Coccidiosis in hihi/stitchbirds (Notiomystis cincta) due to coccidia of the Eimeriidae.

AIM: To describe the pathology of coccidiosis in hihi and to provide preliminary data on the taxonomy of the coccidia involved using molecular methods. METHODS: In an initial study from 1994 to 1997, gross and histopathological examinations were performed on 12 dead juvenile hihi from the National Wildlife Centre (NWC) at Mt. Bruce. In a second study during 2008-2010 DNA from sporulated oocysts and liver tissue was used for PCR analysis and sequencing. Faecal samples were also obtained from infected hihi from the NWC and examined for coccidial oocysts, which were then sporulated in the laboratory in 1994-1997 and 2007-2009. In addition, a post mortem was performed on a dead adult hihi from the NWC in 2008, and 18 archived hihi tissues from 11 individual birds stored at the Institute of Veterinary, Animal and Biomedical Sciences (IVABS) were used for DNA extraction. RESULTS: Severe gross and histopathological changes in the intestine and occasionally in the liver were found in the 12 dead birds examined. The morphological characteristics of the sporulated oocysts suggested that two types of coccidia were present. PCR analysis and sequencing of extracted DNA supported the existence of at least two different coccidia species in hihi. These were genetically more closely related to the genus Eimeria than to the morphologically similar genus Cystisospora (formerly Isospora) of mammals. In addition, one liver tissue sample that was examined post mortem was positive for at least two different coccidia species of the family Eimeriidae according to sequencing results, and the presence of extraintestinal coccidian stages was confirmed. CONCLUSIONS: Preliminary morphological and sequencing results suggest that two types of eimeriid coccidia are present and at least one of these commonly has extraintestinal stages. CLINICAL RELEVANCE: Coccidiosis in hihi is a serious disease capable of causing mortalities in juvenile and adult birds in captive situations. Treatment and control of the disease will be difficult as the extraintestinal stages of the organism are likely to be refractile to oral treatment.

Serratospiculosis in a New Zealand Falcon (Falco novaeseelandiae).

CASE HISTORY: An adult New Zealand falcon was presented with metacarpal fractures in the left wing. CLINICAL FINDINGS: In addition to the fractures, radiographs revealed an area of opacity in the air sacs. A few days after hospitalisation and initiation of treatment of the fractures, the bird developed signs of respiratory disease; the area of opacity was found to have increased in size and density. Treatment with antibiotics and nebulisation was commenced; the bird initially responded but respiratory signs subsequently worsened and the bird died. At necropsy, air sacculitis and bronchopneumonia were associated with numerous nematodes in the air sacs, which were morphologically consistent with Serratospiculum guttatum. DIAGNOSIS: Serratospiculosis CLINICAL RELEVANCE: The discovery of this parasite and the associated disease for the first time in New Zealand indicates that it should be considered in the differential diagnosis of respiratory disease in falcons and possibly other raptors in New Zealand.

The effect of feeding sulla (Hedysarum coronarium) or lucerne (Medicago sativa) on lamb parasite burdens and development of immunity to gastrointestinal nematodes.

Two experiments investigated the efficacy of the legume Hedysarum coronarium (sulla), which contains condensed tannins (CTs), for reducing gastrointestinal nematode infections relative to lucerne. Experiment 1 was aimed to show whether the lower faecal egg count (FEC) and larval establishment previously reported in lambs grazing sulla were due to direct effects of the forage on Ostertagia circumcincta and Trichostrongylus colubriformis or were mediated through an enhanced immune response. Experiment 2 evaluated the impact of feeding sulla relative to feeding lucerne (Medicago sativa), before, at, or after larval challenge on subsequent FECs and nematode burdens. In experiment 1, 64 Romney lambs were fed either freshly cut lucerne or sulla (32 lambs per herbage) for the duration of the trial. Within each herbage there were four treatment groups (n=8 per group). Initial levels of immunity were assessed in uninfected (UN) lambs which were maintained parasite-free until challenged with 15,000 O. circumcincta and 15,000 T. colubriformis larvae on day 63, and slaughtered on day 81. The other three treatment groups were trickle-infected with each of 5000 O. circumcincta and 5000 T. colubriformis larvae three times per week from day 1 to 35. Non-steroid infected (CONTROL) and steroid-treated (STER) groups were treated with anthelmintic on day 49 and challenged with 15,000 O. circumcincta and 15,000 T. colubriformis on day 63 and slaughtered on day 81. The STER lambs were given dexamethasone trimethylacetate from day 49 to 81 to determine effects of immunity on parasite infection. From day 35 an establishment group (EST) on each herbage was fed a common pelleted lucerne diet and slaughtered on day 56 to determine nematode establishment during trickle-infection. Diet did not affect FECs but feeding lucerne increased (P<0.05) numbers of T. colubriformis in CONTROL lambs compared to those fed sulla. O. circumcincta numbers were lower (P<0.05) in UN lambs fed sulla than lucerne. The sulla diet was associated with higher (P<0.05) antibody titres against secretory-excretory antigens to adult O. circumcincta and to adult and larval T. colubriformis, so there appeared to be some immunogenic response to the sulla diet but effects upon T. colubriformis numbers were not significant. The second experiment involved 48 Romney lambs grazing conventional pasture which were infected with 10,000 each of O. circumcincta and T. colubriformis larvae either 7 days before, 7 days after, or at the time they commenced grazing either sulla or lucerne. Lambs which grazed sulla had lower (P<0.05) FEC and lower (P<0.05) O. circumcincta burdens than lambs which grazed lucerne but timing of infection had no effect on FEC or worm burdens. T. colubriformis numbers were not affected by treatment or herbage. In conclusion, the sulla diet resulted in lower O. circumcincta numbers compared to lucerne outdoors and some evidence of an immunogenic response was obtained indoors. However, neither the herbage nor the immunogenic response reduced T. colubriformis numbers in either experiment.

Nematodes and liver fluke in New Zealand.

A general review of the epidemiology, significance and control of nematode parasitism of sheep, goats, cattle and deer in New Zealand, the emergence of anthelmintic resistance and its effects, and the search for parasite control strategies that reduce reliance on anthelmintic use, is provided. The research that has formed the basis for present levels of understanding of this complex and important topic is summarised and sources of further information are indicated. Aspects of nematode infections of horses, pigs, dogs and cats, and the history of the liver fluke, Fasciola hepatica, in New Zealand are also reviewed briefly.

Effect of forage legumes containing condensed tannins on lungworm (Dictyocaulus sp.) and gastrointestinal parasitism in young red deer (Cervus elaphus).

To investigate the effect of feeding forage legumes containing condensed tannins (CT) on internal parasitism, red deer calves were fed either lucerne (Medicago sativa; 0.1 per cent CT), birdsfoot trefoil (Lotus corniculatus; 1.9 per cent CT) or sulla (Hedysarum coronarium; 3.5 per cent CT) and trickle-infected with deer-origin gastrointestinal nematode and lungworm (Dictyocaulus sp.) larvae for 5 weeks, then slaughtered at 7 weeks. There was a significant negative linear relationship between dietary CT concentration and abomasal nematode burdens. No significant differences in faecal egg counts, lungworm burdens or voluntary feed intake were found. Deer fed sulla had higher liveweight gain, carcass weight and carcass dressing-out percentage, higher serum total protein and albumin concentration and lower serum gastrin concentration and faecal lungworm larval count, compared with lucerne-fed deer. Inclusion of sulla in diets for young red deer may reduce the impact of internal parasites and/or reduce the dependence on anthelmintic treatment.

A model for study of lungworm (Dictyocaulus sp.) and gastrointestinal nematode infection in young red deer (Cervus elaphus).

A model of sub-clinical parasitism in young red deer, using concurrent trickle infections of lungworm (Dictyocaulus sp.) and mixed gastro-intestinal (GI) nematodes of deer-origin was evaluated. 20 parasite-free deer calves were artificially reared indoors from 4 days of age. A further five calves were naturally reared on pasture with their dams, treated with anthelmintic and brought indoors at 3-4 months. At 4-4.5 months of age they were individually housed and allocated to five groups (n=5). Groups were dosed 3 x per week, for 9 weeks with 0, 100 and 500, 200 and 1000 (2 groups), 400 and 2000 infective larvae of lungworm and mixed GI nematodes, respectively, cultured from deer faeces. Liveweight and voluntary feed intake measurements and faecal and blood samples were taken weekly. In the fourth week following cessation of trickle infection, deer were euthanased and lung and GI nematodes recovered. Both lungworm and GI nematode infections became patent at Week 4 of infection. Maximum group arithmetic mean faecal egg counts were 100-190 epg. Maximum group arithmetic mean faecal lungworm larval counts were 58-123 lpg. Group arithmetic mean nematode counts at slaughter ranged from 439-806 for GI nematodes and 31-73 for lungworm, respectively. Despite low nematode counts, reduced liveweight gain, voluntary feed intake and serum albumin concentration, elevated serum pepsinogen, gastrin and globulin concentrations and elevated peripheral eosinophil counts and slight haemoconcentration, but no clinical signs, were observed. The reduction in liveweight gain was related to the reduction in voluntary feed intake (r2=0.83; p<0.088). Naturally-reared deer had similar liveweight gains, voluntary feed intake and nematode counts to artificially-reared deer. Thus, methods of infection to produce concurrent sub-clinical lungworm and GI nematode burdens for study of sub-clinical parasitism in young deer have been defined.

Serological study of a dairy herd with a recent history of Neospora abortion.

UNLABELLED: HERD HISTORY: A cluster of 19 abortions occurred in April and May 1994 on a seasonal dairy farm of 277 cows in Taranaki. Lesions consistent with neosporosis were seen in several foetuses. The age of the aborting cows ranged from 2 to 12 years. Pregnant rising 2-year-old heifers had been grazed off the main farm on a run-off' until returning in May, when they were replaced on the run-off by the rising l-year-old calves. None of these pregnant heifers aborted. METHODS: Sera were obtained from all calves (n = 32), 32 of 33 pregnant heifers, 59 of 60 3-4 year-old cows and 101 of 150 older cows, plus all cows that had aborted. Sera were tested using an indirect fluorescent antibody assay. Non-aborting cattle sera were screened at 1:400, and further dilutions were tested from 15 aborted cows. RESULTS: The percentages of seropositive cattle were: 32% of the calves, 3% of the heifers, 31% of the younger cows and 27% of the older cows. The proportion of seropositive heifers was significantly lower (p< 0.01) than in the other age classes. There was no significant difference in the proportion of seropositive animals between these other age classes (p >0.05). Of the 15 cows that aborted, and for which further dilutions were tested, nine had titres of 1: 12,800, four had titres of 1:6400, one had a titre of 1: 1600 and one had a titre of 1:400. Twenty cows were the dams of pregnant heifers. Six of these cows were seropositive but all their offspring were seronegative. Fifteen aborting cows were held over to the following year when 13 became pregnant and calved normally. There were only two to three abortions in the whole herd in this following year. CONCLUSION: These results suggest that either a point-infection occurred sometime in early 1994 with an infective period short enough so that the heifers did not get infected when they returned to the main farm, or some factor precipitated a recrudescence of latent infection with a subsequent rise in titres in about 29% of the animals on. the main farm only. Although neither hypothesis can be proven or disproven in this observational study, the former hypothesis seems more likely.

Variation between hosts in the developmental success of the free-living stages of trichostrongyle infections of sheep.

Variation between hosts of different ages and genotypes in the developmental success of trichostrongylid parasite eggs from sheep was investigated in two trials. The percentage development to infective third-stage larvae of eggs collected from lambs and adult ewes infected with Ostertagia circumcincta was compared in an indoor trial. In addition, sheep previously bred for either high or low faecal egg count and grazed outdoors on parasite contaminated pasture were sampled; egg development, faecal egg count, generic profile and faecal dry matter were measured. In both trials, development to L3 was significantly lower in eggs derived from adult ewes than from lambs and, in the field trial, from animals selected for low faecal egg count. The observed differences could not be accounted for by variations in faecal egg count, faecal dry matter content nor by differences in the generic composition of worm egg output. Although not shown conclusively, the results are consistent with an immune mechanism influencing the development of free-living stages of gastrointestinal nematodes outside the host. These results have significant implications for our understanding of parasite epidemiology, particularly as it relates to sources of pasture contamination.

Establishment and fecundity of Ostertagia circumcincta and Trichostrongylus colubriformis in lambs fed lotus (Lotus pedunculatus) or perennial ryegrass.

Forty four 12-14 week old Polled Dorset ram lambs, raised helminth free from birth, were used to investigate the effects of condensed tannins (CT) in lotus (Lotus pedunculatus) on lamb growth and gastrointestinal nematode establishment and fecundity. Condensed tannins bind to proteins in the rumen and increase the flow of protein to the intestines. Lambs were allocated to either a ryegrass (Lolium perenne) or lotus diet fed ad libitum, with or without twice daily drenching of polyethylene glycol (PEG) which binds with and deactivates the CT. One week after allocation to the diets, each lamb was infected per os with 10,000 Ostertagia circumcincta and 10,000 Trichostrongylus colubriformis larvae and slaughtered 28 or 29 days later. Lambs fed lotus had lower faecal egg counts (P < 0.0001) and lower O. circumcincta burdens (P < 0.001), fewer female O. circumcincta (P < 0.0001) and higher faecal dry matter (P < 0.001) than lambs fed ryegrass, but numbers of T. colubriformis nematodes were not affected. This trial did not ascribe beneficial effects of L. pedunculatus to CT per se and the use of PEG does not seem appropriate to studies of parasitology.

Effect of plant species on the larvae of gastrointestinal nematodes which parasitise sheep.

Faeces containing Trichostrongylus colubriformis and/or Ostertagia circumcincta eggs were used to provide four contaminations in each of 2 years on plots of browntop, Yorkshire fog, ryegrass, tall fescue, lucerne, chicory, cocksfoot, white clover, and prairie grass and in the second year a mixed sward of ryegrass/white clover. Third stage larvae were recovered from faeces and from four strata of herbage, 0-2.5, 2.5-5, 5-7.5 and > 7.5 cm above the soil surface at 2, 4, 6, 8, 11, and 14 weeks after faeces were deposited on the swards. Herbage species had a significant (P < 0.0001) effect on the number of larvae recovered. Greatest numbers of larvae, as indicated by ranking analysis, were recovered from Yorkshire fog, ryegrass, and cocksfoot and lowest numbers from white clover and lucerne. The difference between herbages in numbers of larvae recovered was due to the "development success", the ability of larvae to develop to the infective stage and migrate on to herbage, rather than "survival", the rate of population decline once on the herbage. Faecal degradation was most rapid from white clover and browntop, intermediate from tall fescue, lucerne, prairie grass, cocksfoot, and ryegrass, and slowest from Yorkshire fog swards. The numbers of larvae recovered from herbages were related (r2 = 0.59, P < 0.05) with the faecal mass remaining. A greater proportion of the total larvae recovered from the herbage was recovered from the bottom stratum of Yorkshire fog and prairie grass than from white clover, with the other herbages intermediate, indicating that larvae had greater difficulty migrating up Yorkshire fog and prairie grass than the other herbage species. In most herbage species, despite more larvae being recovered from the lowest stratum, larval density (L3/kg herbage DM) was highest in the top stratum. This study has demonstrated that herbage species can have a significant impact on the population dynamics and vertical migration of T. colubriformis and O. circumcincta larvae.

Production, faecal egg counts and worm burdens of ewe lambs which grazed six contrasting forages.

Lambs were grazed on monospecific swards of one of six forages, some containing condensed tannins (CT); sulla (Hedysarum coronarium), Lotus corniculatus and L. pedunculatus and some without CT (lucerne (Medicago sativa), plantain (Plantago lanceolata) and a ryegrass/white clover pasture (Lolium perenne/Trifolium repens)) to evaluate effects of forage type on gastrointestinal nematode burdens, faecal egg count as well as liveweight gain (LWG), wool growth and dag formation over a 42-day period. A new allotment of feed was provided at seven-day intervals, and feed was available ad libitum. Total worm burdens in a pre-treatment slaughter group were ca. 22000 of which ca. 5000 were in the abomasum. Within each forage type, 10 of the 25 lambs (NP) were given an anthelmintic drench at 14-day intervals and the remaining 15 lambs (P) were not drenched. Daily gains of the NP lambs were double that of P lambs which grazed either L. corniculatus, lucerne or pasture, but parasitism had a lesser effect on performance of lambs which grazed sulla and L. pedunculatus. Plantain was not palatable and all lambs performed poorly. Highest daily gains in P lambs were for those which grazed sulla (175 g/day) and L. pedunculatus (160 g/day) with total worm burdens of 13100 and 23000 for the respective treatment groups. The other forages resulted in lower daily LWG in P lambs, and performance was not related to either worm burdens or worm species. This experiment has shown that, when sulla is fed, there is a reduction in worm burdens and faecal egg count (FEC), but with Maku lotus which also contains CT, the good level of performance was achieved despite in high worm burdens and FEC. The mechanisms by which these forages enable high levels of productivity in the face of a parasitism appear to differ, but both could be incorporated into forage feeding systems to reduce dependence on anthelmintic drenches.

Evaluation of a larval development assay for the detection of anthelmintic resistance in Ostertagia circumcincta.

Two experiments were carried out to evaluate a larval development assay for the detection of anthelmintic resistance in O. circumcincta. In Experiment I, the dose responses to levamisole (LEV), thiabendazole (TBZ) and ivermectin (IVM) of 8 isolates of O. circumcincta were measured 34 days after infection (DAI). Four of these isolates were shown to be resistant to 1 or more anthelmintics. With 2 exceptions, all isolates considered to be resistant had higher LD50 values than the susceptible isolates for that anthelmintic. One exception was isolate RM8, which was considered to be resistant to all 3 anthelmintics based on faecal egg count reduction tests in goats, but the LD50 value for LEV did not differ from that for the susceptible isolates. The other exception was an isolate considered to be susceptible to TBZ which had a relatively high LD50 value. In an unrelated trial that was prompted by this finding, this isolate was confirmed to be benzimidazole-resistant. Isolate RM8 and an isolate susceptible to all 3 anthelmintics (SK2) were used in the second experiment, which was conducted to monitor changes in the LD50 values of LEV, TBZ and IVM over time following a single infection of 35,000 infective larvae in young sheep. Faecal samples were collected weekly from 24 to 115 DAI. With all 3 anthelmintics, the LD50 values increased with time to a peak around 50-60 DAI, and then declined to levels similar to those observed soon after patency. This trend was consistent for both isolates. The highest mean LD50 values for isolates SK2 for IVM and TBZ and RM8 for IVM and RM8, respectively, were 1.7 and 1.8 times, and 2.2 and 2.9 times higher than the initial mean LD50 values. There was a clear distinction in LD50 values between isolates at each sampling day for both IVM and TBZ. However, as a consequence of the changes in LD50 values with time, the peak LD50 values of IVM for isolate SK2 were higher than the minimum LD50 values of isolate RM8. As there was no apparent difference in LEV efficacy between these 2 isolates, the data were pooled. The highest mean LD50 value was 2.3 times higher than the initial LD50 value.

Protozoal encephalomyelitis of dogs involving Neospora caninum and Toxoplasma gondii in New Zealand.

In a retrospective study of 15 cases of encephalomyelitis in dogs, three cases of Neospora caninum and two cases of Toxoplasma gondii infection were identified using immunohistochemical staining of central nervous system sections. All cases of neosporosis showed ataxia and progressive hind limb paralysis due to multifocal non-suppurative meningoencephalomyelitis which was most severe in the spinal cord and base of the brain stem. Neospora tissue cysts could not be distinguished morphologically from those of T. gondii using light microscopy, but electron microscopy confirmed their characteristic features. Although Neospora abortion in cattle has only recently been recognised in New Zealand, this study has shown that neosporosis has been present in dogs since at least 1972.

Controlling internal parasites in grazing ruminants without recourse to anthelmintics: approaches, experiences and prospects.

Interest in ways of raising stock without using anthelmintics has been stimulated by the desire for intensive grazing systems to adopt more sustainable methods of internal parasite control and by potential premiums for "organic" produce. This requires grazing management systems which are practicable and achieve levels of parasite control sufficient to meet realistic production objectives. This paper will summarise 3 years experience of lamb and cattle production without a recourse to anthelmintics on 2 production systems: a mixed cropping, lamb finishing, and cattle rearing unit and an all grass, mixed-stock, hill country unit in which all non-replacement lambs are sold at weaning. On both units parasite control was almost entirely dependent on integrated grazing management of sheep and cattle and strict systems of grazing management had to be rigidly applied. Acceptable productivity could be more readily achieved in sheep than cattle. Rams selected for resistance to nematodes were also used in both sheep flocks. Their influence on lamb production was equivocal. There is a need for more information on factors influencing parasite epidemiology and for consideration of strategies other than alternate grazing. The impact of pasture species on parasite epidemiology needs to be clarified. Current research indicates substantial differences between grasses in terms of parasite burdens acquired and production losses suffered by lambs grazing them. Grazing management may need to vary with pasture species. Also, specialty forage crops, particularly those containing condensed tannins (i.e. Hedysarum coronarium, Lotus pedunculatus and Lotus corniculatus) hold special promise as a means of countering parasite-induced production losses and dagginess. Using biological control of free-living larval stages and vaccination, may in the long term, also prove useful. Developing effective and acceptable systems for raising stock without using anthelmintics presents a considerable challenge, to parasitologists, as well as to plant breeders, agronomists, and farming systems researchers.

Efficacy of ivermectin against benzimidazole-resistant Nematodirus spathiger in sheep.

Eighteen helminth-free lambs were randomly allocated to six groups of three. Each lamb was dosed with 3300 infective larvae pooled from two isolates of Nematodirus spathiger known to be benzimidazole resistant. One lamb from each group was treated with oral ivermectin, one with oral oxfendazole and one left untreated 21 days after infection. All lambs were humanely killed 14 days later and small intestine worm counts performed. No Nematodirus were found in the ivermectin-treated lambs. Nematodirus numbers were reduced by 13% in oxfendazole-treated lambs relative to the control lambs.

Evaluation of the efficacy of doramectin against an artificial infection of Dictyocaulus viviparus in calves.

Three groups of eight Friesian calves, reared parasite-free, were experimentally infected with 1000 infective larvae of Dictyocaulus viviparus. Two groups were injected subcutaneously with 1% doramectin at 0.2 mg/kg body weight, one group 5 days after infection and the other 25 days after infection. A third group served as untreated controls. Faecal samples were examined for lungworm larvae on days 28, 32, 33, 34 and 35 after infection; the calves were killed and necropsied 39 or 40 days after infection and any lungworms present recovered and counted. Doramectin proved 100% effective against both 5-day-old and mature D. viviparus infections.

The outdoor survival of Sarcocystis gigantea sporocysts.

Sporocysts of Sarcocystis gigantea in cat faeces were placed in vented polystyrene tubes and superficially buried in both open and shaded sites. Their survival, as measured by their ability to excyst in vitro, was then monitored during the course of two separate experiments extending over 12-month periods. The results showed that the viability of these sporocysts declined most rapidly over the summer months and suggested that they were unlikely to remain infective for more than 1 year.

Resistance against Taenia hydatigena in sheep after passive transfer of serum or colostrum.

The role of antibody in the resistance of sheep to infection with Taenia hydatigena metacestodes was examined using passive transfer of immunoglobulin. The immunoglobulin either was experimentally transferred in serum, or was transferred from immune ewes to their new-born lambs in colostrum. Pooled serum from donor lambs which had received one, light, oral infection did not protect recipients although the donors themselves were immune. However, transfer of pooled serum from donors which had either received three oral infections, or three immunizations with solubilized T. hydatigena oncospheres in a water-in-oil adjuvant, resulted in 70-80% fewer cysts in the recipients. Colostrum from ewes infected with three high or low doses of T. hydatigena eggs was transferred to their lambs. A short acting protection (one to three weeks) was observed in the lambs. Comparisons by ELISA and Western blot, of the anti-T. hydatigena oncosphere antibody content of the donor sera, the sera of the recipients collected 24 h and seven days after transfer, the sera of the lambs and ewes, and the colostrum of the ewes, indicated that resistance to the challenge infection depends upon a critical level of antibody.

The survival of Sarcocystis gigantea sporocysts following exposure to various chemical and physical agents.

Using in vitro excystation as a measure of viability, it was found that at 4 degrees C Sarcocystis gigantea sporocysts survived considerably better in tap water (85% excystation after 174 days) than in either 2.5% potassium dichromate (15% excystation after 174 days) or 2% sulphuric acid (0% excystation after 5 days). Although they were able to resist 48 h suspension at room temperature in most laboratory reagents and disinfectants tested, six (sulphuric acid, ammonia, methanol, ethanol, potassium hydroxide, sodium hydroxide, Medol) had substantial sporocysticidal properties. Further investigation with three of these showed that sporocyst excystation was reduced from 65% to less than 10% following contact with 2.5% sulphuric acid for 1 h or with 2% ammonia or 4% Medol for 4 h. Sporocysts were either killed or had their ability to excyst severely impaired by heating to 60 degrees C and 55 degrees C for 5 and 60 min, respectively, by exposure to ultraviolet radiation at a dose of 4000 ET, or by prolonged storage in water at 24 degrees C. Sporocysts exposed to either constant or intermittent freezing at -18 degrees C suffered a comparatively slow decline in excystation rate with time, as did those subjected to desiccation. The duration of survival of desiccated sporocysts was inversely related to relative humidity and after 245 days at 33% relative humidity and temperatures of 15 degrees C or 24 degrees C, 60% of such sporocysts excysted.