Inuence of graded levels of baobab oil seed cake on growth performance and enteric methane emissions in Savannah × Boer crossbreed yearling goats.

Background: The recent concern on the emission of methane from ruminant livestock that contributes to climate change has called for a search of feed ingredient that will mitigate the emission of this gas at the same time increase the growth performance of the animal. One of such feed ingredients could be baobab oil seed cake. This study therefore, investigate the effects of feeding graded levels of baobab oil seed cake (BOSC) on growth performance and enteric methane emissions of Savannah × Boer goats at yearling age. Methods: A total of 24 goats weighing 16.63 ± 3.639 kg were used in a completely randomized block design. The study was conducted for a period of 70 days, with an adaptation period of 14 days. The baobab oil seed cake was included in the diets at 0% (control diet), 15%, 30% and 45 % on a mass basis. The experimental diets were iso-nitrogenous and iso-caloric. The parameters measured include the average daily feed intake (ADFI), body weight gain (BWG), and average daily gain (ADG), feed conversion ratio (FCR), and enteric methane emissions using Laser methane detector (LMD). Results The inclusion of baobab oil seed cake (15%, 30%, and 45% BOSC) in goats’ diets signicantly increased (P<0.05) (ADFI). Females in the current study gained more weight than males. It was observed that goats fed diets with 0% baobab oil cake emitted higher methane output (P<0.05). Methane emission decreased as the baobab inclusion levels in the diet increase (P<0.05). Female Savannah ´ Boer goat emitted more methane than male (P<0.05). Both sex and baobab oil cake inclusion levels contributed signicantly to methane emissions individually and interactively. Conclusion: Inclusion of baobab oil seed cake in the diet improved growth performance and could also be used to mitigate methane production in Savannah × Boer crossbreed goats at a range of 15 to 45 % inclusion levels without causing any detrimental effects on the goats.’


Introduction
Livestock production contributes huge amounts of methane (CH4) to the global anthropogenic greenhouse gas (GHG) emission [1]. Also, over 12% of dietary energy ingested by ruminant are been lost through methane emission [2]. These has made greenhouse gas emissions (GGE) from livestock a major concern globally. According to [3], it was estimated that livestock sub sector of agriculture contributes approximately 18% of the greenhouse gases (GHG) annually all over the world [4]. Therefore, strategies for mitigation of methane emission from ruminants are both bene cial to the environment and could improve feed utilization by the animal. Currently, information on enteric methane emissions from Savannah and Boer crossbreed goat of South Africa varies from limited to non-existent. The increase in the population of Savannah and Boer crossbreed goat across Southern African region call for the reasons to estimate their greenhouse gasses contribution to global warming.
Among the factors that affect CH 4 emissions, level of feed intake of the individual animal, type diet as well as presence or non-presence of inhibiting factors in the diets contribute signi cantly to the amount of methane generated in ruminant animals [5]. Recently, numerous reports have shown reduction of methane emission in ruminant as a result of feeding tannin rich ingredient [6]. Tannins have anti-methanogenic effect, through a direct inhibition of methanogens or indirectly inhibition of protozoa [7,8]. Tannins are polyphenolic compounds that bind to protein thereby decreasing ruminal fermentation of proteins in the rumen [9]. They are complex polymers with different linkages and bonds that varies in different species of plants and also within different parts of plants [9,10].One of such non-conventional feed source that is rich in tannin is baobab cake. It been reported severally that baobab oil cake contains 2-12% tannin [11,12]. Baobab seed is rich in protein and contains substantial amount of energy [13]. Also, baobab seed oil cake could be a replacement for the conventional livestock feed sources (such as barley, maize, soya bean meal, sun ower oil cake) that are too expensive for smallholder farmers as protein and energy source. Baobab tree been a drought and re-resistant plant are cheap and very available even during the dry season [14,15].
Several techniques have been developed to measure the enteric methane emissions in ruminant animals [16]. The respiration chamber technique is considered as the gold standard method to quantify the enteric methane output of ruminant animals. However, estimates obtained from chamber measurements are not transferable to grazing ruminants because they are obtained in an arti cial environment [17]. The portable laser methane detector (LMD) technique may offer a reasonable alternative because it allows the quanti cation of CH 4 from animals in their natural environment [18] without interfering with the animal's behaviour [19].
.The LMD has been potentially used in eld-based measurements for detection of CH4 outputs like screening animals for breeding programs or assessment of alternative management strategy [20]. The LMD is a non-invasive technique that allows CH4 measurements on individual animals and may provide advantages over other eld-scale methods, such as the SF6 tracer technique, the polyethylene, tunnel or breath sampling methods [21]. The LMD is a handheld gas detector that measures CH4 concentrations of gaseous outputs from an animal during short periods of time. The objective of the study, therefore, is to evaluate the potential effects of baobab oil seed cake on growth performance and enteric methane emission through LMD in Savannah and Boer cross breed goats at yearling age. It was hypothesised that baobab oil seed cake will positively in uence the growth performance and enteric methane output of Savannah and Boer goat's crossbreed at yearling age.

Ethical approval
The experiment was conducted in compliance with the standards required by the Animal Ethics Committee of the University of Fort Hare (Ethical No: MPE011SSAH01). All trial procedures were conducted in accordance with standards of experimentation given by the committee of ethics on animal use of the Society for the Prevention of Cruelty to Animals (SPCA) (Constitutional Court of South Africa, 2013).

Study site
The study was conducted at the University of Fort Hare Teaching and Research Farm. The farm is 520 m above sea level, located at 32.8° S and 26.9° E with an average annual rainfall of 480 mm and a mean annual temperature of 18.7°C. It is situated in the False Thornveld of the Eastern Cape [22]. The topography of the area is generally at with a few steep slopes. The vegetation is a mixture of several trees, shrubs and grass species [22] 2.3.2. Animal management, diets, feeding and experimental design A total of 24 goats (yearling goats) weighing 16.63 ± 3.639 kg were used in the current study. The goats were de-wormed with Valbazen purchased from Umtiza, in Alice Town in the Eastern Cape province of South Africa, manufactured by Zoetis TM, dosage rate: 5ml for animals with 30Kg body mass, contains Albendazole 1.9% m/v, against internal parasites. The goats were vaccinated against anthrax, pasteurelosis and pulpy kidney using Hi-tet 120 purchased from Umtiza, in Alice Town in the Eastern Cape province of South Africa, manufactured by Bayer (Pty) Ltd animal health care, dosage rate: 2ml for animals with 30kg body mass. The goats were kept in well-ventilated individual pens situated in the animal house equipped with watering and feeding troughs. The experiment was conducted in randomized complete block design (RCBD) where sex is a blocking factor. The goats were assigned into four different treatments: T1, control (0% BOSC); T2 (15% BOSC), T3 (30% BOSC), and T4 (45% BOSC). All 24 goats were ear-tagged. Table 3 showed the experimental diets for each treatment. The feed was formulated to meet the nutritional (energy and protein) requirements of the used goats (NRC, 2007). The goats were given 14 days to adapt to the treatment diets, experimental pens, as well as human handling and presence before the commencement of the trials that lasted for 70 days. The animals were given 1.2 kg of feed twice a day (at 8:00 am and at 16h00 pm daily) across all treatments. The goats were provided with clean and fresh water ad libitum on daily basis using 5 litre buckets. The amounts of protein and energy were supplied based on daily energy and protein requirement on animals.

Average daily Feed intake
Feed intake (FI) was determined by subtracting the total amount of leftover and refused feed weekly from the feed supplied weekly. Rejected feed from feeding troughs was weighed; including refused feed every day in the morning. The amount of feed that disappeared was considered the feed consumed by the goats. Sacs were placed under all feeding troughs to collect feed spillages. Feed spillages and refused feed were sun dried, weighed, recorded and then discarded. Weights of feed refusals and spillages were subtracted from the total weight of the feed given to each goat weekly and divided by 7 to determine average daily feed intake (ADFI) [23]. Calculated body mass in kg was then determined using the formula; Weight gain = Weight in the current week -weight in the previous week. Then average daily gain (ADG) was measured by weighing the goats every week across the whole experiment. The difference in weight of goats at the beginning and end of each week divided by 7 determined the ADG [23].

Feed conversion ratio
Feed conversion ratio was calculated from average daily feed intake divided by average daily weight gain. The following formula was used: Feed conversion ratio = Feed intake/Weight gain Experiment 2 Twenty-four Savannah X Boer crossbreed goats of both sexes, weighing an average of 16.63 ± 3.639 kg, in randomized complete block design (RCBD) where sex is a blocking factor, were used. Enteric methane was measured using a hand-held LMD (Crowcon, Tokyo Gas Engineering Co Ltd 2006) in the 13:00 hours when the animals are no longer eating. The LMD equipment measures the concentration of CH4 between the equipment and the target point. It is based on infrared absorption spectroscopy and measures CH4 values as a plume [24][25][26]. Thus the measurements are in parts per million-metre (ppm-m) [25]. The equipment operates normally in the temperature range between 0 and 40 °C, in the humidity range of 20-90%, with a reaction time of 0.1 seconds. The LMD can detect CH4 concentrations between 1 and 50,000 ppm within a distance of up to 150 m. Gas column density was measured by directing the auxiliary LMD targeting (visible HeNe) laser beam at the nostrils of goats at a distance of 1.5m from the goat [26]. Prior the commencement of the measurements for each day, the LMD was off-set to adjust it to the ambient CH4. Off-setting the LMD is done to account for the CH4 in the environment before the machine records the gas concentrations from the point source [24]. The measurements were taken Weekly for a period of 9 Weeks.
The proximate analysis of the experimental diets was carried out as described by AOAC, 2005 (method 920.85), and is shown in Table 2. The relationships between goats' growth performance (ADFI, ADG, BWG, and FCR), time in weeks and inclusion levels of baobab oil seed cake are shown in (Table 3., Figure 1). As the level of the test diet increases, average daily feed intake increased generally from 1 to week 9 across all treatment diets (Table 3 and Figure 1). Feed conversion ratio in T1 (control diet) decreased linearly (P<0.05) with increase in time (from week1 to week 9). There was a positive quadratic relationship (P<0.05) between time (in weeks), FCR and BOSC inclusion levels between T2 (15% BOSC) and T4 (45% BOSC). There were no signi cant differences (P>0.05) observed on both average daily gains and body weight gains across all treatments. However, the results in Table 3 show that goats amongst all treatments continued to gain body mass linearly from week 1 to week 9 (Figure 2 and 3). Effect of graded levels of Baobab oil seed cake on enteric methane emission.
Enteric methane production is related to DMI whereby the more feed is consumed, the more methane is produced. The results in Table 5 show that goats that consumed more feed emitted signi cantly more methane gas compared to the ones that consumed less feed. It was observed that goats fed on diet with 0% baobab oil cake emitted higher CH 4 gas (CH 4 litres/day, CH 4 g/kg DMI, CH 4 g/day/head, and CH 4 ppm-m) than goats fed on diet supplemented with baobab oil cake (P<0.05). Female goats emitted more methane gas than male animals (P<0.05). The results in Figures: 1, 2,3 and Figure 4, show that methane production was not consistent (P<0.05) with time. Generally, from week 1 to week 4, animals emitted more methane with time; but from week 5 onwards there were a lot of uctuations. Graphical illustration in Figures: 1, 2,3 and Figure 4 also show that methane output was decreasing as the level of the test diet increases (0% BOSC, 15% BOSC, 30% BOSC, and 45%BOSC).    There was a general increase in feed intake across all treatments in the study, this could be attributed to an increase in palatability of the diets [15] which might be probably due to the aromatic characteristics of baobab oil seed cake as reported by [27]. This pattern could also be a re ection of the relative acceptability and palatability as was reported by [28].Similarly, [29] reported that signi cantly higher concentrate intake in treatment groups fed West African Dwarf goats maybe an indication of the palatability of the concentrate with fermented baobab inclusion(P<0.05). Also, [30] noted that young goats fed diet containing 40 % baobab leaf meal had higher dry matter intake when compared to other leaves as Blighia sapida , Entada africana and Gliricidia sepium. The ndings on the increase in ADFI with an increase in time (in week) correspond to that of [31], who observed an increase in feed intake with an increase in age (week) of all broilers fed graded levels of cotton seed meal. This might be a similar case with goats fed graded levels of baobab oil seed cake. Generally, when animals are growing there is a possibility that feed intake also increases due to the fact that feed requirement of an animal is determined by its body weight.
The results on sex and treatment effect correspond with that of [32], who reported that dry matter intake was higher in male castrates than in female kids during the interval of 21 to 24 week , and 17 to 32 week of age. These effects could be largely due to differences in body size between male castrates and female kids. Contrary, a study conducted by [33] reported that sex of goats had no signi cant difference on dry matter intake (P>0.05), this could be attributed to the fact that feed intake on animals is more in uence by feed palatability, as well as appetite on an individual animal, which means that either male nor female, any animal that has high appetite is more likely to have higher feed intake, regardless of sex.
Body weight gains (ADG, BWG) The increase in body weight gains reported in all the treatments is an indication that the goats responded positively to all the dietary treatments and that protein content and dietary energy supply of the experimental diets adequately enhanced growth performance of the goats. There is a general increase in weight gains amongst all treatments, thus implying that the goats were able to convert feed protein into extra muscles. Weekly changes on body weight gains could also be attributed to the adaptability of the animals to dietary treatments. The results obtained in the trial aligns with the report of [34], who observed that there were no signi cant differences (P>0.05) in body weight gains on rabbits fed 5% and 10% BOSC when compared to those that were placed in the control diet. Regardless of the fact that there were no statistical differences (P>0.05) on body weight gains, goats that were subjected to the diet containing 45% baobab oil seed cake inclusion levels were superior in body weight gains compared to other treatments. This might be attributed to the differences in nutritional composition amongst the dietary treatments. In this study, females had higher ADG values than males. However, [35] reported a contrary results that Boer-cross bucks that were fed Marshall Ryegrass had higher ADG compared with Does. Findings in the current study are different from what was expected, male goats were expected to be more superior on body weight gains than females, due to the presence of growth hormone testoterone. This might be explained by the fact that, males' goats were castrated 4 weeks before the commencement of the feeding trial, thus it might happen that some of them were not yet healed, hence growth rate was delayed by castration stress. As shown in table 2, incorporation of baobab oil seed cake in the diet and sex signi cantly increased (P<0.05) body weight gain, with female's notable having higher BWG than males at 0, 15, and 45% BOSC, while only males fed on diet with 30% BOSC have higher BWG than females. The inconsistency found in the current study could be attributed to the above-mentioned reason that male goats that were used in the study were castrated 4 weeks before commencement of the study, hence it could happen that some males had not yet recovered, hence castration stress had negatively affected body weight gains in males.

Feed Conversion ratio (FCR)
There are limited studies to support or dispute results, on weekly changes of feed intake and growth performance parameters. Several studies focus more on treatment effects, but generally, feed conversion ratio is negatively correlated with body weight. Increase in body weight leads to a decrease in FCR. Nevertheless, lower FCR means that animals were able to utilize feed e ciently. The results on FCR can be supported by similar trend that was observed on ADFI, ADG, BWG, this concludes that as the goats were growing on weekly basis, feed intake and growth rate was also increasing, thus goats were able to convert feed into muscle. The results in this study are different from [36] who reported lower FCR in diet with 10% BBSM inclusion level and higher FCR at 15% BBSM inclusion level. This increase in FCR was associated with increase in anti-nutrient content of the diet with increase in BBSM inclusion level.
Effect of Baobab oil seed cake on enteric methane emission.
The results in the current study show that animals that consumed more feed, emitted signi cantly more methane gas compared to those that consumed less feed, dry mater intake being the major determinant [37]. Also, previous ndings reported that approximately 95% of methane that is emitted during enteric fermentation is lost to the atmosphere via nose, while only 2-3% is lost via rectal emissions [38].
Nevertheless, the CH 4 emission varies largely, depending on various factors, with dry mater intake (DMI), digestibility and diet composition being the most important determinants of methane production [38]. [39] reported that there is an inverse relationship between gross energy intake and methane production. In the current study, methane output depletes as the level of baobab oil seed cake increases. This could be attributed to the presence of tannins in baobab seeds. The Baobab oil cake contains 2-12% tannin [40,41]. The decline in enteric methane emissions with increase in inclusion of tannins in ruminant diet was similarly reported by several authors [42,43]. Also, [44] reported that tannins may modulate ruminal fermentation favourably, thereby decreasing protein degradation in the rumen, preventing bloat, as well hindering of methanogenesis. Tannins-containing forages have been an evidence that tannins reduce methane emissions both in vivo and in vitro [16]. Similarly, [45] reported that tannins have a potential to degrade enteric CH 4 production in ruminants as a result of decrease in bre digestibility.
In general, tannins could reduce methanogenesis through reductions of numbers of protozoa, methanogens and depressed bre degradation depending upon the chemical structure of tannins and methanogen species [46]. In its simplest form, propionate is considered to reduce nitrogen. This means it reduces the amount of H 2 available for methogens, thus methane production declines. The three main VFAs are generally produced relatively to each other at a ratio of 70: 20: 10 for acetate: propionate: butyrate. Nevertheless, these portions can be altered by dietary manipulation to reduce the ratio of acetate to propionate in particular to less than 0.5 [47]. This will help to avoid access of H 2 , which might increase methane production. Nevertheless, if hydrogen produced is not used correctly, ethanol or lactate can be formed, hence inhibiting microbial growth, and further production of VFAs. Enteric methane emission is reduced if the ow of hydrogen shifts towards alternative electron acceptors [37].
In the current study, the results show that female goats exhibited more methane gas output than male goats. However, there is limited literature to support or dispute the effects of baobab oil seed cake on enteric methane emissions. The results in the current study could be ascribed to the fact that, female goats generally consumed more feed than male goats; hence DMI was higher in females than in males, as a result of that, male goats emitted less methane gas than female gas. Findings in the current study contradict that of [37], who reported that male goats produced more methane gas than female goats. This could be possible due to the fact that males animals normally, are the ones that usually consume more feed (higher DMI) than female animals, thus that was the case with [37]. The result in gure 1, 2, 3 and 4 show that methane production was not consistent with time. Generally, from week 1 to week 4, animals emitted more methane with time; however, there were lots of uctuations from week 5 onwards. This could be explained by the fact that animals were adapting to the dietary treatments with time, hence they were consuming more feed thus methane gas production was increased.

Conclusion
The incorporation of baobab oil seed cake in the goats' diet improved growth performance of goats. The sex of the animal also affected ADFI, and growth performance of goats. Sex of the animal signi cantly affected methane production in goats, with female animals producing more methane gas than males.
Sex and treatment diets both contributed signi cantly to CH 4 emissions individually and interactively. Therefore, BOSC can be incorporated in the goats' diet up to 45% without causing any detrimental effects on goat's well-being. Also, baobab oil seed cake in goats' diet also signi cantly reduced enteric methane production better than soy bean meal based diet (control diet). Hence, baobab oil seed cake could improve overall performance of cross breed goats and contribute signi cantly in stabilising climate change by reducing methane output in goat production.

Declarations
Authors' contributions Saho Samkelo and Mupangwa conceived the research, Moyo formulated the diets, Saho and Ikusika were involved in the feeding trials, writing and design while Mpendulo and Mupangwa carried out the statistical analysis and supervision of the study.
Data availability Data will be made available on request. Figure 1 The effects of baobab oil seed cake inclusion levels and weeks on average daily feed intake Figure 2 The effects of baobab oil seed cake inclusion levels and weeks on average daily gain Figure 3 The effects of baobab oil seed cake inclusion levels and weeks on body weight gain Relationship between time in weeks and level of baobab inclusion on methane (litres/day) Figure 5 Relationship between time in week and baobab oil seed cake inclusion level on methane (kg/day/head)

Figure 6
Relationship between time in weeks and inclusion level of baobab oil seed cake on methane (g/kgDMI).

Figure 7
Relationship between time in weeks and inclusion levels of baobab oil seed cake on enteric methane (ppm-m).