ABSTRACT
Objective:
This study aims to provide comparative information on the rumen ciliate fauna of goat (Capra aegagrus hircus) and sheep (Ovis aries) living in Zawiya, Libya.
Methods:
We obtained rumen samples from 16 goats and 17 sheep after the slaughter in Zawiya, Libya between June and August 2016. We immediately fixed the well-mixed samples with an equal volume of 18.5% formalin. We filtered and stained the samples in the laboratory with methyl green formalin saline solution to determine the nuclei and added 2% Lugol’s iodine solution to visualize the skeletal plates.
Results:
We found that the mean number (± standard deviation) of ciliates in the rumen contents from goats and sheep was 70.9±61.6×104 cells mL-1 (minimum-maximum value, 4.0-187.0×104 cells mL-1) and 96.3±49.3×104 cells mL-1 (minimum-maximum value, 19.5-235.0×104 cells mL-1), respectively. Results also showed that the total number of species per goat and sheep was 1-17 (mean, 8.2±4.7) and 1-13 (mean, 7.9±3.8), respectively. We identified 10 genera, 19 species, and 11 morphotypes in goats and 9 genera, 16 species, and 13 morphotypes in sheep. Additionally, we found that Entodinium simulans prevalence in all goats and sheep was 100%. On the other hand, we observed Hsiungia triciliata and Ostracodinium gracile in only one goat (6.3% prevalence) and Polyplastron multivesiculatum in only one sheep (5.9% prevalence). Overall, the ruminal ciliate fauna of goat and sheep in Libya comprised Entodinium species (mean for goats, 85.9%; mean for sheep, 83.5%).
Conclusion:
This study recorded Hsiungia triciliata as a new endosymbiont in goats. To our knowledge, this study is the first to report all of the species detected in goats from Libya. Similarly, this is the first to detect Diplodinium anisacanthum, Entodinium bursa, E. ellipsoideum, E. longinucleatum, E. simulans, Isotricha prostoma, Ophryoscolex caudatus, Ostracodinium gracile, and Polyplastron multivesiculatum in sheep from Libya.
INTRODUCTION
The rumen is the main example of an extremely diverse, densely colonized microbial habitat, which contains bacteria, archaea, and protists (1). Rumen ciliated protozoa are the prime eukaryotic component of the rumen ecosystem and can contribute up to 50% of the biomass in the rumen (2). These symbiotic ciliates belong to the subclass Trichostomatia and can utilize a vast array of carbohydrate compounds, such as soluble sugar, starch, and lignocellulose (3,4). Rumen ciliate species composition and concentration vary the type and amount of feed consumed, pH, turnover rate, and frequency of feeding (5,6). Trichostomatid ciliates are classified into two orders Vestibuliferida and Entodiniomorphida. Most of the rumen ciliates are composed of ophryoscolecid ciliates in the order Entodiniomorphida (3,7,8). Despite the general morphology of ophryoscolecid ciliates shows a great uniformity, these ciliates have a monophyletic origin (9,10). Transfaunation of rumen ciliates occurs only by direct contact of hosts throughout their lifetime (11).
Libya is in the African continent, and there has been no study on the rumen ciliates of goats living there. Therefore, the aim of this study to provide comparative information on the rumen ciliate fauna of goat (Capra aegagrus hircus) and sheep (Ovis aries) living in Zawiya, Libya.
METHODS
Rumen samples were obtained from 16 goats (C. aegagrus hircus) and 17 sheep (O. aries) after the slaughter in Zawiya, Libya between June 2016 and August 2016. The well-mixed samples were immediately fixed with an equal volume of 18.5% formalin (12). In the laboratory, they were filtered and stained with methyl green formalin saline (MFS) solution to get differential and total cell counts (13). The MFS solution was used to indicate nuclei and the solution of 2% Lugol’s iodine was added to samples to visualize the skeletal plates (14,15).
The mean number of ciliates in each goat and sheep was determined using the hemocytometer counting chamber. The relative abundances of ciliate species in each goat and sheep were estimated from smear slides (16,17). The prevalence calculation of ciliate species was performed as previously described by Bush et al. (18).
The species were identified and classified based on previously published species descriptions and taxonomic lists (8,13,19-21).
Statistical Analysis
Statistical analysis of rumen ciliates was performed by the SPSS statistics 20 programs.
RESULTS
The mean number (± standard deviation) of ciliates in the rumen contents from the 16 goats and 17 sheep living in Libya was 70.9±61.6×104 cells mL-1 (minimum-maximum value, 4.0-187.0×104 cells mL-1) and 96.3±49.3×104 cells mL-1 (minimum-maximum value, 19.52-35.0×104 cells mL-1), respectively. The total number of species per goat and sheep was 1-17 (mean, 8.2±4.7) and 1-13 (mean, 7.9±3.8), respectively.
The relative abundance and the prevalence of genera and species in the rumen contents of goat and sheep in Libya are shown in Table 1. In goat, 10 genera, 19 species, and 11 morphotypes were identified, whereas, in sheep, 9 genera, 16 species, and 13 morphotypes were detected. Entodinium simulans was found in all goats and sheep with a prevalence of 100%. Hsiungia triciliata and Ostracodinium gracile were observed in only one goat (6.3% prevalence), likewise, Polyplastron multivesiculatum was observed in only one sheep (5.9% prevalence). In general, the ruminal ciliate fauna of goat and sheep in Libya was composed of Entodinium species (mean for goats, 85.9%; mean for sheep, 83.5%). H. triciliata is a new endosymbiont recorded for the goats. All of the species detected from the goats are the first report in Libya. Diplodinium anisacanthum, Entodinium bursa, E. ellipsoideum, E. longinucleatum, E. simulans, Isotricha prostoma, Ophryoscolex caudatus, O. gracile, and P. multivesiculatum are first detected from the sheep in Libya.
DISCUSSION
The present study is the first for the rumen ciliate fauna of goats and the second for the rumen ciliate fauna of sheep in Libya. The mean number of ciliates in the rumen contents of sheep living in Zawiya, Libya (96.3±49.3×104 cells mL-1) was less than that of sheep in Triple city, Libya (220×104 cells mL-1). The number of ciliate species in sheep living in Zawiya, Libya was higher than sheep living in Triple city, Libya (Table 2). These variations may be the differences in the kinds and amounts of food consumed, host animals, geographical location, or a combination of these factors (15,22,23).
In the present study, 10 genera, 19 species, and 11 morphotypes were found in goats and 9 genera, 16 species, and 13 morphotypes were found in sheep. Although this is the first report on rumen ciliates of goat in Libya, no novel species were detected. However, Hsiungia triciliata is recorded for the first time as a new endosymbiont for goats. Before, H. triciliata was detected from the dromedary camels in Zawiya, Libya (24) and also reported from the camels in Triple city, Libya (25). In addition, H. triciliata was found from the dromedary camels and cattle in Tunisia (26). The goat may have obtained this ciliate from close contact with cattle or camels because H. triciliata was not detected from sheep.
In the rumen contents of goats and sheep in Zawiya, Libya, Entodinium spp. were generally predominant, it may be feeding habits of goats and sheep. Although goats and sheep are the intermediate feeders, goats are browsers while sheep are grazers. Goats consume concentrate selection such as leaves and tree sprouts (27-29), whereas sheep eat mostly grass, herbaceous flowering plants, and forbs (30). Goats were fed on steppe shrubs and sheep were fed on mixed-grass steppes and meadows in Libya. Additionally, cereal grain straw (barley, oat, and wheat hay) is the dietary source of both animals. The rumen of the intermediate feeders may have environmental factors more favorable for the growth of Entodinium spp. than those of true browser and true grazer (31). If the host is fed a high concentrate ration, the composition ratio of Entodinium spp. and their total density become higher because Entodinium spp. grow rapidly (32). The starch is the principal source of energy for Entodinium spp. (33). The host species itself may exert some degree of control over the specific fauna in its rumen. The selecting of particular foods and the quantities consumed by different host species are important factors on the rumen ciliate fauna (31,34). In addition, Entodinium spp. are normally predominant in the rumen and have a wide distribution in almost all ruminants, worldwide (35,36).
Of the ciliate species detected from the goats and sheep, Entodinium simulans was the most abundant (100%), it may be the wide distribution of E. simulans or the diet of the host. In the present study, H. triciliata and Ostracodinium gracile were reported only in one goat, likewise, Polyplastron multivesiculatum was found only in one sheep. These ciliates could have had rare opportunities for transmission between hosts, resulting in very limited distribution. In goat and sheep, Diplodinium, Epidinium, Ophryoscolex, Ostracodinium, and Polyplastron were observed. Although Metadinium is present in goats, it was not reported from sheep, whereas Enoploplastron is present in sheep, it was not observed from goats. The species of genera Diplodinium, Epidinium, Ophryoscolex, Ostracodinium, Polyplastron, Metadinium, and Enoploplastron would be favorable for host animals fed mainly fresh and dried grass with low nutritive value (37). These ciliates have been considered to possess the cellulolytic activity and ingest many fragments of plants (21,32,36,38).
In Libyan goats, the density of E. simulans morphotype with caudal spines is high, conversely, the density of E. simulans without caudal spines is high in Libyan sheep. When the hosts are fed a diet with low starch value, ciliates without caudal spines are predominant, whereas ciliates with well-developed caudal spines become predominant when the hosts are fed a diet with rich starch (39). It is considered that the caudal spines of Entodinium and Epidinium served to protect against engulfment of Entodinium bursa and P. multivesiculatum, respectively (40) because they are larger and carnivorous ciliates. However, the exact reason of spine development is not known. The rumen ciliate fauna of a goat and a sheep in Libya is composed of only Entodinium spp. It could be that Entodinium only fauna is the result of an extremely low pH, a rapid rate of passage of fluid and particulate matter through the rumen, ingestion of specific toxic or inhibitory substances, or a combination of all these factors (32,41-43) but it is not confirmed.
Rumen ciliate populations of ruminants have been grouped into four main types (13,21,44-46). All four groups include the genera Entodinium, Dasytricha, and Isotricha but are differentiated from each other based on the presence or absence of specific species. The A-type ciliate population is designated having P. multivesiculatum and usually, but not always, Metadinium affine. The B-type ciliate population contains Epidinium sp., Eudiplodinium maggii, or both. The K-type ciliate population is found in cattle populations containing Elytroplastron bubali. The O-type ciliate population includes only Entodinium, Dasytricha, and Isotricha. The A-type ciliate population and B-type ciliate are not present in the same host, because the predatory activity of P. multivesiculatum can eliminate E. maggii and Epidinium spp. Six of the 16 Libyan goats had the B-type, the remaining five and two of ten were the A-type and, the O-type, respectively. The remaining three goats contained the ciliates of the A-type and B-type together. Twelve of the 17 Libyan sheep had the B-type, the remaining four of the five were the O-type and the other one was the A-type. Besides, seven of seventeen sheep had Epidinium ecaudatum and Ophryoscolex together. Eadie (47) reported that it was not possible to establish Epidinium and Ophryoscolex in the same host. In the present study, the occurring of two species and the A-type and B-type population together in the same host indicate that these animals can live in the same area and they have many opportunities for cross-infection to each other.
CONCLUSION
Geographical distribution, phylogenetic factors, and feeding habits of the host species, PH in the rumen, antagonism, and transfaunation among the ciliate species have significant effects on the occurring rumen ciliate fauna.
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