Direct measurement of dispersal of Dictyocaulus viviparus in sporangia of Pilobolus species

Dispersal of Dictyocaulus viviparus larvae by Pilobolus sporangia was studied on 29 faecal pats deposited between the end of June and late October 1988. Faecal pats were covered daily from day 3 to 4 after deposition with a large petri dish to measure the numbers of sporangia released and the numbers of larvae carried. The yield of both was variable. Dispersal of lungworm larvae was lowest on over-grazed pasture or when Pilobolus growth was very poor. When faecal pats were sheltered by a long sward, 17 per cent or more of larvae present at deposition were transported in this manner. In July and August, peak dispersal of lungworm larvae was on day 5, in September on day 6 and in October on day 7, the increasing time intervals being probably associated with decreasing temperature.     

Spread of equine lungworm (Dictyocaulus arnfieldi) larvae from faeces by Pilobolus fungi

Between 10 and 25% of the Dictyocaulus arnfieldi larvae excreted in faeces from a naturally infected donkey were harvested as infective stages from faecal cultures by means of Pilobolus fungi. The faeces were collected between 24 and 56 hours after drenching the donor animal with Pilobolus spores and kept at 16 +/- 2 degrees C. Most larvae were collected between the 5th and the 8th day of culturing during which period fructification and sporangium discharge also peaked. The sporangia and the adhering larvae were collected in Petri dishes inserted between the faecal mass and a light source. All recovered larvae were viable. A mean larval length of 368 microns (range 312-440 microns) and width of 14.6 microns (range 12-20 microns) was recorded for the infective stage. The method was found suitable for the recovery of infective stages for experimental purposes. The authors suggest that the Pilobolus mechanism play an important part in the spread of equine lungworm infection under field conditions similar to the situation in bovine lungworm (Dictyocaulus viviparus) infection.     

The efficacy of two isolates of the nematode-trapping fungus Duddingtonia flagrans against Dictyocaulus viviparus larvae in faeces.

A series of experiments was carried out to examine the effects of two different isolates of the nematode-trapping fungus Duddingtonia flagrans to reduce the number of free-living larvae of the bovine lungworm, Dictyocaulus viviparus. A laboratory dose-titration assay showed that isolates CI3 and Troll A of D. flagrans significantly reduced (P < 0.05 to P < 0.001) the number of infective D. viviparus larvae in cultures at dose-levels of 6250 and 12,500 chlamydospores/g of faeces. The larval reduction capacity was significantly higher for Troll A compared to CI3 when lungworm larvae were mixed in faecal cultures with eggs of Cooperia oncophora or Ostertagia ostertagi and treated with 6250 chlamydospores/g of faeces. Both fungal isolates showed a stronger effect on gastrointestinal larvae than on lungworm larvae. Two plot trials conducted in 1996 and 1997 involved deposition of artificial faecal pats containing free-living stages of D. viviparus and C. oncophora on grass plots. Herbage around the pats was collected at regular intervals and infective larvae recovered, counted and identified. These experiments showed that both D. flagrans isolates reduced the number of gastrointestinal as well as lungworm larvae in faecal pats. During both plot trials, the transmission of C. oncophora larvae, but not D. viviparus, from faecal pats to the surrounding herbage was clearly affected by climatic conditions. After collection of faecal pats from the grass plots one month after deposition, the wet and dry weight of pats as well as organic matter content were determined. No differences were found between the fungus-treated and non-treated control pats. This indicated that the rate of degradation of faeces was not affected by the addition of the fungus.     

Spread of lungworm (Dictyocaulus viviparus) infection by Pilobolus fungi among stabled calves

Pilobolus kleinii spores isolated from cattle faeces were administered daily by the oral route to two donor calves excreting Dictyocaulus viviparus larvae in their faeces.Faeces were collected at daily intervals from the donor calves, and on each day a pooled sample of 500 g was placed outside a pen containing two parasite-free experimental calves. A wire screen prevented the experimental calves from coming into contact with the artificially deposited faecal portions.Pilobolus kleinii fruit bodies were observed on the faecal surface in large numbers 5–10 days after deposition.Sporangial discharge was observed to be directed against a window which served as the only light source. Sporangia were found up to 110 cm away from the faeces, and it was estimated that at least 29% of them had been transmitted to a feeding trough placed close to the wire screen. Infective D. viviparus larvae were observed on some of the discharged sporangia.The experimental calves started excreting low numbers of D. viviparus larvae in their faeces 40–49 days after the first portions were deposited. These results indicate that P. kleinii was reponsible for the infection of the experimental calves with D. viviparus, through the discharge of sporangia carrying infective lungworm larvae.     

Seasonal translation of infective larvae of gastrointestinal nematodes of cattle and the effect of Duddingtonia flagrans: a 3-year pilot study

A study was conducted over 3 years (1998-2000) to investigate larval availability of gastrointestinal nematodes from faeces of cattle reared under different parasite control schemes. These cattle were part of a parallel, but separate grazing trial, and were used as donor animals for the faecal material used in this experiment. At monthly intervals, faeces were collected and pooled from three groups of first-season grazing cattle. These groups were either untreated, ivermectin bolus treated or fed the nematophagous fungus Duddingtonia flagrans. The untreated and fungus treated animals were infected with gastrointestinal nematodes and the number of eggs per gram (epg) pooled faeces ranged between 50 and 700 in the untreated group and between 25 and 525 epg in the fungus treated group. Each year between June and September, artificial 1 kg dung pats were prepared and deposited on pasture and protected from birds. The same treatments, deposition times and locations were repeated throughout the study. Larval recovery from herbage of an entire circular area surrounding the dung pats was made in a sequential fashion. This was achieved by clipping samples in replicate 1/4 sectors around the dung pats 4, 6, 8 and 10 weeks after deposition. In addition, coinciding with the usual time of livestock turn-out in early May of the following year, grass samples were taken from a circular area centred where the dung pats had been located to estimate the number of overwintered larvae, which had not been harvested during the intensive grass sampling the previous year. It was found that recovery and number of infective larvae varied considerably within and between seasons. Although the faecal egg counts in 1999 never exceeded 300 epg of the faecal pats derived from the untreated animals, the abnormally dry conditions of this year generated the highest level of overwintered larvae found on herbage in early May 2000, for the 3 years of the study. Overall, biological control with D. flagrans significantly reduced larval availability on herbage, both during and between the grazing seasons, when compared with the untreated control. However, the fungus did not significantly reduce overwintered larvae derived from early season depositions (June and July), particularly when dung pats disappeared within 2 weeks after deposition. Very low number of larvae (<3 per kg dry herbage) were sporadically recovered from grass samples surrounding the ivermectin bolus faecal pats.     

Spread of infective Dictyocaulus viviparus larvae in pasture and to grazing cattle: Experimental evidence of the role of Pilobolus fungi

Four calves experimentally infected with Dictyocaulus viviparus were made Pilobolus-free by hygienic measures and by feeding them irradiation sterilized feed. Two of the calves were orally administered laboratory cultured Pilobolus sporangia daily. As a result, the faeces from one air contained D. viviparus larvae and Pilobolus spores, and the faeces from the other pair contained D. viviparus larvae, but no Pilobolus spores.

Two identical plots were used for deposition of the two kinds of faeces, and one of them remained free of Pilobolus fructification. Herbage sampling and the use of tracer calves revealed that on this plot the larval contamination and the infectivity of the pasture were greatly reduced. A mean larval count of 1321 near the faecal pats (0–5 cm) in the plot where Pilobolus was observed was reduced to 69 per kg of herbage on the Pilobolus-free plot. At a distance of 100 cm from the pats, a reduction from 99 to 3 larvae per kg herbage was found.

Each plot was grazed by four parasite-free tracer calves for 3 days. During the subsequent stabling period of these calves, the lungworm larval excretion of those from the Pilobolus-free plot was reduced by 90% and the clinical symptoms were milder than those which grazed the plot which contained the fungus. The mean post mortem worm counts after 4 weeks of stabling showed a reduction from 167 to 25 worms.

A more marked effect of Pilobolus fungi on the transmission of D. viviparus infection is to be expected under field conditions where calves are grazing more selectively than in the present study.     

Dispersal of Dictyocaulus viviparus larvae from bovine faeces in Ireland

An involvement of Pilobolus species fungus in the dispersal of Dictyocaulus viviparus third stage larvae from dung to surrounding herbage under Irish conditions was investigated. The presence of Pilobolus kleinii on artificial dung pats containing first stage larvae of D viviparus was associated with a 19-fold increase (P less than 0.05) in numbers of third stage larvae recovered from the surrounding herbage. A subjective examination of natural dung pats showed that the presence of Pilobolus species was significantly correlated with hours of bright sunshine (r = -0.5, P less than 0.01), total rainfall (r = 0.41, P less than 0.05) and the height of herbage surrounding the pats (r = 0.31, P less than 0.001). A multiple regression analysis showed that meteorological parameters and the height of surrounding herbage accounted for 38 per cent of the variation in growth of Pilobolus species on dung pats. The incidence of extensive damage to natural dung pats within five days of deposition, caused by biotic factors, another possible cause of D viviparus third stage larvae dispersal, varied from 0 to 92 per cent of the pats depending on their degree of dryness.     

Epidemiology of bovine dictyocaulosis in Denmark

A field experiment was conducted over two grazing seaons with calves on a permanent pasture in order to follow the pattern of infection with Dictyocaulus viviparus. Infective larvae persisted during the first, but not during the second, winter of observation. By means of the agar-bile herbage technique, a moderate first peak of infection was demonstrated in the pasture 2–3 weeks before the appearance of respiratory signs in the calves. Fluctuations in faecal larval output were reflected in the herbage contamination with infective larvae close to faecal pats. This, as well as the horizontal dispersion of larvae in the pasture, took place in less than a week. The proportion of lungworm larvae recovered away from faeces was low during a period of dry and hot weather while herbage sampling at two-hour intervals during two days showed an increase in herbage contamination with lungworm larvae, but not with trichostrongyle larvae between 10 a.m. and 12 noon.The infectivity of the pasture was monitored by tracer calves and compared with the results of the pasture sampling. The general course of the infection in the calves and in the pasture was the results of interaction between them. In addition, the pasture infection was influenced by climate and the infection in the calves by the development of immunity. The course of infection in individuals appeared to have an influence on the general course of the infection through the contamination of the pasture.     

A new isolate of the nematophagous fungus Duddingtonia flagrans a biological control agent against free-living larvae of horse strongyles

An experiment was carried out in 1997 to test the efficacy of an isolate of the microfungus Duddingtonia flagrans against free-living stages of horse strongyles under conditions in the field and to assess the eventual effect of the fungus on the normal degradation of faeces. Faecal pats were made from faeces of a naturally strongyle infected horse, which had been fed fungal material at a dose level of 106 fungal unit/kg bwt. Control pats without fungi were made from faeces collected from the same animal just before being fed fungi. Faecal cultures set up for both groups of faeces to monitor the activity of the fungus under laboratory conditions showed that the fungus significantly reduced the number of infective third-stage larvae (L3) by an average of 98.4%. Five faecal pats from each batch of faeces were deposited on pasture plots at 3 times during spring-summer. The herbage around each pat was sampled fortnightly to recover L3 transmitted from faeces. The results showed that the herbage infectivity around fungus-treated pats was reduced by 85.8-99.4%. The remaining faecal material at the end of each sampling period was collected, and the surviving L3 were extracted. Significantly fewer larvae were recovered from the fungus-treated pats. Analysis of wet and dry weight of the collected pats, as well as their organic matter content, were performed to compare the degradation of faeces of both groups. The results indicated that the presence of the fungus did not alter the degradation of the faeces.     

The origin and overwintering survival of the free living stages of cattle parasites in Sweden

During the 1997 Swedish grazing season, faeces were collected every 3 weeks on 7 occasions from young grazing cattle with moderate nematode parasite infections. From this source 12, 400 g dung pats were set up on each sampling occasion on a specially designated area of pasture. Half of these pats were placed on pasture where it was aimed to prevent snow cover during the subsequent winter. During the grazing season, herbage growth was kept at reasonably uniform height by clipping and the dung pats were protected from destruction by animals and birds. At the time of animal turn-out the following year (7th April 1998), it was observed that all dung pats had disappeared. Assessments of the survival of infective larvae, both on pasture and in soil, were made in a circular area encompassing the location of each pat. These sampling procedures were completed within a 3 week period. All faecal deposits yielded infective larvae at turn-out the following year, with proportionally greater numbers developing from nematode eggs deposited in cattle dung during the mid third of the previous grazing season. The surface layer of soil was found to be an important reservoir for infective larvae, with numbers recovered being approximately half those found in the overlying pasture samples. No significant differences were found between the normal pasture and snow excluded pasture in the number of infective larvae recovered from both pasture and soil samples. The epidemiological consequences of these findings are discussed.     

Observations on the free-living stages of cattle gastrointestinal nematodes

A 4-year study on the free-living stages of cattle gastrointestinal nematodes was conducted to determine (a) the development time from egg to infective larvae (L3) inside the faecal pats, (b) the pasture infectivity levels over time, and (c) the survival of L3 on pasture. Naturally infected calves were allowed to contaminate 16 plots on monthly basis. Weekly monitoring of eggs per gram of faeces (epg) values and faecal cultures from these animals provided data for the contamination patterns and the relative nematode population composition. At the same time, faecal pats were shaped and deposited monthly onto herbage and sampled weekly to determine the development time from egg to L3. Herbage samples were collected fortnightly over a 16-month period after deposition to evaluate the pasture larval infectivity and survival of L3 over time. The development time from egg to L3 was 1-2 weeks in summer, 3-5 weeks in autumn, 4-6 weeks in winter, and 1-4 weeks in spring. The levels of contamination and pasture infectivity showed a clear seasonality during autumn-winter and spring, whilst a high mortality of larvae on pasture occurred in summer. Ostertagia spp., Cooperia spp. and Trichostrongylus spp. were predominant and a survival of L3 on pasture over a 1-year period was recorded in this study.     

Disintegration of dung pats from cattle treated with the ivermectin anthelmintic bolus, or the biocontrol agent Duddingtonia flagrans

An experiment was performed during the grazing seasons of 1998, 1999 and 2000 to study the influence of the antiparasitic drug ivermectin and the nematophagous fungus Duddingtonia flagrans on cattle dung disintegration. The faeces originated from groups of animals that were part of a separate grazing experiment where different control strategies for nematode parasite infections were investigated. Each group consisted of 10 first-season grazing cattle that were either untreated, treated with the ivermectin sustained-release bolus, or fed chlamydospores of D. flagrans. Faeces were collected monthly on 4 occasions and out of pooled faeces from each group, 4 artificial 1 kg dung pats were prepared and deposited on nylon mesh on an enclosed pasture and protected from birds. The position of the new set of pats was repeated throughout the 3 years of the study. Each year, the dung pats were weighed 4, 6, 8 and 10 weeks after deposition and immediately afterwards replaced to their initial positions. Results showed that there was no difference in faecal pat disintegration between groups. However, the time-lag between deposition and complete disintegration of the faeces varied significantly between deposition occasions. Dung pats disappeared within 2 weeks (visual observation) when subjected to heavy rainfall early after deposition, whereas an extended dry period coincided with faeces still remaining 12 months after deposition.     

Factors affecting recovery of Dictyocaulus viviparus third stage larvae from herbage and growth of Pilobolus on dung pats

Significantly greater Dictyocaulus viviparus stage three larval recoveries from herbage samples adjoining first stage larval infected dung pats with Pilobolus, and the effects of biotic and mechanical factors on dung pat integrity supported previous findings. Several meteorological factors including sunshine hours, relative humidity, rain total and wind speed showed significant correlations with growth of Pilobolus on dung surfaces. The influence of monitored variables on the recovery of third stage larvae from herbage and the growth of Pilobolus was modified by height of the sward surrounding the dung pat. Multiple regression analysis showed that 60 per cent of variation in D viviparus third stage larvae recovery from herbage was accounted for by known variables.     

Survival of infective Ostertagia ostertagi larvae on pasture plots under different simulated grazing conditions

This study was carried out to examine the survival of infective Ostertagia ostertagi larvae (L(3)) on pasture under different simulated conditions of grazing, i.e. mixed grazing of cattle and nose-ringed sows, or grazing by cattle alone. Standardised pats of cattle faeces containing O. ostertagi eggs were deposited on three types of herbage plots, which were divided into zone 1: faecal pat; zone 2: a circle extending 25cm from the edge of the faecal pat; zone 3: a circle extending 25cm from the edge of zone 2. For "tall herbage" (TH) plots, the herbage in zone 2 was allowed to grow naturally, while the herbage in zone 3 was cut down to 5-7cm fortnightly, imitating a cattle-only pasture. For "short herbage" (SH) plots, the herbage in both zones 2 and 3 were cut down to 5-7cm fortnightly, imitating mixed grazing of cattle and sows. The grass in the "short herbage and scattered faeces" (SH/SF) plots were cut as for SH plots, and the faeces were broken down 3 weeks after deposition and scattered within zone 2, imitating the rooting behaviour of co-grazing sows. Five faecal pats from each plot group were collected on monthly basis, along with the herbage from zones 2 and 3 cut down to the ground. Infective larvae were then recovered from both faeces and herbage. The numbers of L(3) recovered from zone 1 were higher in the TH plots than in the other two groups and, furthermore, the larval counts from SH plots were always higher than from SH/SF plots. The three groups followed a similar pattern during the season regarding numbers of L(3) in zone 2, and no clear patterns between plot types were obtained. The presence of L(3) in zone 3 was almost negligible. Important differences were seen throughout the study from the biological point of view; more L(3) were able to survive in faeces on the TH plots, presumably reflecting a better protection from heat and desiccation compared to those in the other plots. The overall results support the idea that mixed grazing of cattle and pigs favour the reduction of O. ostertagi larval levels in pasture. This reduction is mainly due to the grazing behaviour of pigs, which by grazing up to the very edge of the cattle faeces, will either expose the larvae in faeces to adverse environmental summer conditions or ingest cattle parasite larvae, or both.     

Viability and pattern of emergence of Dictyocaulus viviparus larvae in sporangia of the fungus Pilobolus kleinii

Recoveries of third stage Dicytocaulus viviparus larvae (L3) from Pilobolus species sporangia ranged from 23 per cent at 21 days to 3 per cent after 90 days for sporangia attached to polythene discs positioned on pasture. There was a continuous release of L3 for up to 16 days from sporangia which were placed under conditions simulating those occurring on pasture.     

Development and survival of infective larvae of gastrointestinal nematodes of cattle on pasture in central Kenya

On a series of pasture plots, 2 kg pats of bovine faeces containing known numbers of strongylid (Haemonchus, Cooperia, Oesophagostomum and Trichostrongylus) eggs were deposited at intervals of 4 weeks from July 1995 to June 1996. The plots were sampled every 2 weeks after contamination and infective larvae were identified and counted. Larvae of all the genera developed throughout the year, but the pats exposed during the rainy season yielded more abundant larvae on the herbage. Irrespective of the season of deposition of the pats, larvae were found in larger numbers from 2 to 6 weeks after deposition and generally declined to below detectable levels within 12 to 16 weeks of contamination. The comparatively short survival times noted in this experiment may present opportunities for manipulation of the population dynamics of the gastrointestinal nematodes in the tropical environment of Kenya.     

Development of free-living stages of equine strongyles in faeces on pasture in a tropical environment

The development of the free-living stages and yields of infective third stage strongyle larvae in faeces from a horse with a mixed natural infection deposited on pasture plots were studied over a 2-year period in a coastal area in tropical north Queensland. Two sets of faecal masses (one exposed to, and the other protected from the action of a natural population of dung beetles) were deposited monthly and after 7 days faecal samples were taken for larval recovery and counts. Hatching and development of the free-living stages occurred in faeces on pasture throughout the year. Development was rapid as infective stages were reached within a week of faecal deposition in all months. Yields of infective larvae were affected by the season and the action of dung beetles on the faecal masses. Highest yields were obtained from both beetle-exposed and protected faeces during winter (June to August) and lowest yields were in spring (September to November). High temperatures in spring and summer resulted in low yields of larvae, however, the dry conditions in spring made this season the most unfavourable period. In autumn and winter the temperatures were never low enough to stop or markedly slow down the rate of development, and allowed the development of large numbers of infective larvae. Dung beetle activity was observed throughout the year, and exposed faeces were usually completely dispersed within 24 h of deposition. This resulted in lower yields of infective larvae from these than from protected faeces. Though larval yields were lower, the actual numbers were still substantial so as to cast doubt on the usefulness of these beetles as biological control agents for equine strongylosis in the dry tropics.     

THE EPIDEMIOLOGY OF EQUINE STRONGYLOSIS IN SOUTHERN QUEENSLAND 1. The Bionomics of the Free-Living Stages in Faeces and on Pasture

SUMMARY Development of the free-living stages of strongylid nematodes of the horse to the infective stage occurred in faeces in all months of the year in southern Queensland, at a rate which depended on the season. Most rapid development to the infective stage occurred in the warmer months, with the hatching of strongyle eggs being completed in 2 days in summer. During the winter, egg hatching continued for over 2 weeks. Larval moults proceeded at a faster rate in summer—all larvae were infective in 7 days during the hottest months, but it was as long as 5 weeks before all were infective in winter. However, even though development was rapid in summer, survival rates varied from 1 to 10%, in contrast to the spring and autumn, when over 80% reached the infective stage. One percent of larvae in faeces survived for up to 20 weeks in autumn and winter, but for only 4 weeks in summer. These results highlight the inadequacy of short-term pasture spelling for all but the hottest months. Infective larvae were found on herbage in all months of the year, but greatest numbers were recovered in spring and early summer, and in autumn and early winter. The relationship of pasture infestation to migration of larvae from Paecal reservoirs in response to rain was clearly shown. Most infective larvae were found within 30 cm of faecal masses, and in fact 89% of all larvae isolated from herbage in this study were found within 15 cm of faeces. Migration of larvae from faeces to herbage occurred with falls of rain as small as 25 mm. Horse faecal masses dried out completely in 6–8 days in summer and in 14–16 days in winter. Strongyle larvae developed to the infective stage in faeces in the absence of rain, although many remained in the pre-infective stage and completed their development when rain fell. This study shows that massive contamination of pastures with the eggs of strongylid nematodes must be prevented in spring and autumn if susceptible young horses are not to be at serious risk.     

Studies on the epidemiology of bovine parasitic bronchitis.

In the West of Scotland the epidemiology of parasitic bronchitis in grazing calves was studied over a two year period with the aid of tracer calves and herbage examinations for Dictyocaulus viviparus larvae. The observations of both years emphasised the importance of overwintered lungworm larvae as a source of disease. In the first year it was shown that the ingestion and development of these overwintered larvae were, by themselves, directly responsible for severe morbidity, high faecal larval counts and deaths. In the second year it was shown that pasture ungrazed during the winter and spring and from which a hay crop was removed in mid-summer was still capable of producing clinical parasitic bronchitis in susceptible calves within three to four weeks of their introduction in later summer. In both years there was some evidence that the outbreaks appeared to be associated with the sudden availability of infective larvae on the herbage. The possibility that such larvae may have survived for many months in the soil is discussed. Despite the heavy challenge with lungworm larvae experienced by the grazing calves in the first year those vaccinated with lungworm vaccine survived, their clinical signs were mild and of short duration and their faecal larval output was greatly reduced.     

Nematode burdens of alpacas sharing grazing with sheep in New Zealand

Sheep and alpacas of similar age groups (6, 18 and 36+ months) were grazed for 16 weeks on pasture contaminated by lambs. Faecal egg counts, bulked larval cultures, lungworm larvae in faeces, dag scores, liveweight changes and nematode larvae on pasture were measured. Chabertia, Oesophagostomum, Cooperia, Ostertagia and Haemonchus and Trichostrongylus larvae were cultured from both the sheep and the alpacas. For the respective age groups, the alpacas had lower liveweight gains (10, 32 and 47 g/d vs 88, 84 and 120 g/d), peak faecal egg counts (384, 50 and 60 epg vs 1500, 500 and 140 epg) and faecal contamination of the perineum than the same ages of sheep. These results suggest alpacas became less affected with gastrointestinal nematodes than sheep.