Key determinants affecting sheep wool biodegradation directed by a keratinase-producing <Emphasis Type="Italic">Bacillus subtilis</Emphasis> recombinant strain

OVAT (one variable at a time) approach was applied in this study to screen the most important physicochemical key determinants involved in the process of sheep wool biodegradation. The process was directed by a keratinase-producing Bacillus subtilis DB 100 (p5.2) recombinant strain. Data indicate that, sheep wool could be degraded efficiently in cultures incubated at 30°C, with initial pH of 7 with agitation at 150 rpm. Two times autoclaved alkali treated and undefatted chopped sheep wool is more accessible to biodegradation. B. subtilis recombinant cells could utilize sheep wool as a sole source of carbon and nitrogen. Sheep wool-based modified basal medium II, lacking NH₄Cl and yeast extract, could greatly support the growth of these bacterial cells. Sheep wool biodegradation was conducted efficiently in the absence of kanamycin consequently; high stability of the recombinant plasmid (p5.2) represents a great challenge upon scaling up this process. Three key determinants (sheep wool concentration, incubation time and inoculum size) imposing considerable constraints on the process are highlighted. Sheep wool-based tap water medium and sheep wool-based distilled water medium were formulated in this study. High levels of released end products, produced from sheep wool biodegradation are achieved upon using these two sheep wool-based water media. Data indicate that, sheep wool hydrolysate is rich in some amino acids, such as tyrosine, phenylalanine, lysine, proline, isoleucine, leucine, valine, aspartic acid and glutamic acid. Moreover, the resulting sheep wool hydrolysate contains soluble proteins of high and intermediate molecular weights. The present study demonstrates a feasible, cheap, reproducible, efficient and rapid biotechnological approach towards utilization of raw sheep wool waste through a recombinant bacterium.     

Genetic control of alopecia in sheep

A hypothesis is suggested which turns down the opinion that alopecia or pathological wool shedding in sheep is due to some pathological processes or disease. We suppose that this trait is controlled by a gene of natural wool shedding inherited from sheep far ancestors, and we propose to designate the recessive alopecia gene as Alp3 and its dominant allele, a wool alopecia resistance gene, as AlpA. Alopecia will only be revealed in homozygotic Alpa/Alpa state, due to recessivity of alopecia gene, but heterozygotic AlpA/Alpa and homozygotic AlpA/AlpA sheep will have a normal wool coat. A new polymorphic trait is supposed to exist which is conditioned by simultaneous presence of two genetic alleles in a sheep population with descrete phenotypic manifestation.     

Influence of hydrothermal factors on wool development of Tan sheep in China

Hydrothermal factors in Ningxia, an arid and semi-arid area in Northwestern China, were examined to determine their influence on the geographical differentiation of the wool quality of Tan sheep lambskin. Prospective areas for extension of the Tan sheep industry were also investigated. Two sheep groups were sampled; one consisted of artificial insemination flocks (1241 lambs sampled from 25 flocks) and the other of natural mating flocks (1009 lambs from 38 flocks). Six phenotypical traits were measured for each lamb and six hydrothermal factors were collected from the meteorological stations located in the sampling areas. A significant correlation was found between wool characteristics of the lambskin and hydrothermal conditions. Data further indicated that among all the hydrothermal factors measured, those causing the geographical differentiation of soil and vegetation were the major factors responsible for the corresponding differentiation of the lambskin and wool quality of Tan sheep. Thermal factors were mostly positively correlated with wool quality, while the influence of moisture was negative. Three eco-geographical regions were defined based on a combination of the hydrothermal conditions and corresponding wool characters: (i) typical region or super-suitable region, (ii) sub-typical region or suitable region and (iii) transitional region. The delimitation could be used as a basis for the extension of the Tan sheep industry. It is also suggested that the wool quality of lambskin of Tan sheep in these three regions could be improved by means of controlled breeding and selection.     

The dietary-regulated biosynthesis of high-sulphur wool proteins

1. When the diet of sheep is supplemented by the abomasal infusion of sulphur-containing amino acids or casein, a special group of proteins, with a very high content of sulphur (about 8.3%), is incorporated into the high-sulphur proteins of wool. These special proteins cannot be detected in control wool from the same sheep. 2. This is a naturally occurring process, as these special proteins are found in wool from sheep on a high level of nutrition under ordinary conditions of feeding, and in wool of an inherently high sulphur content. 3. This represents a control mechanism in protein synthesis that has not previously been observed, and may be further evidence that the high-sulphur proteins of wool are produced by an unusual synthetic route.     

Effects of phenylalanine and analogues of methionine and phenylalanine on the composition of wool and mouse hair

Administration of the methionine analogue methoxinine (O-methyl-DL-homoserine) to sheep substantially changed the composition of wool; in addition wool fibres were weakened and the staple crimp frequency was reduced for a prolonged period. The proportions of high-tyrosine proteins were reduced by 40-45% whereas the high-sulfur proteins were usually slightly increased. The content of high-tyrosine proteins in wool was still depressed in most sheep 70 days after dosing with methoxinine. These experiments supported a previous finding that the cystine content of wool and its crimp frequency are not causally related. Ethionine, another methionine analogue, did not consistently change the composition of wool. In some sheep there was no change in the proportions of high-tyrosine proteins following administration of ethionine, even though weak wool was produced. This result, together with the lack of association between the content of high-tyrosine proteins and the strength of wool fibres in a sheep given methoxinine plus methionine, indicates that a reduction of the high-tyrosine proteins is not a prerequisite for the production of weak wool. Neither a threefold increase in the phenylalanine intake by mice nor the administration of three analogues of phenylalanine (4-fluoro-DL-phenylalanine, 4-chloro-DL-phenylalanine and beta-(2-thienyl)-DL-alanine) to sheep altered the composition of hair or wool. Fluorophenylalanine was incorporated into all the constituent proteins of wool to the extent of c. 2% of phenylalanine residues. The other analogues studied could not be detected in wool.     

Wool proteins of New Zealand Romney sheep

Proteins extracted from the wool of 65 Romney ewes were analysed qualitatively by one- and two-dimensional polyacrylamide gel electrophoresis. Romney wool proteins could be classified into the low-sulfur, high-sulfur, and high-tyrosine protein groups described for wool from other breeds. The wool protein pattern of an individual sheep remained constant despite changes in season, age or nutritional status of the sheep, and did not vary between different body positions. There were between-sheep differences in protein pattern, most variation occurring in the high-sulfur protein group. These differences were presumed to reflect genetic differences between the sheep.     

THE OLFACTORY BEHAVIOUR OF LUCILIA SPECIES (DIPTERA) UNDER NATURAL CONDITIONS

Using an apparatus of the 'two-choice' type under field conditions, it has been demonstrated that a wool-factor forms an important part of the mechanism whereby female Lucilia sericata may be attracted to sheep. The 'wool-factor' could not wholly be replaced by ammonium- or sulphydryl-type attractants.
L. sericata was strongly attracted by the combination of ammonium carbonate/indole with sheep wool but less strongly attracted when ethyl mercaptan/hydrogen sulphide was used with sheep wool. L. caesar and L. illustris , on the other hand, were not markedly responsive to the ammonium-type attractants but were strongly attracted by the sulphydryl mixture used either alone or in combination with sheep wool.
Temperature is an important element in the oviposition stimulus. The amount of oviposition by L. sericata was increased when substances known to induce oviposition were exposed at 30–40† C.
Sheep wool kept moist and incubated at approx. 38† C. under field conditions for 3 weeks did not attract L. sericata and oviposition did not occur on such wool. This suggests that bacteria normally present on the wool fibre as distinct from the skin, do not play an important part in rendering sheep susceptible to blowfly attack.     

Rapid morphological change in an insular population of feral sheep

Artificially selected qualities can reduce fitness in a wild setting, thus feral domesticates should experience strong selective forces. Domestic sheep Ovis aries have frequently become feral on islands, which differ substantially from mainland environments. We examined changes in body mass and wool traits in feral sheep inhabiting Santa Cruz Island (SCI), California for ≥90 years. To elucidate the influence of nutrition, we compared the mass of feral island sheep with that of island sheep raised in farm conditions. We found that feral sheep on SCI were smaller than purported founder breeds, and that most documented populations of insular feral sheep worldwide have converged to similar body sizes (within 6 kg). SCI rams attained greater mass in farm conditions but ewes did not, suggesting phenotypic plasticity in ram body mass. Ewes exhibited self-shedding of wool at a greater frequency than rams, and sex differences and shedding patterns were consistent with thermoregulation and the risk of fly strike disease as benefits of wool loss. Pigmentation rates did not increase, further supporting the influence of heat stress on wool traits. These changes occurred in <25 generations and may have had a genetic basis, representing a potential example of rapid evolution in insular feral sheep.     

How to predict attachment potential of seeds to sheep and cattle coat from simple morphological seed traits

Dispersal is a process that determines many aspects in the life-history of plants. Up to now, however, it is difficult to quantify. Many studies rather assess it as a categorical trait, i.e. assuming a species is dispersed by a certain vector or not. Gradual differences in the dispersal potential between species are rarely considered.
In this paper we focus on the key process of epizoochory: the attachment potential of species to animal coats. We present two simple models (GLMs) how to quantify and to predict attachment potentials to sheep wool and to cattle hair from easily measurable seed traits. To calibrate the models, we investigated the attachment potential of seeds of 130 plant species on mechanically shaken coats of sheep and cattle and measured traits describing the seeds. We found that seed mass and seed morphology were correlated with attachment potentials. For sheep wool, the combination of the logarithm of seed mass and a constant for the different seed morphology types explained 85% of the variation of attachment potentials. For cattle hair, 71% were explained. To validate the models, they were applied to 36 additional species. Predicted and experimentally measured attachment potentials were correlated with r=0.84 for sheep wool and r=0.61 for cattle wool. Thus, the attachment potential of seeds to sheep or cattle coat can be assessed for a large set of species using the described models if seed mass and seed morphology are known.     

Investigation of some amino acid analogues and metabolites as inhibitors of wool and hair growth

Sheep were given intravenous infusions of ethionine together with cycloleucine or reduced glutathione, in attempts to prevent the inhibition of wool growth by ethionine. Other sheep were given cycloleucine alone to measure effects on wool growth. Twenty-two compounds related to cystine, methionine, ethionine, lysine, phenylalanine and tyrosine were given as intravenous infusions to sheep to investigate their potential as depilatory agents. Nineteen of these compounds were also tested in mice during their first cycle of hair growth. The concurrent administration of cycloleucine with ethionine prevented the weakening of wool fibres caused by ethionine, but reduced glutathione was ineffective. Cycloleucine weakened wool fibres, as judged subjectively, and caused a small reduction in fibre diameter. Selenocystine and selenomethionine caused some hair loss in mice but selenocystine was also toxic. Both seleno-amino acids were toxic for sheep; selenocystine was lethal at 0.025 mmol kg-0.75 and selenomethionine at 0.09 mmol kg-0.75. Doses that permitted survival of sheep did not have depilatory effects. However, the presence of autophagic vacuoles in the cytoplasm of follicle bulb cells of sheep indicated that a toxic dose of selenocystine had potential depilatory activity. Other compounds investigated did not induce loss of wool or hair. Some compounds, notably 3-methylthiopropionic acid and S-(2-aminoethyl)-L-cysteine, were toxic to mice but not sheep. The methionine analogue, methoxinine (O-methyl-DL-homoserine), caused a substantial reduction in the strength of wool fibres and a prolonged alteration of the crimp pattern. It is suggested tentatively that cycloleucine inhibits methionine adenosyltransferase and thereby reduces or prevents the formation of S-adenosylethionine. The failure of various compounds related to methionine and ethionine to have any depilatory activity in sheep supports the view that ethionine influences wool growth via the formation of S-adenosylethionine.     

Modified techniques for the isolation of Dermatophilus spp. from infected material.

Two improved techniques are described for the isolation of Dermatophilus spp. The first is suitable for samples of scab from lesions of the disease in animals, except for sheep affected with lumpy wool. The second is for the isolation of the organism from samples of lumpy wool in sheep. Both techniques are simple, rapid and reliable and are based on releasing the organism by pulverizing the scab and then by suppressing contaminants by including polymixin B sulphate in the medium. Excellent results are obtained with or without pre-incubation exposure to CO2.     

Responses of hair and wool sheep to a single fixed dose of infective larvae of Haemonchus contortus

Body weights, fecal egg counts (FEC), and packed cell volumes (PCV) of hair sheep and wool sheep crossbred lambs were compared over 8 weeks following administration of a single dose of approximately 10,000 third-stage larvae of Haemonchus contortus. Hair sheep lambs (n=17) were reciprocal crosses between mainland USA populations of Barbados Blackbelly (BB) and Virgin Islands White (VIW) sheep. Wool sheep lambs (n=64) were from a crossbred composite of 50% Dorset, 25% Rambouillet, and 25% Finnish Landrace breeding. Lambs of both breed types continued to grow during the period of infection. Mean weights were higher for wool lambs (39.7±0.8 kg) than for hair lambs (28.2±1.5 kg). FEC increased to week 5 in both breed groups and remained elevated in wool lambs through week 7 but declined sharply in hair lambs after week 5. Mean FEC for weeks 4 through 8 were 4011±361 eggs per gram of feces (epg) in wool lambs, and 1135±196 epg in hair lambs. PCV declined through week 7 in wool lambs but stabilized and then increased after week 4 in hair lambs. Mean PCV in weeks 4 through 8 were 22.4±0.3% in wool lambs and 24.3±0.5% in hair lambs. These results suggest that Caribbean hair breeds may be able to contribute significantly to development of parasite-resistant sheep populations.     

Volatiles from Merino fleece evoke antennal and behavioural responses in the Australian sheep blow fly Lucilia cuprina

To identify flystrike‐related volatile compounds in wool from Merino sheep, the attractiveness of wool to Lucilia cuprina Wiedmann (Diptera: Calliphoridae) was examined. First, a selection of wool samples guided by previous knowledge of sheep lines, predicted to be more susceptible or more resistant to flystrike, was tested. The attractiveness of the 10 samples selected was not associated with field susceptibility: two samples from the more resistant line were identified as most attractive and two samples from the more susceptible line were identified as least attractive, based on the behavioural assays with gravid flies. Comparison of the headspace volatiles of these samples, using solid phase microextraction and gas chromatography‐mass spectrometry‐electroantennographic detection, revealed octanal and nonanal to be present in the attractive wool samples that elicited responses from the fly antenna. Furthermore, the two compounds were not present in wool that was least attractive to L. cuprina. In laboratory bioassays, octanal and nonanal evoked antennal and behavioural responses in gravid L. cuprina, thus confirming their potential role as semiochemicals responsible for attracting L. cuprina to Merino sheep.     

Identification of microRNAs in Wool Follicles during Anagen,Catagen, and Telogen Phases in Tibetan Sheep

Background

Wool quality is one of the most important economic traits in sheep. The wool fiber is derived from specialized skin cells that are referred to as wool follicles. To understand the roles of microRNAs (miRNAs) in wool fiber growth, we detected the expression patterns of miRNAs in wool follicles at the anagen, catagen, and telogen stages from Tibetan sheep through Solexa sequencing.

Results

A total of 244 mature miRNAs were identified. Of these, only five miRNAs are listed in the database of sheep miRNAs (miRBase Database V19), and the other 239 miRNAs have not been previously described in this species. Further analyses indicated that 204 miRNAs are evolutionarily conserved among mammal species, whereas 35 of the identified miRNAs were first found specifically in sheep. The expression pattern analyses showed that the expression levels of 39, 34, and 20 of the miRNAs significantly change between anagen and catagen, between anagen and telogen, and between catagen and telogen, respectively. The results of the bioinformatics analysis show that these differentially expressed miRNAs might regulate wool follicle development by targeting genes in many different pathways, such as the MAPK and Wnt pathways, as well as the pathways that regulate the actin cytoskeleton, focal adhesion, and tight junctions. Furthermore, we identified six differentially expressed miRNAs (oar-miR-103-3P, oar-miR-148b-3P, oar-miR-320-3P, oar-miR-31-5P, oar-novel-1-5P, and oar-novel-2-3P) that might target the key genes of the Wnt pathway. It has been reported that the Wnt pathway is critical for wool follicle development. Therefore, these miRNAs may regulate wool development through the Wnt pathway.

Conclusions

Our results provide new information on the identification and expression pattern of miRNAs in wool follicles. Our data might therefore aid in the understanding of the mechanisms of wool follicle development in sheep.     

绵羊皮肤中GHR、IGF-1和IGF-1R基因表达的发育性变化及品种特点

采用相对定量反转录多聚酶链式反应 (RT-PCR)方法, 以18S rRNA作内标, 研究了罗米丽(Romilly Hillys)×中国美利奴(新疆军垦型)杂交一代优质细毛羊和哈萨克粗毛羊皮肤中生长激素受体(GHR)、胰岛素样生长因子1(IGF-1)和胰岛素样生长因子1受体(IGF-1R) mRNA发育性变化并进行了品种间比较。分别于30、60、90、135、180和255日龄称重、采毛样, 并于30、90、135和255日龄采皮样。结果表明: 粗毛羊和细毛羊体重、羊毛生长的发育模式没有明显的差异, 30~135日龄体重迅速增加, 135~255日龄增重十分缓慢; 30~135日龄羊毛日增长逐渐增加, 135~180日龄羊毛生长十分缓慢, 而180~255日龄又上升到较高水平。粗毛羊皮肤中GHR mRNA在30~90日龄显著增加 (P<0.05), 90日龄达到高峰, 此后显著下降(P<0.05); 细毛羊在135日龄时GHR mRNA极显著地升高(P<0.01), 此后又极显著地下降。粗毛羊皮肤中IGF-1、IGF-1R mRNA 30~90日龄上升, 90日龄之后极显著下降(P<0.01); 细毛羊皮肤中IGF-1、IGF-1R mRNA出生时较高, 然后逐渐下降。品种之间比较, 细毛羊GHR mRNA出现高峰晚于粗毛羊, 135日龄高峰时显著地高于粗毛羊; 粗毛羊IGF-1、IGF-1R mRNA在90日龄出现高峰, 并极显著或显著地高于细毛羊; 粗毛羊90日龄前GHR、IGF-1和IGF-1R mRNA高于细毛羊, 之后低于细毛羊。结果提示: 绵羊皮肤中GHR、IGF-1和IGF-1R基因表达有特定的发育模式, 并存在品种差异。     

Effects of intravenous infusion of mimosine on wool growth of Merino sheep.

Merino sheep were given continuous intravenous infusions of L-mimosine for periods of 1 1/2, 2 or 21 days; efficacy as a defleecing procedure and effects on subsequent wool growth were measured. In addition, the amino acids tyrosine, phenylalanine and cystine were investigates as antagonists to the effects of mimosine. Infusions for 1 1/2 or 2 days at the daily rate of 80-120 mg/kg caused a cessation of wool growth by 1 1/2-2 days from the start of infusion, and all sheep were subsequently defleeced. It was estimated that, on average, fibre growth stopped for 10 1/2-13 days in four sheep after a 2-day infusion, and for 5 1/2 and 9 1/2 days in two sheep after an infusion for 1 1/2 days. There was considerable variation in the time taken for new fibres to recommence growth. During the period 3-5 weeks after infusion of mimosine, length growth rate was consistently greater than the pretreatment rate. Likewise, fibre diameter was greater in three out of the four sheep. As a result, the volume growth rate of fibres was greater post-treatment than it was pretreatment. Infusion for 3 weeks at the daily rate of 21-24 mg/kg did not stop wool growth. However, both length growth rate and fibre diameter were considerably depressed, and after 12 days' infusion, fibre diameter and volume growth rate were reduced to less than half the pretreatment values, and wool fibres were very weak. After the mimosine infusion stopped, fibre diameter increased to above pretreatment values and remained ther for the period of 2-3 weeks studied. The concurrent infusion of tyrosine, phenylalanine or cystine with mimosine failed to prevent any of the effects of mimosine on wool growth.     

Coarser wool is not a necessary consequence of sheep aging: allometric relationship between fibre diameter and fleece-free liveweight of Saxon Merino sheep

The mean fibre diameter (MFD) of wool is the primary determinant of price, processing performance and textile quality. This study determines the primary influences on MFD as Saxon Merino sheep age, by allometrically relating MFD to fleece-free liveweight (FFLwt). In total, 79 sheep were grazed in combinations of three stocking rates and two grazing systems (GS: sheep only; mixed with Angora goats) and studied over 3 years. Measurements were made over 14 consecutive periods (Segments), including segments of FFLwt gain or FFLwt loss. Using shearing and liveweight records and dye-bands on wool, the FFLwt and average daily gain (ADG) of each sheep were determined for each segment. The mean and range in key measurements were as follows: FFLwt, 40.1 (23.1 to 64.1) kg; MFD, 18.8 (12.7 to 25.8) μm. A random coefficient restricted maximum likelihood (REML) regression mixed model was developed to relate the logarithm of MFD to the logarithm of FFLwt and other effects. The model can be written in the form of β(A) is a random animal effect, γ(Segment.Plot) a random effect associated with Segment.plot combinations, and κ a constant that depends on GS, random animal effects, random Segment.plot combination effects, Segment and ADG. Thus, MFD was allometrically related to the cube root of FFLwt over seasons and years for sheep, but to the square root of FFLwt for sheep grazed with goats. The result for sheep grazed alone accords with a primary response being that the allocation of nutrients towards the cross-sectional growth of wool follicles is proportional to the changes in the skin surface area arising from changes in the size of the sheep. The proportionality constant varied systematically with ADG, and in sheep only grazing, was about 5 when sheep lost 100 g/day and about 6 when sheep gained 100 g/day. The proportionality constant did not systematically change with chronological age. The variation in the allometric coefficient between individual sheep indicates that some sheep were more sensitive to changes in FFLwt than other sheep. Key practical implications include the following: (a) the reporting of systematic increases in MFD with age is likely to be a consequence of allowing sheep to increase in size during shearing intervals as they age; (b) comparisons of MFD between sheep are more likely to have a biological basis when standardised to a common FFLwt and not just to a common age; (c) wool quality (MFD, staple strength) are most likely to be optimised in management systems that maintain constant FFLwt of adult sheep within and between years.     

LABORATORY STUDIES ON THE CHEMO-SENSORY REACTIONS OF BLOWFLIES

Attraction of various blowflies to sheep wool has been demonstrated under laboratory conditions using a choice-chamber technique. The attractive factor in wool is intimately associated with the wool fibre for it does not disappear during storage and it is not completely removed by washing.
Only female flies were attracted to wool. The highest degree of attraction was obtained from fertilized females which had daily access to meat. Even in the absence of fertilization and meat, attraction was not completely abolished.
The species and strains tested can be placed in the following order in terms of strength of response to sheep wool: Lucilia sericata (British), L. cuprina (tested on damp floor)>L. sericata (Danish: country)> L. sericata (Danish: city), L. sericata (Australian) > L. illustris, L. caesar. No response was given by Calliphora vomitoria.
The olfactory organs involved in the response to wool are mainly on the antennae but antennaeless L. sericata and L. cuprina with some tarsi removed were still able to show some response to wool provided that they were tested on a damp floor. In the course of testing the olfactory responses it was also shown that the main humidity receptors are present on the antennae.
The existence of a 'wool-factor' to which only certain strains of L. sericata and L. cuprina are specially sensitive offers a possible explanation for the greater efficiency of these species as myiasis-producing forms.     

Studies on the in vitro metabolism of [3H]cortisol and [3H]oestradiol by sheep skin and wool roots.

The in vitro metabolism of [3H]cortisol, [3H]cortisone and [3H]estradiol-17 beta by adult sheep skin and wool follicle tissue (wool roots) was examined. The main metabolic product of the incubation of [3H]cortisol with sheep skin was [3H]cortisone, and the conversion was reversible. Wool roots were unable to carry out detectable interconversion, nor did this tissue give rise to other significant metabolites. Sheep skin and wool roots both rapidly converted [3H]oestradiol-17 beta to [3H]oestrone and the conversion could be carried out by follicle and non-follicle skin structures. It is suggested that sheep skin contains both 11 beta- and 17 beta-hydroxysteroid dehydrogenases, but that wool follicles contain only the latter enzyme.