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Feeding Dayak onion (Eleutherine palmifolia) extract and Lactobacillus acidophilus mixture on blood biochemicals, meat quality characteristics and growth performance in broiler chickens

Iis Yuanita1,2, Dwi Sunarti1, Hanny Indrat Wahyuni1 and Nyoman Suthama1

1 Department of Animal Science, Faculty of Animal and Agricultural Sciences, Diponegoro University, Semarang 50275, Central Java, Indonesia
2 Animal Science Study Program, Faculty of Agriculture, Palangka Raya University, Palangka Raya 73112, Central Kalimantan, Indonesia


The study was conducted to evaluate the effect of feeding Dayak onion (Eleutherine palmifolia) extract and probiotic Lactobacillus acidophilus (EpLa) mixture on blood biochemicals, meat quality characteristics, and growth performances in broiler chicken. A total of 192 birds of unsexed one-day old broiler (CP 707 strain) were randomly divided into 4 dietary treatments and 6 replications with 8 birds each. The dietary treatments were basal diet as a control diet (EpLa0), basal diet added with 0.1% EpLa (EpLa1), 0.2% EpLa (EpLa2) and 0.3% EpLa (EpLa3). Blood antioxidant enzyme superoxide dismustase (SOD) in broiler was significantly enhanced (p<0.01), while the malondialdehyde (MDA) was significantly decreased (p<0.05) with the increasing level of feeding EpLa (Tabel 2). Increasing dietary inclusion of EpLa also significantly (p<0.01) improved meat characteristics as indicated by higher phenolic and flavonoid content, and antioxidant activity, and significantly (p<0.01) lowered meat cholesterol mass (Table 3). The improved meat characteristics was followed by the increased (p=0.06) body weight gain and significantly improved ( p<0.05) the feed conversion ratio when compared to control group. In conclusion, the mixture of Dayak onion extract and Lactobacillus acidophilus at 0.3% (EpLa3 treatment) effectively enhances meat characteristics and growth performance through the increase in antioxidant activity of broiler chickens.

Keywords: antioxidant enzyme, broiler, flavonoid, probiotic


Dietary inclusion of antibiotics as growth promoters (AGPs) in poultry production has dramatically increased for the last decades worldwide. It has been reported previously that AGPs have long been used as growth promoter and disease control (Plail 2006; Akinleye et al 2008). Recently, AGPs have been banned in many countries due to the negative consequences for health and food safety with special concern to the development of resistant pathogenic bacteria and residue in animal products. The ban on AGPs in poultry feed and feeding has prompted nutritionists to look for and to investigate an alternative substance beneficial for the improvement of poultry production performance with “clean product” without residue.

Various alternative feed additives derived from natural sources have been studied in the effort to improve feed efficiency and to enhance the health status of poultry. The active compounds originated from plants have several health benefits with its respective biological function as prebiotic, antioxidant, immunostimulant or antimicrobial activity (Patra 2012). Similarly, other natural compound has been reported that the use of probiotics in poultry feed improved the immune response and balanced the intestinal microbes (Das et al 2012). The increased intestinal lactic acid bacteria and the decreased total Coliform in broiler fed diet with probiotic Lactobacillus sp (Cholis et al 2018), and in native chicken given a combination of Lactobacillus sp and inulin (Purbarani et al 2019) are the indication of bacterial balance improvement.

Dayak onion (Eleutherine palmifolia), also called as either berlian, sabrang or tiwai onion, is a kind of plant-producing bulbs that belonging to the genus of Iridaceae. This onion was reported to have high active compounds, specifically rich in phenolic and flavonoid contents (Wang et al 2010). Phenolic compounds and flavonoids serve as very strong antioxidants and also contain an antibacterial agent (Scalbert et al 2005; Vauzour et al 2010). Some flavonoid derivatives such as phenolic acid had no inhibiting effect on growth of lactic acid bacteria, but it depressed the growth of gram negative bacteria like E. coli and Salmonella sp (Puupponen-Pimia et al 2001). In addition to antioxidant compound, onion extract also contain FOS and inulin prebiotics as much 22.96 and 15.59 g/100g dry matter, respectively (Mabrok et al 2018).

Currently, Lactobacillus sp is the most widely used probiotic that can be used for broilers (Gaggia et al 2010; Cholis et al 2018; Wulandari et al 2018), one of them is Lactobacillus acidophilus (Shoeib and Madian 2002; Sugiharto 2016). Lactobacillus acidophilus lives in the digestive tract of animal. It may degrade the simple carbohydrates into lactic acid, that has an impact to reduce the pH of the digestive tract and inhibit the growth of pathogenic bacteria (Gaggia et al 2010; Sugiharto 2016). It was reported that the reduced intestinal pH due to the presence of SCFA produced by fermentation activity of Lactobacillus sp on dietary low molecular weight carbohydrate, lead to the decreased growth of Coliform (Cholis et al 2018; Purbarani et al 2019). Other study showed that probiotic bacteria has another function as a source of antioxidants as reported by Sugiharto et al (2018). According to those described above, the treatment using probiotic bacteria is expected to improve the antioxidant status of broiler chickens.

Apart from the positive effects of a single use of bacterial species as probiotic, its effect when mixtured with Dayak onion extract is needed to be clarified. Although an in vitro study has been previously conducted with the best result was the mixture of Dayak onion extract at 75% and Lactobacillus acidophilus counts at 108 cfu/ml, based on total phenolic, antioxidant activity, and Lactobacillus liveability test (unpublished data). However, the information on the use of Dayak onion as a source of active compound additive in poultry diet is still very rare, so as the mixture of Dayak onion and Lactobacillus acidophilus may function as sources of prebiotic and probiotic that would be beneficial for broiler health and productivity. Therefore, research needs to be conducted to clarify the effect of Dayak onion extract and probiotic mixtured in broiler, in the effort of increasing productivity.

The present study was conducted to evaluate dietary mixture of Dayak onion extract and Lactobacillus acidophilus with the expectation that both ingredients could synergiticaly work, as prebiotic and probiotic additive, and become to be more beneficial for the health status and productivity of broiler chickens. Thus, the specific aim of this study was to evaluate the effect of feeding a mixture of Dayak onion extract and Lactobacillus acidophilus on blood biochemicals, meat quality characteristics, and growth performances of broiler chickens.

Material and methods

Dayak onion extract and Lactobacillus acidophilus mixture preparation

The bulbs of onion layers were separated and thinly sliced, ​​ air dried at room temperature thereafter. It was further mashed using blender and extracted with methanol solution at the ratio of 1: 4 (w/v) for 3 days. The extract liquid was stirred at least once a day and filtered (Whatman No. 1) on day 4. The extract liquid was evaporated in a rotary vacuum evaporator, until paste form was obtained and then stored at refrigerator temperature until further use.

The pure isolat of Lactobacillus acidophilus FNCC 0051 was obtained from Biotechnology Laboratorium Gadjah Mada University. To obtain the target population of Lactobacillus acidophilus (108 cfu/mL) the culturing procedure was performed at the Laboratory of Animal Nutrition Physiology and Biochemistry Diponegoro University. The levels of Dayak onion extract and Lactobacillus acidophilus mixture were based on the result of the previously-conducted in vitro study by evaluating 3 levels Dayak onion extract (25, 50, 75%), and 3 different counts of Lactobacillus acidophilus (106, 10 7, 108 cfu/ml). The best result found was the mixture of 75% Dayak onion extract and 108 cfu/ml Lactobacillus acidophilus, based on the total phenolic, flavonoid, antioxidant activity, and Lactobacillus liveability test. Therefore, Dayak onion extract (75% extract in sterile water) and Lactobacillus acidophilus bacteria (10 8 cfu/mL) were mixed with the ratio of 1:1 (v/v) becoming a single formulation, called EpLa as a unified mixture that was applied for the treatments material as assigned in the present study.

Experimental animal and diet

A total of 192 birds of unsexed one-day old broiler (CP 707 strain) were randomly divided into 4 dietary treatments and 6 replications (8 birds each) with initial body weight of birds was 46.43+1.65 gram. The chickens per replication (8 birds) were kept in a 1x1 m in size of litter-floored cage, and given diet ad libitum and free access of drinking water until 42 days of age. For the first 7 days, the chicks were fed with commercial diet (3,070 kcal/kg ME, 20.5% CP, 5% fat, 0.9% Ca and 0.6% P) and continued with basal diet. Basal diet was formulated according to the requirements recommended by Indonesian National Standards (SNI) No 01-3930-2006 as shown in Table 1. The levels of EpLa at 0, 0.1, 0.2, and 0.3% were added into the basal diet and were created as dietary treatments with the code of EpLa0, EpLa1, EpLa2, and EpLa3, respectively.

Table 1. Composition of basal diet


Composition (%)

Starter (<21days)

Finisher (22-42 days)







Soybean meal



Meat bone meal






Mineral and vitamin premix*






Proximate analysis

Metabolisable Energy, kcal/kg



Crude protein, %



Crude fiber, %



Ether extract, %



Ash, %



Calsium, %



Phosphor, %



Metionin, %



Lysine, %



Arginine, %



* Content per kg: calcium 32.5%, phosphor 1%, iron 6 g, mangan 4 g, iodine 0.075 g, copper 0.3 g, zinc 3.75 g, vitamin B12 0.5 mg, vitamin D3 50,000 IU
** Based on analyzed value at the Laboratory of Animal Nutrition and Feed Science, Faculty of Animal and Agricultural Sciences, Diponegoro University

Sampling and parameters measurement

Broiler performances measurement were feed consumption, body weight gain (BWG), and feed conversion ratio (FCR). Feed consumption was the sum of daily recording, and BWG was obtained by substracting final body weight with initial body weight. Feed consumption divided by BWG can be found FCR. On day 42, three birds of each replication were randomly selected. One bird was slaughtered to obtain carcass weight as well as whole meat samples, while the rest were used for collecting blood sample. The blood sample was taken from the wing vein with a 3 ml syringe and put into a vacuum tube. The blood samples were then centrifuged at 2500 x g for 10 minutes to obtain serum.

The measurement of serum level of malondialdehyde (MDA) was performed using thiobarbituric acid (TBA) reagent according to the Esterbauer and Cheeseman (1990) method. A half mL serum (0.5 mL) and 4.5 ml phosphate buffer saline (PBS) was mixed and centrifuged for 15 minutes, and 4 ml supernatant was taken. The supernatant was mixed with 1 mL trichloroacetic acid (TCA) 15% and 1 mL TBA, and heated in a waterbath at 80° C for 15 minutes, then cooled at room temperature for 60 minutes. The solution was centrifuged for 15 minutes, and then the absorbance was read in a spectrophotometer at λ 532 nm. MDA concentration (nmol/mL) was determined based on the standard curve of 1,1,3,3-tetramethoxypropane (Erdemir et al 2008). Superoxide dismutase (SOD) was analyzed using Ransod Superoxide Dismutase Manual Rx Monza reagent, and the measurement was performed using spectrophotometer UV-Vis at 505 nm according to the manufacturer’s instruc­tions. Enzyme activity was expressed as units per milliliter (U/mL) of the sample.

Breast meat was analyzed for its total phenolic content according to Folin-Ciocalteu method (Tabasum et al 2016) with a slight modification. The meat extract (0.5 mL), Folin-Ciocalteu reagent (1:10 in deionized water) and sodium carbonate (Na 2CO3, 4 mL) were mixed, and incubated at room temperature for 30 minutes with intermittent shaking. The absorbance of the solution was spectrophotometrically measured at 765 nm. A standard curve was subsequently plotted using gallic acid as phenolic standard.

The total flavonoid content (TFC) of meat was determined by spectrophotometric method according to Silva et al (2015) and Ramos et al (2017). The meat sample was extracted under reflux conditions (80°C) with 20 mL water‑ethanol solution 60% (v/v) (pH = 5.06) during 60 min. The extract was cooled to room temperature and filtered. The residue was re‑extracted under equivalent conditions. Both hydroalcoholic-disolved extract and re‑extract were combined, and the volume was completed to 50 mL of water‑ethanol solution 60% (v/v), resulting in the stock solution. A part of aliquot of the stock solution was transferred to a 10.0 mL volumetric flask and made to volume with methanol, resulting in the blank solution. A second part of aliquot of the stock solution was transferred to another 10.0 mL volumetric flask, a volume of the 2% AlCl3 was added and made to volume with methanol to obtain test solution. After 25 min the absorbance of the test solution was measured at 430 nm against blank solution. The results of TFC of meat sample were calculated, as quercetin, and were expressed as the amount of flavonoid (mg/g) after moisture content correction.

Antioxidant activity was determinated by the DPPH test (Sochor et al 2010). 15 μL meat sample solution was incubated in a 150 μL volume of reagent consisting 0.095 mM 2,2-diphenyl-1-picrylhydrazyl - DPPH•. Absorbance was measured at 505 nm for 10 minutes, thereafter.

Cholesterol contents of mixed meat were analyzed according to enzymatic in vitro method for the quantitative determination using specific reagent of cholesterol. Meanwhile, determination of protein content of meat was performed based on a slight modification of AOAC method (AOAC 2007). The percentage of meat protein content was then multiplied by the whole weight of meat to find protein meat mass as it has been reported elsewhere (Suthama et al 2018).

Experimental design and statistical analysis

The level of Dayak onion extract and Lactobacillus acidophilus mixture (EpLa) at 0.2% was based on the value after standarization with the level of total phenolic and antioxidant compounds from Echinacea purpurea for broiler as recommended by Lee et al (2012). Due to the different source (Dayak onion) used in the present study, EpLa was then developed into lower (0.1%) and higher (0.3%) levels compared to group without any additive addition. Therefore, a completely randomized design (CRD) was assigned in the present study with 4 treatments and 6 replications. The treatments tested were dietary additional levels of EpLa at 0, 0.1, 0.2, 0.3% with the code of EpLa0, EpLa1, EpLa2, and EpLa3, respectively. Data were statistically analyzed using GLM procedure of SAS version 9.0 and the differences among treatment means (p<0.05) were determined using Duncan’s multiple range test.

Results and Discussion

Blood biochemicals

Dietary inclusion of EpLa did not affect blood blood levels of cholesterol and triglyceride because no differences among treatment means could be observed (Table 2). Blood circulating levels of cholesterol and triglyceride are known to depend on either endogenous sources or exogenous factor derived from feed source availability. The endogenous source is considered to be not different since the same strain and age of the birds were used. However, dietary inclusion level of EpLa was the important factor that affects the blood concentrations of cholesterol and triglyceride. The maximum dietary addition of EpLa in the present study was until 0.3%, (EpLa3) and with this level the combining effects of flavonoid from Dayak onion extract and Lactobacillus acidophilus is assumed to be not enough in lowering pH of intestinal tract. It can be compared to the previously reported results that dietary inclusion of prebiotic inulin (Krismiyanto et al 2014) and Lactobacillus sp. (Cholis et al 2018) were able to reduce intestinal pH that aided to stimulate growth of endogenous lactic acid bacteria, leading to the increased production of bile salt hidrolase (BSH) enzyme. It well documented that BSH enzyme is able to inactivate the work of bile salt from pancreas to be deconyugated form so that decreases fat digestion and absorption, and finally impact on reduced sirculating levels of cholesterol and trigliseride. However, the change in gastrointestinal physiology condition did not happen in the present investigation. As the results the unchange blood concentration of cholesterol and trigliseride can be found (Table 2). This study was supported by the results reported by Aditya et al (2017), and Goodarzi et al (2013) that dietary inclusion of onion did not exert any change on blood serum levels of cholester­ol and triglyceride of broiler.

Table 2. Effect of Dayak onion extract combined with Lactobacillus acidophilus on blood biochemical parameters







p value

Chol., mg/dL







TG, mg/dL














MDA, ng/mL







a-b Means within row followed by different superscript differ at p<0.05, Chol (Cholesterol), TG (triglyceride),
SOD (superoxide dismustase), MDA (malondialdehyde),EpLa0 (basal diet), EpLa1 (basal diet added with 0.1%
EpLa), EpLa2 (basal diet added with 0.2% EpLa), EpLa3 (basal diet added with 0.3% EpLa)

However, the present result was in contrast to the previous studies that dietary inclusion of herbal ingredients containing flavonoid can reduce serum cholesterol and triglycerides in broiler (Hu et al 2003; Attia et al 2014). Some other researchers (Kim et al 2009; Yan et al 2019; Ao et al 2011; An et al 2015) also reported that dietary addition of garlic lowered blood concentrations of cholesterol and triglyceride in broilers. The varying effect of feeding dietary onion on serum cholesterol is certainly possible to be due to the variation of onion species, extraction method, additional level, and combination of treatment.

Although serum cholesterol and triglyceride concentrations were unchange, dietary inclusion of EpLa increased serum SOD and decreased MDA levels ( p<0.05). The effect of Dayak onion extract combined with Lactobacillus acidophilus containing phenolics and flavonoids could increased enzyme activities may suggest that poultry had greater capacity in scavanging oxygen free radicals, resulting in lower MDA level. The results of the present investigation were the same to those previously reported (Rahman 2007; Goliomytis et al 2014; Iskender et al 2016).

SOD as an antioxidant enzyme is an important index of the antioxidant capacity of animal tissue (Jiang et al 2007). Previous studies indicated that herbal extracts, such as Echinacea purpurea-containing flavonoid increased plasma level of SOD in broilers (Lee et al 2012). MDA, as a by-product of lipid peroxidation, reflects the degree of body cell oxidation. The increased serum antioxidant enzyme by feeding dietary Dayak onion extract and probiotic Lactobacillus acidophilus could be due to the active compounds of phenolic and flavonoid constituents which function as a strong antioxidant. Zhou et al (2016) reported that several herbs showed greatly stimulatory promoting effects on Lactobacillus acidophilus. It seems like there was a positive correlation between the antioxidant activity of Dayak onion extract and probiotic Lactobacillus acidophilus that would exert stimulating effect on health condition of the host animal and supports the productivity.

Meat quality characteristics

Flavonoid content, antioxidant activity, and meat cholesterol mass were different (p<0.05) among the treatment groups (Table 3). The increasing level of EpLa inclusion enhanced total phenolic, flavonoids, and antioxidant activity of meat, while meat cholesterol mass decreased with the slightly the increased meat protein mass. The present study indicated that total phenolic, flavonoid, and antioxidant activity of meat increased with the increasing levels of dietary EpLa treatment. Onion has been known as a source of bioactive phenolic com­pounds, and these compounds have paid attention due to their antioxidant properties. Total phenolic concentration of Dayak onion extract in the present experiment was 16,284 ppm (16.28 g/kg), which was higher than that has been examined in the previous studies, such as Aditya et al (2017), Kavalcová et al (2014), and Bouba et al (2012) was 0.39, 0.397, and 0.62 g/kg, respectively. Thus, the higest level of feeding EpLa produced the best meat characteristics indicated by the significantly increased total phenolic, flavonoids, and antioxidant activity of meat.

Table 3. Effect of Dayak onion extract combined with Lactobacillus acidophilus on meat quality characteristics







p value

Meat protein mass, g/bird







Meat cholesterol mass, g/bird







Phenolic total, mg/100g







Flavonoid, mg/100g







Antioxidant activity, % discoloration







a-b Means within row followed by different superscript differ at p<0.05, EpLa0 (basal diet), EpLa1 (basal diet
added with 0.1% EpLa), EpLa2 (basal diet added with 0.2% EpLa), EpLa3 (basal diet added with 0.3% EpLa)

The increased total phenolic, flavonoids, and antioxidant activity of meat were closely related to the enhanced SOD and the educed MDA levels in blood serum. Bioactive phenolic compounds of onion extract due to their antioxidant properties was completed by the combination of Lactobacillus acidophillus in preventing and scavenging the possible presence of free radical attacting body tissue. The decreased meat cholesterol mass and the slighly increased meat protein mass with the increasing level of EpLa were the evident of synergistic effects of the two components, antioxidant properties and Lactobacillus acidophillus. The present results were in accordance to the report of Sohaib et al (2015) indicated that retardation of meat lipid oxidation in broiler was brought about by dietary antioxidant quercetin, other active substance of plant source. However, it is not fully known how and what antioxidative substances in onion would impact the anti-oxidative action. The above result proved that the dietary flavonoid from Dayak onion can be absorbed and deposited into broiler meat by dose dependent manner. In addition, flavonoid deposition in meat has contributed to the better meat quality with higher anti-oxidationt content. This result may be caused by the increase in intestinal absorption of antioxidant compounds as an inclusion effect of probiotics Lactobacillus acidophilus. It is known that some plant active components may get quickly absorbed after oral administration and metabolized (Lee et al 2004). The current study implied that antioxidant properties can induce meat quality improvement was also due to the induction of antioxidant enzyme activities. However, further study is needed to specifically clarify the mechanism.

Growth performances

Feeding Dayak onion (Eleutherine palmifolia) extract and Lactobacillus acidophilus mixtured (EpLa) slightly increased body weight gain (p=0.06), and significantly improved (p <0.05) feed conversion ratio (Table 4). The best BWG and FCR found in the present study was due to the addition of EpLa at 0.3%.

Tabel 4. Effect of Dayak onion extract combined with Lactobacillus acidophilus on growth performance







p value

BWG, g/bird







FI, g/bird

2517 1













Carcass percentage, %







a-b Means within row followed by different superscript differ at p<0.05, BWG (body weight gain),
FI (feed intake), FCR (feed conversion ratio),EpLa0 (basal diet), EpLa1 (basal diet added with 0.1%
EpLa), EpLa2 (basal diet added with 0.2% EpLa), EpLa3 (basal diet added with 0.3% EpLa)

The antioxidant properties and probiotic may mediated the improvement of nutrient digestibility via the enhance digestive juices secretion caused by the microbial growth control in the intestine. These results were in accordance to the report of Purbarani et al (2019) that villi growth, protein digestibility, BW, and FCR improved in broiler fed diet fortified with a combination of Lactobacillus sp. and dahlia inulin. Previous studies indicated the positive influence on BW, FI and FCR of broiler fed diet containing fresh onion compared to that given diet without any addition of either onion or antibiotics (Goodarzi et al 2013). In connection with the levels of antioxidant properties, dietary addition of herbal plant extract at 0.2% containing phenolic and flavonoid as much 5041 mg/100g and 7102 mg/100g, respectively, was able to increase and improve carcass quality (Park et al 2014). It is camparable to the present study that feeding EpLa at 0.3% containing 9459 ppm flavonoids and 16284 ppm phenolic derived from Dayak onion extract was also able to increase BWG and improved FCR. It was supported by Karadas et al (2014) that a combination of phytonutrient improved growth and feed efficiency of broilers. Similar result reported by Goodarzi and Nanekarani (2014) that the addition of onion extract to the drinking water increased body weight and decreased feed conversion. However, these findings were in contrast to the results of Daneshman et al (2015) that the combination of propolis (a phenolic-rich source plant) and probiotic did not significantly affect broiler chickens performance.

The synergistic activity of active compounds of the mixture of Dayak onion extract and Lactobacillus acidophilus at 3% (EpLa3) increased SOD and decreased MDA (Table 2) with the enhanced active compounds (total phenolic and flavonoid) and antioxidant activity in meat (Table 3). The positive relationship of those parameters provided an impact on the slightly increased meat protein mass, the decreased meat cholesterol mass, and the improvement of growth performance. Therefore, it can be assumed that the addition of a combination of Dayak onion extract and Lactobacillus acidophilus is possible to indirectly increase the efficiency of nutrients utilization, especially protein, for body tissue syntesis. The result is supported by the previous reports that Lactobacillus sp. has a positive influence on the intestinal health (Hassanpour et al 2013) as indicated by the increased LAB population with lower coliform counts (Cholis et al 2018; Wulandari et al 2018). Similarly, improvement of growth performance in relation to the better balance of intestinal bacteria (higher LAB and low coliform counts) was found in broiler fed diet with addition of plant extract as glucomannan derived from porang (Amorphophallus onchophyllus) tuber (Perdinan et al 2019). Condusive gut condition through the improvement of intestinal morphology, such as better villi growth, and supported by protein digestibility (Purbarani et al 2019) ensured the promoting effect on growth performance of chicken. It seems like there was a positive correlation between antioxidant activity derived from Dayak onion extract and probiotic Lactobacillus acidophilus that would exert their stimulating effect on health condition of the host animal and supports the productivity. Unfortunately, there is very limited report on the effects of Dayak onion on broiler performance. Therefore, further study is needed to clarify the positive effects of synergistic mechanism between active compounds of Dayak onion and Lactobacillus acidophilus.



This study was funded by the Doctoral Dissertation Research Grant of the Ministry of Research, Technology and Higher Education of Republic of Indonesian through Domestic Graduate Scholarship (BPP-DN) program on the fiscal year 2019 with contract No 258-42/UN7.P4.3/PP/2019.


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Received 4 August 2019; Accepted 16 August 2019; Published 1 September 2019

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