Freezing of sheep faeces invalidates Haemonchus contortus faecal egg counts by the McMaster technique

Faecal pellets from a sheep that was artificially infected with a monoculture of Haemonchus contortus were collected over a 2-h period in the morning. In the laboratory the faeces were thoroughly mixed by hand and 48 by 1 g aliquots of the pellets were sealed in plastic bags, from which the air had gently been expressed. The faecal worm egg count of the sheep was about 14,000 g(-1). Varying numbers of the bags were either processed for faecal worm egg counting (FEC) by the McMaster technique on day 0, or were stored at one of the following temperatures: about 4 degrees C, -10 degrees C or -170 degrees C before processing. The faecal aliquots that were frozen were thawed at room temperature after having been frozen for either 2 h or 7 days, and processing of aliquots maintained at 4 degrees C proceeded shortly after the samples had been removed from the refrigerator. A dramatic reduction in egg numbers was found in all the aliquots that were frozen at -170 degrees C before faecal worm egg counts were done, as well as in those frozen for 7 days at about -10 degrees C. Numerous empty, or partially empty, egg shells were observed when performing the counts in faeces that had been frozen. In contrast, there was no significant reduction in the numbers of eggs in aliquots maintained for 7 days in a refrigerator at +/- 4 degrees C before examination, when compared with others examined shortly after collection of the faeces. Since H. contortus eggs in faeces are damaged by freezing, some methods that can be used for short term preservation are outlined. It is concluded that all nematode egg counts from cryopreserved faeces (whether in a freezer at -10 degrees C or in liquid nitrogen) should possibly be regarded as being inaccurate, unless the contrary can be demonstrated for different worm genera. However, exceptions are expected for the more rugged ova, such as those of the ascarids and Trichuris spp.     

Effect of temperature on development of the free-living stages of Ostertagia circumcincta

Hatching of the eggs of Ostertagia circumcincta was studied by recovering them from faeces and incubating them in distilled water at temperatures of 4, 16, 25 and 35 degrees C. Hatching occurred at all the temperatures. The rate of hatching increased with the rise in temperature. Development of larvae to the infective third stage (L3) was studied in faecal cultures incubated at 4, 16, 25, 30, 35, 40 and 45 degrees C. Except at 4 degrees C, L3 developed at all temperatures, the optimum temperature being 16 degrees C. The rate of development of L3 increased with the rise in temperature. This resulted in a corresponding decrease in the percentage recovery of larvae.     

Effect of pre-incubation storage conditions on hatchability,chick weight at hatch and hatching time in broiler breeders

1. Eggs were stored for two different times at varying temperatures. The effects on hatchability, chick weight at hatch and hatching time were examined in two broiler breeder lines from 33 to 58 weeks of age. 2. Short storage (1 to 3 d). Storage at 20 degrees C compared with 16.5 degrees C reduced hatchability of all eggs set. No effect was observed on hatchability of fertile eggs, hatching time or chick weight. 3. Long storage (9 to 11 d). Storage at 16.5 degrees C compared with 10 degrees C decreased both hatchability of fertile eggs and chick weight at hatch. Incidence of early embryonic death increased and incubation time decreased at 16.5 degrees C compared with 10 degrees C. 4. Chicks from morning eggs were heavier than those from afternoon eggs irrespective of storage conditions. 5. Hatchability (all eggs set and fertile eggs) and chick weight varied with hen age irrespective of storage conditions. During long storage, hatching time varied with hen age independently of breeder line, storage temperature or egg laying time. 6. Hatchability (all eggs set and fertile eggs) was higher in line A than in line B. Line B eggs hatched later and produced heavier chicks than line A eggs irrespective of storage time.     

Studies on ticks of veterinary importance in Nigeria. VI. Comparisons of oviposition and the hatching of eggs of Hyalomma species

Fully engorged Hyalomma spp. in Nigeria oviposited greater numbers of eggs than those partially engorged. Hyalomma impressum was a more prolific egg layer than H. rufipes, H. impeltatum and H. truncatum. The variations in the egg output as well as the recognizable peaks in the number of eggs during oviposition were described for each species. No species of Hyalomma below the engorged weight of 0.2 g oviposited; oviposition started with ticks of weight 0.3 g. Eggs produced by ticks weighing below 0.3 g did not hatch; the highest percent egg eclosion occurred with ticks of weight 0.6 g (H. rufipes) and 0.7 g (other Hyalomma species). The pre-oviposition, oviposition and eclosion periods were shortened when eggs were laid and incubated at high temperatures, although the number of oviposited eggs did not increase significantly. At the standard temperature of 24 degrees C, the longest eclosion period was seen in the eggs of H. rufipes (41 days) while those of H. truncatum, H. impressum and H. impeltatum were similar to each other (29 days). Only eggs of H. rufipes hatched at an incubation temperature of 15 degrees C. Eggs of Hyalomma species laid at the same time hatched over a 2--4 day period, except at 15 degrees C when the hatching period of H. rufipes lasted 10 days. The eclosion period was longest in the earlier ovipositions and shorter in the later ones. It is suggested that some intrauterine larval development might have started in the eggs before they were released at a later oviposition period. The percentage mortality of eggs at various temperatures showed that eggs of H. rufipes were more tolerant of low temperatures than those of H. impressum, H. truncatum and H. impeltatum, while the eggs of the latter 3 species were more tolerant of high temperatures than those of H. rufipes. The relevance of these results of the distribution and abundance of the Hyalomma species in Nigeria was discussed.     

Effect of temperature on the growth of Penicillium from moldy feed

178 Penicillium strains were isolated from moulded feedstuffs (mainly corn and wheat after ambient air drying, wheat after refrigeration, corn after treatment with propionic acid), and investigated for the effect of temperature on growth rate. Temperatures between 4 and 27 degrees C were used. 159 strains proved psychrotrophic, they were able to grow at 4 degrees C on malt extract agar (MA) as well as in wheat; the growth in wheat was indicated by the production of ergosterol. On MA all psychrotrophic strains were able to grow also at 10-27 degrees C. The maximum rate of growth was reached at 10-15 degrees C, 15-20 degrees C, 20-27 degrees C, or must be assumed for temperatures greater than or equal to 27 degrees C. At 4 and 10 degrees C the mean growth rate on MA was 24 and 51%, respectively, related to the mean rate at 20 degrees C; the lower temperature limit of growth was found at - 1 degree C by graphic extrapolation. The Q10 value of growth on MA is about 2, if temperatures between 20 and 10 degrees C are compared; with decreasing temperature higher Q10 values up to 3.7 are obtained. The growth rate of a strain of Penicillium aurantiogriseum on wheat was affected by temperature nearly in the same manner as on malt extract agar. The results are discussed with respect to the risk of moulding during refrigerated storage of feedstuffs.     

Effect of cool storage of faecal samples containing Haemonchus contortus eggs on the results of an in vitro egg development assay to test anthelmintic resistance

An in vitro egg development assay was used to test the resistance of eggs from a benzimidazole susceptible and a resistant strain of Haemonchus contortus to four benzimidazoles. Storage of eggs contained in faecal samples at 4 degrees C for four, 24 or 72 hours did not appear to impair the results from the in vitro egg development assay or alter the LD50. Storage for one week, however, proved to be lethal for the eggs. The average LD50 values for thiabendazole, albendazole, mebendazole and cambendazole were 0.06, 0.07, 0.96 and 2.47 ppm for the susceptible strain. For the resistant strain the values averaged 0.26, 0.24, 1.07 and 5.11 ppm.     

Development of Hyalomma lusitanicum under laboratory conditions

At the constant temperature of 25 degrees C and relative humidity (RH) of 84%, the average pre-oviposition period of Hyalomma lusitanicum was 47 days, the oviposition lasted an average of 26 days and the total egg production was an average of 6320 per female. At 16 degrees C the females did not lay eggs at all, but those which survived for 1 year and were transferred thereafter to 25 degrees C and 84% RH laid viable eggs. At 35 degrees C, the oviposition was identical at all levels of RH tested (25, 62 and 93%). At 25 degrees C, the pre-oviposition period was shorter at 93% RH, and the number of eggs laid was fewer at 25% RH. The eggs hatched in 32-40 days, the hatching percentage being lower in batches of eggs laid at the beginning and at the end of the oviposition period. The larval and nymphal moultings were not influenced by the type of host. As the temperature increased, the pre-moult period became shorter. The engorged larvae were more sensitive to the low RH than the engorged nymphs, whose moulting percentage was always greater than 72 in all regimes. Low temperature and high humidity had a favourable effect on the survival of unfed nymphs. The female-to-male ratio was 1:2. Hyalomma lusitanicum always behaved as a 3-host tick. The adults did not engorge on rabbits. The female ticks engorged on calves weighed an average 543 mg. Ticks maintained at 25 degrees C and 84% RH and engorged on calves completed the life cycle in 138-196 days, which does not include the period of chitinization of about 30 days. More than half of this period was spent in egg laying and hatching.     

Effect of short periods of high incubation temperature on hatchability and incidence of embryo pathology of turkey eggs

1. Turkey eggs were incubated at 38.0 degrees and 38.5 degrees C at different stages of embryo development and for periods of 3 to 25 d. Results were compared with control eggs incubated at 37.5 degrees C. The age of mortality, the incidence of malpositions and the incidence of morphological abnormalities were recorded from all unhatched eggs. 2. Eggs incubated at 38.5 degrees C for 5 or more days hatched significantly less well than eggs incubated at 37.5 degrees C. Eggs incubated at 38.5 degrees C for 3 d hatched worse than controls but not significantly so. Eggs incubated between 0 and 25 d and 7 and 12 d but not between 0 and 6, 13 and 18 and 19 and 25 d, at 38.0 degrees C had significantly lower hatchabilities than eggs incubated at 37.5 degrees C. 3. Lowest hatchabilities were obtained when the eggs were incubated at high temperatures between 7 to 12 d and 6 to 10 d, indicating that embryos at this stage of development are more likely to succumb to high temperature than at other ages. 4. Increases in embryo mortality due to overheating were seen in weeks 3 and 4 of incubation and at the pipping stage. This was observed even when the period of overheating occurred in weeks 1 and 2 of incubation. 5. Embryos malpositioned with their head in the small end of the egg were seen at a higher incidence when overheating in the 2nd or 3rd quarter of the incubation period.     

Calibration and diagnostic accuracy of simple flotation, McMaster and FLOTAC for parasite egg counts in sheep

The present study was aimed at carrying out a calibration and a comparison of diagnostic accuracy of three faecal egg counts (FEC) techniques, simple flotation, McMaster and FLOTAC, in order to find the best flotation solution (FS) for Dicrocoelium dendriticum, Moniezia expansa and gastrointestinal (GI) strongyle eggs, and to evaluate the influence of faecal preservation methods combined with FS on egg counts. Simple flotation failed to give satisfactory results with any samples. Overall, FLOTAC resulted in similar or higher eggs per gram of faeces (EPG) and lower coefficient of variation (CV) than McMaster. The "gold standard" for D. dendriticum was obtained with FLOTAC when using FS7 (EPG=219, CV=3.9%) and FS8 (EPG=226, CV=5.2%) on fresh faeces. The "gold standard" for M. expansa was obtained with FLOTAC, using FS3 (EPG=122, CV=4.1%) on fresh faeces. The "gold standard" for GI strongyles was obtained with FLOTAC when using FS5 (EPG=320, CV=4%) and FS2 (EPG=298, CV=5%). As regard to faecal preservation methods, formalin 5% and 10% or freezing showed performance similar to fresh faeces for eggs of D. dendriticum and M. expansa. However, these methods of preservation were not as successful with GI strongyle eggs. Vacuum packing with storage at +4°C permitted storage of GI strongyle eggs for up to 21 days prior to counting. Where accurate egg counts are required in ovine samples the optimum method of counting is the use of FLOTAC. In addition, we suggest the use of two solutions that are easy and cheap to purchase and prepare, saturated sodium chloride (FS2) for nematoda and cestoda eggs and saturated zinc sulphate (FS7) for trematoda eggs and nematoda larvae.     

Replication of reptilian paramyxovirus in avian host systems

The reptilian paramyxovirus GOV replicated in chicken embryo fibroblasts, in embryonated chicken eggs and in explanted chorio-allantoic membrane with titres of up to 10(8.2) TCID50/ml at 28 degrees C. The virus did not multiply above 30 degrees C. GOV re-isolated from the avian host systems was identified by immunofluorescence and by immunogold-electron microscopy.     

Effects of short-term exposure to condensed tannins on adult Trichostrongylus colubriformis

Twelve parasite-naive sheep were used to study the possible direct anthelmintic effect of a condensed tannin extract (quebracho) on the population and fecundity of the intestinal nematode Trichostrongylus colubriformis. The sheep were infected with a single dose of 20,000 L3 of T colubriformis. Twenty-eight days later, six of them were drenched daily for a week with quebracho extract at 8 per cent by weight of their food intake. All lambs were then slaughtered, and their small intestines removed to estimate the worm burdens and the numbers of eggs in utero. Two days after the first drench with tannin extract the faecal egg counts of the treated sheep were approximately 50 per cent of those of the control sheep (P<0.01), but there was no further reduction with continued drenching. In the treated sheep the worm burdens and number of eggs per gram faeces per worm were reduced by 30 per cent compared with the controls (P<0.05), but the sex ratios, the number of eggs in utero and length of the worms were not affected by drenching with tannin.     

Effect of time and temperature on growth of Salmonella enteritidis in experimentally inoculated eggs

Influence of time and temperature on Salmonella enteritidis multiplication in experimentally injected eggs was examined. There was an increase in the number of S. enteritidis with the increase in temperature of egg storage. There was less increase of S. enteritidis in eggs stored at 4 degrees C than in eggs held at temperatures higher than 4 degrees C (P less than 0.05). These results suggest a possible method for monitoring commercial eggs for the presence of S. enteritidis. It was concluded that the chances of recovery of S. enteritidis can be increased 10(6)-fold or more by holding the eggs at temperatures of 21 or 27 degrees C for more than 20 days and culturing their contents.     

Development and survival of free-living stages of equine strongyles under laboratory conditions

In a series of laboratory studies the optimum conditions for the development and survival of the free-living stages of strongyle parasites occurring in horses in tropical north Queensland were determined. No differences in behaviour were noted between the strongyle species. Development to the infective stage occurred only between 10 and 35 degrees C. The rate was affected by temperature, taking 15-24 days and 3 days, respectively, at the lowest and highest temperatures for the developing stages to reach the infective third stage. Yields of infective larvae were very low outside the range 20-33 degrees C, and were highest at 28 degrees C. Survival of infective larvae was good between 20 and 33 degrees C, and large numbers were recovered after 3 months in faeces incubated at 20-28 degrees C. At 33 and 37 degrees C larval survival was affected by the moisture content of the faeces, with infective larvae surviving better in dry than in moist faeces; even a residual moisture level of 40% significantly reduced the number of larvae recovered from faeces incubated at 37 degrees C for 1 month. Moisture also affected larval development, especially at the higher temperatures of 25-39 degrees C. When faecal moisture content fell to less than or equal to 20% by 3 days, larvae which had not yet reached the infective stage were still pre-infective at 7 days, while all larvae in faeces with adequate moisture had reached the infective third stage. It was not possible to determine the critical faecal moisture level below which larval development ceased, however, 28 degrees C (range 25-33 degrees C) was found to be the optimum temperature. Larval development was very rapid and yields of infective larvae highest at this temperature.     

MILK PRODUCTION OF GRAZING DAIRY CATTLE AFTER A SINGLE ANTHELMINTIC TREATMENT

The prevalence of nematode infections in lactating dairy cattle was determined by faecal egg counts and cultures from over 350 cows selected at random from twelve herds. The mean egg count was 3 eggs per gram (epg) and 43% of the cows sampled were passing detectable concentrations of eggs (greater than 6 eggs) in their faeces. In June 1976 171 cows in 10 herds were treated with 7.5 mg/kg fenbendazole and their milk production compared over the following 2 months with that of 164 untreated cows in the same herds. Anthelmintic treatment had no significant influence on milk production in any of the herds. The contrasting results of this study and recent studies in the United States of America are discussed.     

Seasonal comparisons of anthelmintic activity of levamisole pour-on in cattle in the USA

Anthelmintic activity of a pour-on formulation of levamisole, applied during warm weather (16 degrees to 36 degrees C) at 10 mg/kg bodyweight, was evaluated in groups of naturally parasitised calves. This activity was compared to that obtained in similar groups of calves treated in the winter (-4 degrees to +7 degrees C). Controlled efficacy of the pour-on formulation was determined for each season by comparing mean worm burdens in treated calves sacrificed seven to nine days after treatment to non-treated controls. In these trials, burdens of Bunostomum phlebotomum, Cooperia species, Haemonchus placei, Nematodirus species, Oesophagostomum radiatum, Ostertagia ostertagi and Trichostrongylus axei in treated calves were reduced by 83.3 to 100 per cent in the summer and 89.2 to 100 per cent in the winter. Field investigations at nine locations across the USA compared changes in faecal egg counts for cattle treated and evaluated during warm summer months (27 degrees to 36 degrees C) to those treated during cold winter months (-18 degrees to +10 degrees C). Overall, faecal egg counts were reduced by 90.2 per cent in the summer trials and 94.0 per cent in the winter trials. The results of these trials indicate that there is no seasonal variation in the anthelmintic activity of this pour-on formulation of levamisole.     

Optimization of a culture technique for the isolation of Listeria monocytogenes from faecal samples

A culture technique employing cold enrichment at 4 degrees C followed by selective enrichment and plating at higher temperatures (30 degrees C) was used to isolate Listeria monocytogenes from faecal samples. The samples were held at 4 degrees C for 15 weeks and cultured weekly to assess the sensitivity of the culture after cold storage for different lengths of time. No media, Listeria selective enrichment broth (LSEB), nutrient broth (NB) and saline were used as cold storage medium. Cold storage increased the frequency of Listeria positive samples. The sensitivity of the culture for Listeria spp. and L. monocytogenes was 72 and 94%, and 56 and 61% after third and seventh week of cold storage, respectively. When the results of third and seventh week of cold storage were combined, the sensitivity was 100% for Listeria spp. and 94% for L. monocytogenes. LSEB and NB as storage medium increased Listeria positive samples after the first week of cold storage but did not maintain the increase thereafter while saline had an adverse effect on the growth of the bacteria. However, samples held in no media in a pilot study involving monthly sampling of a herd revealed better results. Detection limit of the culture media was also investigated. The lowest concentration detected by culture media was 3.17 organisms/ml. This was seven organisms/g for known Listeria positive sample. The faecal samples spiked with 10-fold dilutions of L. monocytogenes and held at 4 degrees C revealed that the sample spiked with 3.17 x 10-1 cfu/ml organisms resulted in growth after the second week of cold storage. The results suggest that the culture technique employing cold enrichment followed by selective enrichment and plating is more sensitive, the storage of faecal samples in no media when compared with the samples in storage medium, LSEB, NB and saline, during cold enrichment is a better application and culture of faeces, immediately after collection, at third and seventh week of cold enrichment produce more satisfactory results.     

Some effects of temperature on the adults, eggs and pupae of Stomoxys calcitrans Linnaeus (Diptera: Muscidae)

Adults could only live and reproduce to their full capacity at temperatures between 20 degrees C and 30 degrees C. At 15 degrees C the females laid no eggs, the adult life span was relatively short and the reproductive capacity of females kept at 35 degrees C was low. The thermal histories of the flies had no apparent effect on their later reactions to temperature in any of the parameters tested. The viability rates of S. calcitrans eggs exposed to temperatures between 10 degrees C and 40 degrees C exceeded 84%, but 45 degrees C was lethal. The optimum temperatures for incubation of the eggs was 30 degrees C. Pupae of S. calcitrans seemed to tolerate temperatures between 20 degrees C and 30 degrees C, but their mortalities increased markedly outside this temperature range. Tests showed that pupal mortalities increased linearly with increasing periods of exposure to a temperature of 15 degrees C.     

Improving turkey poult quality by correcting incubator humidity to match eggshell conductance

One of the most important factors determining hatchability of avian eggs is their proper water budget during incubation. We show here that water budget changes of turkey eggs effect not only hatchability but also poult quality. In addition to an increase in hatchability of 3.3%, poult quality was increased by 7.3%. This was achieved by sorting eggs into low (less than 18.5), medium (18.5 to 22.0) and high (greater than 22.0) eggshell mass-specific water vapour, conductance categories and incubating them in matching incubation humidities of 21.8, 26.6 and 31.4 Torr, corresponding to relative humidities of 45, 55 and 65% at 37.5 degrees C, respectively. A total diffusive water loss of 11.5% (range 10 to 14%) of the initial egg mass in 25 d of incubation yielded maximal hatchability and poult quality.     

The effect of temperature and relative humidity on survival of eggs of the cestode, Monieza expansa (Rudolphi, 1810)]

The survival of Moniezia expansa eggs in the droppings of lambs was investigated at various temperatures in laboratory conditions and on test plots outdoors. The optimum temperature of the livability of M. expansa eggs in laboratory conditions is 5 degrees C; at this temperature 10% of oncospheres survived after 161 days. At the temperatures of 10, 25, 35 degrees C the oncospheres survived 105, 28, 46 days respectively, at -12 degrees C it was 28 days. It was for 21 and 35 days that on the test grassy plots the eggs of M. expansa survived in the droppings of lambs in the summer months of July and August at the average air temperatures of 15.7-18.2 degrees C and relative humidity of 67.7-74.3%. In autumn in September and October, at the average temperatures of 5.8-14.6 degrees C and relative humidity of 65.3-76.7% the oncospheres survived for 49 to 91 days. The M. expansa eggs in the droppings of lambs were able to survive on the test plot. The living oncospheres were demonstrated for 119 days from November 1987 to March 1988, and for 175 days till May by means of experimental infection of intermediate hosts.     

Anthelmintic resistance and sheep management practices in south western Victoria

Twenty-eight farms in 7 shires in south western Victoria were selected and tested for presence of benzimidazole-resistant nematodes between November 1979 and June 1981. Mean faecal egg counts of sheep were less than 100 strongyloid eggs/g on 11 farms. Faecal egg count reduction tests were conducted on the remaining 17 farms and thiabendazole was less than 90% efficient in reducing egg counts in sheep from 5 (29%) of these farms. Thiabendazole-resistant Teladorsagia circumcincta were identified at necropsy of experimentally infected treated sheep. In further studies a survey of 104 farms was conducted in the Mount Rouse and Dundas shires of western Victoria in 1981 and 1982 respectively to determine the prevalence of anthelmintic resistance in these shires. Mean faecal egg counts among weaner sheep in the winter-spring of both years were less than 100 eggs/g which indicated low levels of parasitic nematode populations. A faecal egg count reduction test was conducted on 10 farms and thiabendazole was less than 90% efficient on 3; levamisole was greater than 90% efficient in all 10 tests. Most of the surveyed farms carried Merino or Merino crossbred sheep at 10 to 15 dry sheep equivalents per ha and weaners were treated with anthelmintics 3 to 6 times per year. Management procedures based mainly on anthelmintic therapy were effective in controlling nematode populations in weaner sheep, although many producers alternated between different groups of anthelmintics within the same year contrary to current recommendations for long-term preservation of anthelmintic efficacy. It was concluded that anthelmintic resistance was not of practical importance to the majority of sheep producers in the region.