific review from research governance at all New South Wales public health sites based on their distinction in the National Health and Medical Research Council National Statement. This separation allowed for single-site ethical review of multicentre studies. AIMS: To investigate the time taken for governance approval of multicentre studies through the site-specific approval (SSA) process. METHODS: A retrospective audit of the SSA process for five non-interventional studies proposed by a university cancer research unit. RESULTS: The median total governance approval time for all submissions (n= 28) was 12 weeks (range 2.5-64); median time from starting the SSA to submission was 8 weeks (range 1-48) and median time for governance approval was 5 weeks (range 0.3-40). Approval times were shorter for public compared to private institutions. Reasons for delays in finalising submissions for approval were the absence of institutional governance officers, lack of clarity regarding signatories, the need to identify a principal investigator employed by the institution, and lack of recognition of ethical approval by private institutions. The need to develop legal agreements between the university and hospital was the main reason for lengthy delays in obtaining approval. CONCLUSIONS: The advantages of a harmonised single ethical review process were undermined by the coexistence of a fragmented, complex and lengthy governance approval process. This experience has implications for the success of the national Harmonisation of Multi-Centre Ethical Review (HoMER) model. A harmonised and fully supported national approach to research governance should be developed contemporaneously with HoMER.

Predacious activity of Duddingtonia flagrans within the cattle faecal pat.

Two studies were conducted to investigate the growth and activity of the fungus, Duddingtonia flagrans, within cattle faecal pats. Artificial faecal pats were constructed with the centre separated from the outer layer by a nylon mesh. Eight treatments were tested, by varying the presence/absence of Cooperia oncophora eggs and fungal spores within each layer. With parasite eggs in the centre layer, a statistically lower recovery of larvae was observed compared to both pats with parasite eggs in the periphery and pats with parasite eggs throughout both layers. Regardless of location within the pat, if co-located with the parasite egg, D. flagrans was found to be effective in trapping developing larvae. The reduction in recovery of larvae from pats with parasite eggs and fungal spores in the centre was found to be significantly higher than when parasite eggs were in the centre and fungal spores in the periphery. In the second study, pats were made up in two treatments: pats containing fungal spores and C. oncophora eggs (fungus) and pats containing C. oncophora eggs (control). The pats were incubated at low or high humidity. Ten pats were used in a cross over where five pats incubated at low humidity for 7 weeks were removed, water added and then incubated at a high humidity for 1 week. Another five pats were incubated at a high humidity for 7 weeks, aerated and incubated at a low humidity for 1 week. There was no apparent growth of fungus in faecal pats incubated at a high humidity and less than 20% of larvae were recovered. The growth of D. flagrans was observed in faecal pats incubated at a low humidity, but a corresponding reduction in the percentage recovery of larvae did not occur, except in week 4. No statistical difference between fungal and control pats was seen in the change over pats. Nematophagous activity was assessed throughout the study and observed in the first 4 weeks within the pats containing fungus.

Possible application of a nematophagous fungus as a biological control agent of parasitic nematodes on commercial sheep farms in South Africa.

Biological control of parasitic nematodes of livestock is currently under development and represents another tool that may be integrated into helminth parasite control strategies. This paper presents a brief introduction to commercial sheep farming in South Africa and currently available nematode parasite control methods. These include the FAMACHA clinical assay, strategies of pasture management, dilution of resistant worm species by introduction of susceptible worms, breed resistant sheep and nutritional supplementation. The purpose of this paper is to outline the principles of biological control using nematophagous fungi and how it may be applied on sheep farms in South Africa.

The potential of nematophagous fungi to control the free-living stages of nematode parasites of sheep: towards the development of a fungal controlled release device.

Studies showed that chlamydospores of the nematophagous fungus, Duddingtonia flagrans, are capable of surviving pressures of several tonnes when incorporated into matrices and pressed into tablets for the manufacture of prototype intraruminal controlled release devices (CRDs). They remain viable in this tabletted form for at least 9 months when stored at 4 degrees C. In vitro studies demonstrated that there was no effect on spore viability of prolonged exposure to either room or elevated temperature (40 degrees C) in air, or under an atmosphere of either of the major ruminal gases, carbon dioxide and methane. In vivo, studies showed that viable chlamydospores could be detected at the erosion surface of prototype CRDs recovered from the rumen and also in faeces of fistulated sheep, for up to 3 weeks after administration. Further studies have shown that chlamydospores released from such devices can substantially reduce the number of infective larvae that develop in cultures of faeces collected from sheep infected with the nematode parasite, Haemonchus contortus. This work demonstrates, in principle, that the deployment of chlamydospores of D. flagrans in intraruminal CRDs, is another possibility in the development of a range of methods for the biological control of parasites in livestock.

The potential of nematophagous fungi to control the free-living stages of nematode parasites of sheep: feeding and block studies with Duddingtonia flagrans.

A series of feeding trials was conducted with penned sheep harboring Trichostrongylus colubriformis infections. They were offered barley grains supporting the growth of the nematophagous fungus Duddingtonia flagrans. It was shown that as little as 5g of grain/sheep per day was sufficient to virtually eliminate larval numbers from faecal culture. This effect persisted for the time that the fungal grains were fed, and for up to 2 days following cessation of feeding this material. Macerated fungal grains were also incorporated into a range of feed block formulations. In all these, D. flagrans was found to survive the manufacturing process and resulted in significant reductions in larval numbers in faecal cultures set up during the feeding period to sheep. This was observed even for sheep that showed only modest and irregular block consumption. These studies demonstrate that supplementary feeding or block administration offer potential deployment options for D. flagrans as a means of biological control of nematode parasites of livestock.

Biological control of field infections of nematode parasites of young sheep with Duddingtonia flagrans and effects of spore intake on efficacy.

A field study was undertaken to determine the effects of feeding Duddingtonia flagrans to young Merino sheep on pasture. A total of 60 mixed sex lambs 4-5 months old were divided into six even groups on the basis of liveweight. On Monday to Friday, each week for 6 months, three groups were offered barley grains on which D. flagrans had been cultured while the other three groups remained untreated. Every 4 weeks liveweights were recorded and faecal samples collected for nematode egg count estimation. Feeding D. flagrans reduced faecal egg counts and tended to improve liveweight gains, but considerable differences were observed between groups within treatment. These differences are thought to result from variations between the groups in consumption of the treated barley with the "best" consumers showing the greater effects of treatment.

Effect of different times of administration of the nematophagous fungus Duddingtonia flagrans on the transmission of ovine parasitic nematodes on pasture--a plot study.

Investigations were made into the timing of administration of Duddingtonia flagrans as a biological control agent against ovine parasitic nematodes including stongylid and Nematodirus spp. Faeces from 3-4 months old male lambs were deposited onto pasture plots that had never been grazed by sheep. The trial was conducted over two consecutive years (1998 and 1999). For both years, the following three plot types were involved: Sim plots had faeces containing nematode eggs and Duddingtonia flagrans spores deposited simultaneously; Post plots had faeces containing nematode eggs followed 2 weeks later by faeces containing D. flagrans spores alone; Control plots had faeces containing only nematode eggs; Prior plots (included in 1999) had faeces containing D. flagrans spores alone followed 2 weeks later by faeces containing nematode eggs. In each year, two deposition periods were involved: July and August in 1998 and June and July in 1999. During the first year pasture samples were collected at 2, 4, 6, 8 and 12 weeks after initial deposition. In 1999, additional samples were collected at 10, 16 and 20 weeks. Larvae were extracted from the pasture samples and counts performed to estimate the number and species of infective third-stage (L(3), larvae) present. The number of third-stage strongylid larvae on pasture was significantly lower on Sim plots compared to the remaining plot types for both years at all deposition times (P<0.001). This was also the case for the number of Nematodirus infective larvae in August deposition plots in 1998 (P<0. 02). There was no significant difference between treatments in both deposition times in 1999 and July deposition plots in 1998 for the Nematodirus data. These results suggest that D. flagrans, if deposited at the same time as parasite eggs prevents transmission of third-stage larvae from the faecal deposit onto pasture, including occasionally Nematodirus species, but does not have an effect on third-stage parasitic nematode larvae in the surrounding soil.

Biological control of nematode parasites of livestock in Fiji: screening of fresh dung of small ruminants for the presence of nematophagous fungi.

Approximately 2500 faecal samples were collected per rectum from sheep and goats from 26 farms located on four of the Fijian islands where most of the small ruminants in this country are raised. The purpose was to screen these samples to isolate nematode-trapping fungi that had been acquired by these animals during the course of their feeding and which had remained viable following passage through their gastrointestinal tract. From these samples, 23 examples of nematophagous fungi were noted in the initial appraisal, from which 12 pure isolates (all of the genus Arthrobotrys) were made. A number factors emerged from this work which may have restricted the opportunities in which nematophagous fungi were detected, or isolated.

The potential of nematophagous fungi to control the free-living stages of nematode parasites of sheep: studies with Duddingtonia flagrans.

The nematophagous fungus, Duddingtonia flagrans, isolated from a fresh sheep faecal sample obtained from a farm in northern New South Wales, Australia, was subjected to a number of in vivo investigations in both surgically modified and normal sheep to determine its capacity to survive passage through the gastrointestinal tract. Single and sustained dosing experiments established that between 5 x 10(5) and 10(6) chlamydospores/day resulted in a substantial (> 80%) reduction in the number of infective larvae derived from nematode eggs in faeces. This effect can be maintained if dosing continues. These results demonstrate for the first time the potential of nematophagous fungi to be deployed by means of sustained release technology in the biological control of nematode parasites of livestock.

The potential of nematophagous fungi to control the free-living stages of nematode parasites of sheep: pasture plot study with Duddingtonia flagrans.

The potential of nematophagous fungi to control the free-living stages of nematode parasites of sheep: Pasture plot study with the fungus Duddingtonia flagrans. Vet. Parasitol. The experiment was designed to test the ability of D. flagrans to reduce infective larval populations on pasture after passage through the gastrointestinal tract of sheep. Merino sheep were given chlamydospores by intra-ruminal infusion at a rate of 5 million chlamydospores/sheep/day and faeces collected from these sheep was deposited on pasture plots. Numbers of larvae recovered from faeces and pasture were both lower on plots from sheep dosed with fungus (P < 0.001 and P < 0.01, respectively) than on plots from control sheep.

The potential of nematophagous fungi to control the free-living stages of nematode parasites of sheep: comparison between Australian isolates of Arthrobotrys spp. and Duddingtonia flagrans.

Nine isolates of Duddingtonia flagrans and eight isolates of Arthrobotrys spp. which originated from a field survey for the presence of nematophagous fungi in fresh dung of livestock in Australia were used in this study. Comparisons were made between the ability of the different isolates to survive gut passage and subsequently reduce infective larval numbers in sheep faeces. Fungal spores (conidia and/or chlamydospores) were administered orally to sheep in doses ranging from 1 X 10(5) to 4.5 X 10(6) spores. There was no apparent consistent survival of Arthrobotrys spp., whereas D. flagrans showed excellent survival capacity which resulted in profound reductions in Trichostrongylus colubriformis larval numbers in culture. This provides clear evidence that D. flagrans is an ideal candidate as a potential biological control agent for nematode parasites of sheep.

The prospects for biological control of the free-living stages of nematode parasites of livestock.

Control of nematode parasites of livestock is focused almost entirely on the parasitic stages within the host. Current methods rely on anthelmintic drugs, but these are under increasing threat with the development of resistance covering the whole spectrum of anthelmintics amongst the important nematode species of a range of livestock. However, invariably the greatest proportion of the parasite biomass resides not within the animal hosts, but in the external environment-commonly on pasture. It is in this environment that the free-living stages are vulnerable to a range of abiotic factors (extremes in temperature and desiccation) and biotic factors (macro- and micro-organisms) that may decimate their numbers. Of the latter, there are organisms, which exert their effects either indirectly by rendering faecal deposits inimical for the development of nematode eggs through to infective larvae, or directly by acting as pathogens or by exploiting the free-living stages as a food source. Within this vast assemblage of organisms, which include microarthropods, protozoa, viruses, bacteria and fungi, could well emerge a variety of biological control agents of nematode parasites. At present, greatest interest lies with the nematode-destroying fungi. Work has progressed from Petri dishes, to plots, to paddocks with several species of the genus Arthrobotrys and Duddingtonia flagrans. These studies indicate that the voracious nematophagous capabilities of these fungi, clearly demonstrated in vitro, translate to reductions in the number of infective larvae on pasture and indicate that levels of control, comparable to conventional schemes using anthelmintics, can be achieved. The challenge now lies in developing methods of administration of fungi to animals which can be applied under practical farm conditions. However, the pursuit of candidates for biological control of nematode parasites of livestock should not involve just a few species of nematophagous fungi. More than 100 species of fungi have been identified as possessing nematode destroying capabilities. These need to be more specifically investigated for their effects on free-living stages of nematode parasites of livestock, together with other classes of organisms, particularly bacteria, which have proved successful as biological control agents of arthropods.

The potential of nematophagous fungi to control the free-living stages of nematode parasites of sheep: survey for the presence of fungi in fresh faeces of grazing livestock in Australia.

In the course of 12 months, 1742 fresh faecal samples from grazing livestock, principally ruminants, from various States of Australia were examined for the presence of nematophagous fungi. In total, 48 separate isolations were made representing various species from the genus Arthrobotrys and also 16 isolates of the single Duddingtonia species, Duddingtonia flagrans. These isolates will be used in continuing studies to select the most suitable species, on the basis of nematophagous capability and ruminant gut survival capacity, for development of a biological means of controlling nematode parasites of livestock.

The potential of nematophagous fungi to control the free-living stages of nematode parasites of sheep: in vitro and in vivo studies.

Following in vitro screening investigations on approximately 100 nematophagous fungi reported previously, eight species were selected for further investigation. Fungal elements (mycelium and conidia) were subjected to in vitro stress selection designed to simulate rumen and abomasal conditions. From these studies, three species, namely, Arthrobotrys oligospora, Arthrobotrys oviformis and Geniculifera eudermata, were selected for in vivo survival studies in sheep surgically fitted with abomasal and ileal cannulae. Doses of fungal conidia were administered orally or via the abomasal cannulae and samples of digesta were taken from the abomasum, the terminal ileum and faeces. The viability of the three fungal species at these sites was demonstrated. The abundance of fungi throughout the gut was dose-dependent but in all cases only very small volumes of fungal suspension containing unprotected conidia were used. These results demonstrate that a practical means of orally administering nematophagous fungi to control free-living stages of nematodes in faeces may become an achievable objective.

The potential of nematophagous fungi to control the free-living stages of nematode parasites of sheep: screening studies.

Ninety-four species of fungi with known nematophagous activity were tested for their ability to reduce the number of infective larvae of sheep nematodes in faecal cultures, and also for their ability to produce nematode-attractant and nematocidal substances against these free-living stages under in vitro conditions. Reductions of infective larval numbers exceeding 80% were consistently recorded when 100-250 conidia g-1 faeces of various species from the genera Arthrobotrys, Geniculifera and Monacrosporium were used. Even concentrations as low as 10 conidia g-1 faeces resulted in a significant reduction in infective larval numbers compared to control cultures. This study demonstrates that whilst many fungal species exhibit nematophagous activity against a variety of free-living nematodes, few show efficient activity against the free-living stages of parasitic nematodes in the sheep faecal environment. For the most active, there were six species of Arthrobotrys, two species of Geniculifera and two species of Monacrosporium which showed comparable activity to the extensively studied species A. oligospora.

Fungi causing thread mould spoilage of vacuum packaged Cheddar cheese during maturation.

Thread mould is a defect which occurs sporadically in maturing vacuum packaged Cheddar cheese, caused by the growth of fungi in folds and wrinkles of the plastic film in which the cheese is packaged. Fungi were isolated and identified from 110 Cheddar cheese blocks exhibiting typical thread mould defects. The major causative species were found to be Cladosporium cladosporioides, Penicillium commune, C. herbarum, P. glabrum and a Phoma species. Yeasts were also frequently isolated from the cheese, the majority belonging to the genus Candida. Fungal species which can cause thread mould defects were also found in the cheese factory environment, on cheesemaking equipment, in air, and in curd and whey, providing a wide range of potential sources of contamination.