INTRODUCTION

Pork is considered to be one of the most complete foods to meet the protein needs of the population, as it is a stable source and has positive effects on human health, thus contributing to food security (1). In pig production, the incidence of environmental and management factors leads to constant situations of physiological disorder, unleashing an imbalance in its intestinal microbiota, which has a negative impact on health and productivity (2). An alternative to increase the productive yield in animals is the use of microbial additives such as probiotics (3); because they have the capacity to: a) improve health, as well as the digestive and absorbent processes in the gastrointestinal tract, b) increase resistance of the mucus layer to infections, c) modulate the immune system, d) resist adverse conditions and adapt to sudden changes in the intestinal environment (4). However, the availability and cost of these products in developing countries can limit their use and minimize profit for small and medium producers.

However, the use of microbial preparations in the daily intake could be an alternative to increase production yield in piglets (5). a) It participates in vitamin synthesis, mainly B complex (7), b) improves intestinal immunity by avoiding the action of certain pathogens, such as Rotavirus, Escherichia coli, Salmonella and others (8), c)reduces the plasma concentration of certain harmful metabolites such as ammonia and endotoxins (9). Considering all of the above, this study aimed to evaluate the impact of two microbial additives on production and zoometric behavior, incidence of diarrhea and post-weaning piglet mortality.

MATERIALS AND METHODS

Area of study. The experimental work was carried out at the pork production unit, Gahuijón Alto, Cantón Colta, Ecuador. The unit is located at 1° 53’ 12.248 “LS longitude 78° 43’ 22.454 “LW, 3 510 masl (meters above sea level), with annual rainfall between 500-1 000 mm, minimum temperature 3°C, maximum 14°C, average 10°C, annual relative humidity 80% and annual evapotranspiration 69.03.

Experimental Design and treatment. A completely randomized design with four repetitions per treatment was used, where each experimental unit was composed of 40 piglets. The treatments evaluated were: control (T1); microbial preparation A (T2) and microbial preparation B (T3).

Animals and basal diet. 120 Duroc x Landrace/Yorshire cross piglets were used, descendants of 12 sows (Landrace/Yorkshire) with 135.6±2 kg live weight (LW) and 235±3 d of age. Four primiparous sows were used per treatment and, after childbirth, the piglets were randomly distributed to form the experimental groups of 40 animals per treatment (20 females and 20 males). The mother pigs were housed in collective pens of 6 x 6.5 m and cement floor, with a density of 1.8 m2 per animal from 15 days before reproduction to 110 days of gestation. From there, until weaning, they were placed in the maternity ward. From weaning onwards, the piglets were regrouped (without altering the groups of origin) into groups of 20 piglets in collective pens of 4 x 4.50 m, cement floor. The feed used for all the animals was Bioalimentar ® (Ambato, Ecuador), which meets the nutritional requirements for pigs according to the animal category recommended by NRC, (10 ). It was offered to the mothers twice a day, at 7:00 am and 4:00 pm. and ad libitum to the piglets from 7 days of age until weaning and from then up to 42 days of age, they were fed at the same time as their mothers. Water was also supplied ad libitum in nipple-type drinkers.

Animal management system. Maternity was kept at 28°C for the first two weeks after delivery, then reduced by 1.5°C each week until weaning. The photoperiod was controlled with 12 hours of light and 12 hours of darkness. The litters in each treatment were placed distant from each other (with an intermediate quarter on both sides of the aisle) to avoid self-inoculation. The piglets were weaned at 33 days of age. Piglets from each study group received the appropriate veterinary care according to the Manual for the Management of Females and Newborns (11).

Microbial preparations. The strains used in the preparations were: Kluyveromyces fragilis (L-4 UCLV) from the Microorganism Bank of the Universidad Central “Marta Abreu” de Las Villas and four ATCC microorganisms (American Type Cultures Collection, USA): Lactobacillus acidophilus, Lactobacillus bulgariccus, Streptococcus thermophilus and Saccharomyces cerevisiae. These were activated in skim milk at 37ºC for 24 hours. To obtain the microbial preparations, a mixture of sugar cane molasses and orange vinasse was used as a substrate and fermented at 37ºC for 24 hours, according to the technique described by Miranda et al (3). In preparation A, the bacteria L. acidophilus, L. bulgariccus and S. thermophilus were used, and in preparation B, the above bacteria plus yeasts S. cerevisiae and K. fragilis (L-4 UCLV) were used. The chemical composition and microbial concentration of each microbial preparation are shown in Table 1.

Supply of microbial additives to animals: The microbial preparations were supplied at 7:00 am every three days to the sows in groups T2 and T3. 15 mL of biopreparation was provided by mixing in 0.3 kg of dietary mixture plus 500 mL of water to the breeding sows from one day after gestation confirmation until weaning, according to the treatment assigned for each group and their offspring continued to receive the same additive. The first dose supplied to the piglets was 1 mL in single doses before taking colostrum. Dosage in oral form varied according to age: 1 mL in the first week; 1.5 mL in weeks two and three; 2 mL in weeks four and five and 2.5 mL in the following weeks until 42 days of age. The control group received initially physiological serum in the same quantity as the treated groups. Piglets over 7 days of age received the additive inoculated in 25 g of diet mixture.

Response variables. In order to evaluate the influence of microbial additives on piglets, the response variables determined were the productive indicators (live weight and average daily gain) and size of the animals at 0, 7, 14, 21, 28, 35 and 42 days of age. The incidence of diarrhea and mortality rates were also determined from weaning to 42 d. The experimental procedure for determining the indicators is presented below.

Production indicators. For the weighing of the piglets, a 20 kg capacity Roman manual scale (URKO M95460, Switzerland) was used, previously calibrated with a margin of error ±0.25 g. With this information, live weight (LW) and average daily gain (GMD) by age and sex (male and female) were calculated.

Size measurement (cm). Using a measuring tape (SUNBOO, China), the zoometric parameters were determined: height at withers (AZ), height at croup (AP) and body length (DL), according to the methodology described by Castro et al (12). With this information, the size of the animals at different ages was evaluated.

Diarrhea and mortality control. All the animals under study underwent rigorous clinical monitoring to detect behavioral changes, diarrheal disorders and deaths, and each animal was tested independently because they had been individually identified. With this information, the daily incidence and percentage of mortality were determined.

Statistical analysis. The experimental data were processed with the statistical package Statgraphic plus 15.1 for Windows. Analysis of variance by completely randomized design was performed and where necessary, Duncan’s docile comparison (13) was applied to discriminate between mean differences at p<0.05.

RESULTS

Production indicators of piglets at different ages. Table 2 shows the behavior of live weight and average daily gain in the different stages of the animal’s life. The indicator values, for all ages, were higher in the groups treated compared to the control group. The offspring (males and females) from the mothers who consumed the microbial additives were born with a higher weight and were larger at T3. In this same treatment, the highest PV values of the animals were found at 14, 35 and 42 d of age. This was also observed for males at 7, 21 and 28 days, while for females no differences between T2 and T3 were detected.

For ADG, the largest increases were found at 7, 14 and 35 d of age in the piglets (both sexes) in the T3 group. At 21, 28 and 42 days, no differences between T2 and T3 were detected for females.

Zoometric values of piglets at different ages. Tables 3 and 4 show the behavior of the size of male and female piglets from birth to 42 days of age in the different stages of the animal’s life. The offspring (males and females) of the mothers who consumed the T2 and T3 microbial additives were born with higher zoometric indicators, except for DL, females were larger in T3.

The values for the AZ indicator, for all ages 7 and 14, were higher in the treated groups compared to the control groups. In the measurement made at 21 and 42 d of age, a variation between groups was evident, being greater (p<0.05) in T3 compared to the others; it was greater (p<0.05) in T3 compared to T1 and T2, at 28 and 35 d of age in male piglets, while in females it was less (p<0.05) in T1 compared to T2 and T3, with no difference between the latter.

Concerning the AP values, at 7 d of age it was lower (p<0.05) in T1 compared to T2 and T3, with no differences between the latter. The female offspring treated with the microbial preparations were found to have greater values at 14, 28 and 35 d of age. This effect was also observed for males at 21 and 42 days, with no difference between T2 and T3.

For DL, the highest values were found at 14, 21, 35 and 42 d of age for the piglets (both sexes) in the T3 group. At 7 and 28 days after birth, there was no difference between T2 and T3 for males (Table 3 and 4).

Diarrhea rates and percentage of post-weaning piglet mortality. Table 5 shows the incidence of diarrhea and the percentage of mortality from weaning (33 days) to 42 days of age. At baseline, the incidence of diarrhea was lower (p<0.05) in T3 (0.11) than in T2 and T3, while the mortality rate did not vary between treatments. From day 34 to 40, the incidence of diarrhea was higher (p<0.05) in the control group compared to treated animals (T2 and T3). When comparing the piglets that consumed the microbial additives, it was lower (p<0.05) in Q3 than in Q2. With respect to the percentage of mortality, on days 34 to 37, it was higher (p<0.05) in T1 with respect to T2 and T3, without differing between those treated with microbial preparations. On days 41 and 42, there were no diarrheal disorders or deaths from diarrhea in any of the treatments studied.

DISCUSSION

The results in terms of production parameters (Table 2) were given possibly due to the use of microbial strains, simultaneously, this could be related to orange vinasse and sugar cane molasses (2). Increased birth weight and weight gain at different stages may be due to improved digestion and nutrient absorption, which may have manifested in productive behavior (14). Rondón et al (15) supplemented Lactobacillus salivarius C-65 in lactating pigs with increased weaning weight, which corroborates the hypothesis that the use of mixed cultures (lactic bacteria and yeasts) positively influences live weight and average daily gain. Similar results were also reported by Zhao et al (5) when supplementing microbial preparations to young pigs. Patil et al (6) observed improved expression of growth potential and reduced pathogen burden in the gastrointestinal tract by supplementing probiotics in the diet of newborn piglets. This could corroborate that microbial preparations are directly related to the factors that influence weight gain and average daily gain in the first few days of life (8).

The values of the zoometric parameters obtained in piglets could be related to the microbial preparations used in the different production stages of the piglets (Table 4 and 5). The contribution of probiotics to animal health and productivity is attributed to the micro-organisms used, and they may manifest themselves in the expression of morphometric characteristics in young pigs (16). On the other hand, Castro et al (12) preliminarily deduce that piglets with a larger withers height, croup height and body length have large, muscular hams when comparing commercial hybrid pigs with Ibero-American pigs. In this same sense, Sun et al (1) observed the changes in zoometric characteristics in relation to breed, handling and feeding, which leads us to believe that microbial preparations could have a positive effect on the values obtained. Ihara et al (7) reported the effect of probiotics in pigs independently of geographical area and variability within livestock production systems.

The decrease in diarrheal disorders and mortality in the piglets that consumed the microbial preparations (Table 5) were probably directly related to the microorganisms used, due to their capacity to adhere to the intestinal epithelium, colonize the lumen and exercise their function in the duodenum, was able to contribute to the reduction of diarrheal disorders caused by the stress of changing the diet. Simultaneously, the micro-organisms used helped regenerate atrophied microvilli and, as a consequence improved the health of the piglets at the critical stage of the animal (4, 8). In addition, Pajarrilla et al (9) reported beneficial effects of probiotic microorganisms, which include the ability to colonize the gastrointestinal tract, production of short-chain organic acids, production of secondary metabolites such as bacteriocins and thus preserve protective functions and inhibit cellular functions of E. coli and Salmonella spp., agents that promote diarrhea in young animals. In addition to this, Begum et al (17) also observed a reduction in digestive problems and mortality, improved digestion and nutrient absorption, as well as a notable improvement in the productive response, similar to the results obtained in this study (5,9).

In conclusion, the use of the microbial additives evaluated had a positive effect on the productive and zoometric behavior of the piglets. In addition, the incidence of diarrhea and animal mortality was reduced. Preparation B (a mixture of lactic bacteria and yeast) had a greater beneficial effect on piglets.

Acknowledgements

The main author thanks the Instituto de Fomento a Talento Humano (IFTH), Secretaria Nacional de Educación Superior Ciencia Tecnología e Innovación (SENESCYT) for the doctoral training grant (Ph.D.). Also, to the Faculty of Sciences, Superior Polytechnic School of Chimborazo (ESPOCH), Riobamba, (Ecuador) for enabling the use of the equipment and laboratories.

REFERENCES

1. Sun Y, Park I, Guo J, Weaver A, Woo S. Impacts of low level aflatoxin in feed and the use of modified yeast cell wall extract on growth and health of nursery pigs. J Anim Nutr. 2015; 1(3):177-183.

2. Datt C, Malik S., Datta M. Effect of probiotics supplementation on feed consumption, nutrient digestibility and growth performance in crossbred pigs under Tripura climate. Indian J Anim Nutr. 2011; 28(3):331-335.

3. Miranda JE, Marin A, Sánchez D, García Y. Obtención, caracterización y evaluación de dos preparados candidatos a probióticos desarrollados con residuos agroindustriales. Rev MVZ Córdoba. 2018; 23(1):6487-6499.

4. Haffner F, Diab R, Pasc A. Encapsulation of probiotics: insights into academic and industrial approaches. AIMS Materials Science. 2016; 3(1):114-136.

5. Zhao PY, Jung JH, Kim IH. Effect of mannan oligosaccharides and fructan on growth performance, nutrient digestibility, blood profile, and diarrhea score in weanling pigs. J Anim Sci. 2015; 90(3):833-839.

6. Patil AK, Kumar S, Verma AK, Baghel PS. Probiotics as feed additives in weaned pigs. LRI 2015; 3(2):31-39.

7. Ihara Y, Hyodo H, Sukegawa S, Murakami H, Morimatsu F. Isolation, characterization, and effect of administration in vivo, a novel probiotic strain from pig feces. Anim Sci J. 2013; 84(5):434-441.

8. Li J, Kim IH. Effects of Saccharomyces cerevisiae cell wall extract and poplar propolis ethanol extract supplementation on growth performance, digestibility, blood profile, fecal microbiota and fecal noxious gas emissions in growing pigs. Anim Sci J. 2014; 85(6):698–705.

9. Pajarillo E, Chae J, Balolong M, Kim H, Kang D. Assessment of fecal bacterial diversity among healthy piglets during the weaning transition. J Gen Appl Microbiol. 2014; 60(4):140‒146.

10. National Research Council (NRC). Nutrient Requirements of Swine: Eleventh Revised Edition. Washington, DC: The National Academies. 2012

11. Coates J, Corns PJ, Juarez A, MacDonald R, McCulley N, Melody B, Minton A, Molinari R, Montes De Oca H, Mosqueira P, Neill C, Pinilla JC, Piva J, Teuber R. Manual PIC de Manejo de Hembras y Primerizas. 100 Bluegrass Commons Blvd: Suite 2200: Hendersonville, EEUU; 2013.

12. Castro G, Montenegro M, Barlocco N, Vadell A, Gagliardi R, Llambí S. Caracterización zoométrica en el cerdo pampa rocha de Uruguay (descriptiva primaria). AICA. 2012; 2:83-86.

13. Duncan DB. Multiple range and multiple F tests. Biometrics 1955;11:1.

14. Ayala L, Bocourt R, Castro M, Milian G, Oliva D, Herrera M. Suministro de un cultivo de Bacillus subtilis a cerdas gestantes. Respuesta productiva en su descendencia. Rev Comput Prod Porc. 2012; 19(4):260-263.

15. Rondón A, Ojito Y, Arteaga F, Laurencio M, Milián G, Pérez Y. Efecto probiótico de Lactobacillus salivarius C 65 en indicadores productivos y de salud de cerdos lactantes. Rev Cub Cien Agr. 2013; 74(4):401-407.

16. Lähteinen T, Rinttilä T, Koort MK, Kant R, Levonen K, Jakava-Viljanen M, Björkroth J, Palva A. Effect of a multi species lactobacillus formulation as a feeding supplement on the performance and immune function of piglets. Livest Sci. 2015; 180:164–171.

17. Begum M, Li HL, Hossain MM, Kim IH. Dietary bromelain-C.3.4.22.32 supplementation improves performance and gut health in sows and piglets. Livest Sci. 2015; 180:177-182.