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Pig production is one of the main sources of animal protein available to humanity, so that in the world there is a growing demand for this food source and other species of economic and nutritional interest (USDA 2018). Pigs have characteristics that differentiate them and make them preferential for many producers. Among them highlighted, the heterogeneity of the diet and its good conversion, adaptability and high prolificacy, as well as the carcass yield, made up of representative levels of proteins and lipids (Pexas et al. 2020).
It is common to observe in the pig sector that due to semi-intensive or intensive production, these animals have microbial dysbiosis and, in turn, gastrointestinal disorders, being this one the main cause of deaths in the farrowing area. Given these conditions, preventive measures have been applied to reduce this unwanted parameter, with the objective of improving the technological flow and optimizing the productive indices of farms (Tkacheva and Medvedev 2020). Antibiotics are used from the first days of life in the diets of monogastric animals, with the objective of increasing competitive exclusion in the microflora of the gastrointestinal tract (GIT), which controls enteric processes of a subclinical nature, frequent in intensive production. However, antibiotics can increase the number of resistant strains, as well as transfer cross-resistance to other microorganisms (Más et al. 2016).
Several researches have been directed to orally include zootechnical additives to improve food conversion rates and reduce diarrheal syndrome and mortality in the enterprise (Ojeda-García et al. 2016). Probiotics have been used in all pig categories, although with more emphasis on suckling and weaned pigs. They are considered one of the most viable alternatives to eliminate the use of medicated feed for pigs. Studies have confirmed that the use of beneficial microorganisms improves the digestibility of nutrients, intestinal integrity, antioxidant capacity and competitive exclusion of the gastrointestinal tract and therefore directly impacts the productive response of pigs (Ojeda-García et al. 2016).
Currently, the international scientific community has great interest in the study and applicability of efficient microorganisms (EM) in agricultural and livestock production. Although the results are not conclusive, these microorganisms stimulate the diversity of the intestinal flora, reduce inflammation of the intestine, improve the use of nutrients, decrease the excretion of N and improve the biological indicators of the animals (Kim and Baik 2019). However, little research has been done about its effect on suckling pigs, taking into account that it is the most susceptible pig category. That is why the objective of this research was to evaluate the oral effect of efficient microorganisms on bioproductive indicators, diarrheal syndrome and blood count in piglets.
Material and Methods
Experimental location. The research was carried out in the Enterprise Cría Pedregales, belonging to Empresa Porcina Granma, Cuba. This facility is located in the geographic quadrant 86-148-26. The average annual temperature is 25.7 ºC and the average relative humidity is 78 %. The purpose of this enterprise is to produce and commercialize pre-fattening at weaning.
Efficient microorganisms GERMEVIT. The efficient microorganisms GERMEVI were used, which come from the enterprise Patio Integral Agroecológico La Rosita, located in
Manzanillo municipality, Granma, Cuba. Worm humus, whey, molasses and rice powder were used to make GERMEVIT, and a liquid fermentation was developed. The chemical and microbiological characteristics of GERMEVIT were determined, according to the classic methods proposed by the AOAC (2006) and the Cuban Standard 585 (2017), respectively. This additive (EM) has 6.46 % of dry matter, 53.76 g / L of chemical oxygen demand (COD), 392 mg/L of Nt, 2100 mg/L of K, 290 mg/L of Na, 15.86 % of CF, 3.77 of pH, 9 x 105 of mesophilic bacteria and 2.33 x 101 of coliform organisms; 2.33 x 103 of proteolytics, 0.66 x 103 of total fungi, 45.66 x 107 of Lactobacillus spp. and absence of pathogenic bacteria.
Animals and treatments. From a total of 20 litters, two were selected at random, with 10 piglets each, one day old, from the Yorkshe-Yorkland x CC21 cross. The breeders had three furrowing and each suckling piglet constituted an experimental unit. Two experimental treatments were established: T1) basal diet (DB); T2) oral supply of 5 mL of EM on 7 consecutive days, from 24 h after born, and after ingestion of colostrum. The experiment lasted 21 days and the methodology proposed by Betancur et al. (2021) was used. To select the dose and supplying frequency, Zamora et al. (2020) results were considered.
Experimental conditions. The feeding of the suckling sows was restricted at a rate of 2 kg / animal/d and 450 g were added for each piglet (De Bettio et al. 2016).The food was supplied twice a day (8:00 a.m. and 3:00 p.m.) in 52 cm x 16 cm tubular feeders and water ad libitum in automatic nipple drinkers. After three days, the piglets were offered a pre-start diet in pellet form, at a rate of 10 g/animal/day, which was consecutively increased until reaching 300 g/animal/day in two frequencies (8:00 a.m. and 3:00 pm) (Betancur et al. 2021). Heating lamps were used to manage the temperature of the suckling piglets, in a range of 30 ºC (at the entrance) with progressive reduction to 26 ºC (at the exit). Table 1 shows the ingredients of the diets for suckling sows and suckled piglets.
Table 1 Ingredients of diets for suckling sows and suckled piglets
| Ingredients, % | Suckling | Pre-start |
|---|---|---|
| Corn meal | 63.00 | 36.00 |
| Soybean meal | 26.00 | 20.00 |
| Wheat bran | 4.00 | - |
| Soybean oil | 3.00 | 2.00 |
| Sugar | 2.00 | |
| Suckling1 nucleous | 4.00 | |
| Pre-start2 nucleous | 40.00 |
1Suckling nucleous per kilogram of product: folic acid 37.5 mg, pantothenic acid 300 mg, BHT 3.750 mg, biotin 5 mg, Ca 205000 mg, Co 6 mg, Cu 250, choline 10000 mg, Fe 2000 mg, P 51000 mg, Yo 25 mg, Mn 1250 mg, niacin 800 mg, Se 9 mg, Na 44000 mg, Zn 3125 mg; vitamins 250000 UI (B1 60 mg, B12 600 mcg, B2 150 mg, B6 80 mg, C 1250 mg, D3 50000 UI, E 1250 mg, K3 100 mg) Zn 3125 mg.
2Pre-start nucleous per kilogram of product: whey and powdered milk, skimmed milk powder, choline, extrude soybean, corn, sugar, fumaric acid, vegetable oil, dicalcium phosphate, threonine, tryptophan, calcium, mineral premixture, L-lysine, vitamin premixture, sodium chloride, DL-methionine, butyl-hydroxytoluene (BHT); vitamins 360.000 UI (D3 7.500 UI, E 450 mg, K3 18 mg, B1 12 mg, B2 29.5 mg, B6 13.5 mg, B12 0.01 mg, C 300 mg), niacin 118 mg, pantothenic acid 47.5 mg, folic acid 3.25 mg, biotin 0.75 mg, choline 1.800 mg, Fe 875 mg, Cu 625 mg, Mn 180 mg, Zn 625 mg, Co 3.25 mg
Productive indicators and diarrheal syndrome. The individual body weight of piglets was determined at birth, at 7, 14 and 21 d, according to Cheng et al. (2019) methodology using a digital balance with a precision of ± 0.1 g, scale (OSBORNE®, model 37473®, Kansas, Missouri, USA). With these data, the daily weekly weight gain was determined. The incidence of diarrhea (ID) in piglets was determined during the experimental period using the formula ID = number of diarrhea/ (number of animals × total days) × 100 (Liu et al. 2016).
Blood count. Five pigs, 21 d old, identified by an ear tag, were used for each treatment. After previously disinfecting the jugular area, 5 mL of blood samples were collected, whose containment was adequate. The extraction was carried out between 7:00 and 8:00 a.m., with 21 G x 1½ needles. To obtain blood plasma, the anticoagulant EDTA K2 was used, at a rate of 1 mL for every 2 mL of blood. Blood samples were determined using a Mindray hematology analyzer (Model BC-280. Vet. 2016, China). The values of red blood cells, white blood cells, hemoglobin, hematocrit and platelets were determined and compared with the reference values for this species and category.
Statistical analysis. A Student’s t test was performed for two independent samples and the results were expressed as mean and ± SE. The initial weight of pigs was processed through an analysis of covariance (P <0.05) to determine the effect on live weights at 7, 14 and 21 d of age. The data were processed using the statistical program SPSS 23.0.1.2014 (SPSS Inc., Chicago, IL, USA).
Results and Discussion
Table 2 shows the effect of oral supply of EM (GERMEVIT) on live weight and mean daily gain (MDG) of suckling pigs. Live weight at born of pigs showed significant differences between treatments (P <0.05). The covariance confirmed that the initial live weight did not significantly influence (P> 0.05) on the productive indicators analyzed in the following experimental weeks. However, the oral use of EM directly influence on live weight and MDG in suckling pigs during the periods of 7-14, 14-21 and 1-21 d of life (P <0.05). There was not morbidity and mortality processes (data not shown) in suckling pigs, due to the experimental treatments.
Table 2 Effect of oral supply of efficient microorganisms (GERMEVIT) on the productive performance of suckling pigs
| Items | Experimental treatments | SE ± | P value | Covariate P value | |
|---|---|---|---|---|---|
| Control | Efficient microorganisms | ||||
| Live weight (kg) | |||||
| at born | 2.08 | 1.58 | 0.07 | <0.001 | |
| 7 days | 3.35 | 2.99 | 0.05 | 0.014 | 0.072 |
| 14 days | 4.90 | 4.90 | 0.07 | 0.468 | 0.384 |
| 21 days | 6.41 | 6.61 | 0.08 | 0.095 | 0.241 |
| MDG (g/pig/d) | |||||
| 1-7 days | 181.34 | 201.70 | 5.70 | 0.080 | |
| 7-14 days | 193.09 | 240.17 | 9.54 | 0.013 | |
| 14-21days | 251.83 | 285.97 | 4.71 | <0.001 | |
| 1-21 days | 205.95 | 239.89 | 5.23 | <0.001 | |
One of the objectives of this study was to verify if the oral use of EM could have a positive effect on the two most important productive indicators in suckling pigs, especially for the susceptibility of these newborn animals, due to the recurrence of dysbiosis, as a consequence of the low colonization of lactic acid bacteria in the large intestine (Liu et al. 2016). These results show that EM (mainly Lactobacillus spp.) could colonize the GIT, improve intestinal health and therefore, weight gain in 24.38, 13.56 and 16.48 %, for the second and third week of life and throughout the experimental period, respectively.
Montejo-Sierra et al. (2017) reported that the dose of 3 mL/animal of EM stimulated the MDG from the second week of life with respect to what was registered in the control. Hou et al. (2016) and Michiels et al. (2016) stated that oral supplementation with Lactobacillus spp. in suckling pigs, improved nutrient absorption, competitive exclusion of bacteria in the colon and intestinal villi morphometry, which promoted MDG. Betancur et al. (2021) showed that oral probiotic use improved acid base status and weight gain in pigs up to 21 d after born. This study shows that the supply of EM after born, in early stages, could be the optimal time to manipulate the microbiota and improve the intestinal health and immunity of pigs, before and after weaning. The results confirm that the mineral-rich GERMEVIT with Lactobacillus spp. (45.66x107) and low pH (3.77), it is safe for oral use in suckling piglets. It should highlighted that there are still contradictions about the use of this zootechnical additive, with regard to the dose, the way of supplying, the preparation, the types of substrates and the productive categories (Montejo-Sierra et al. 2017 ).
As shown in figure 1, the oral use of efficient microorganisms reduced the incidence of diarrhea by 2.86 % with respect to control during the 21 days of the study.
Figure 1 Effect of oral supply of efficient microorganisms (GERMEVIT) on the incidence of diarrhea in suckling pigs (P <0.05).
The oral use of EM decreased the incidence of diarrheal syndrome, if it is taken into account that it is the main condition in suckling and weaned pigs (Cai et al. 2016), mainly caused by microbial dysbiosis. In this way, the oral use of EM can modulate the intestinal microflora, cause competitive microbial exclusion and, in turn, decrease the loss of electrolytes and water. Miranda et al. (2017) showed that oral supplementation with EM in piglets and fattening piglets reduced the presence of diarrhea. These authors recommended this practical strategy to reduce the proliferation of pathogens and the risk of intestinal infections. The presence of Lactobacillus spp. in the GERMEVIT it could also influence on the lower incidence of diarrhea in piglets, because these bacteria are common probiotics in the GIT, which play an important activity in defense against pathogens, acting as an intestinal barrier and immunity in pigs (Yang et al. 2017).
Song et al. (2010) showed that the use of allochthonous microorganisms (bacteria and yeasts) in pig production reduced the incidence of diarrhea and promoted bioproductive parameters. Zamora et al. (2020) reported that the oral use of EM and the probiotic VITAFER, with respect to the control diet without additives, markedly reduced the diarrheal syndrome. Rodríguez et al. (2013) showed that the oral use of a preparation with autochthonous microorganisms in pigs decreased the diarrheal syndrome, due to the reduction of the adhesion of pathogenic bacteria in the intestinal lumen. Valdés-Suárez et al. (2019) reported that EM supplementation caused significant changes in the caecal bacterial flora, with a substantial increase in the count of lactic acid bacteria and a reduction in mortality. According to the results obtained and in accordance with the information reported by various authors (Miranda et al. 2017 and Valdés Suárez et al. 2019), it is confirmed that the oral addition of EM can prevent and control diarrheal processes in pigs.
Table 3 shows that the oral administration of efficient microorganisms (GERMEVIT) did not affect the blood count, whose values are within the normal parameters for suckling pigs.
Table 3 Effect of oral supply of efficient microorganisms (GERMEVIT) on the blood count of suckling pigs
| Blood count | Experimental treatments | P value | Reference values* | ||
|---|---|---|---|---|---|
| Control | Efficient microoorganisms | SE± | |||
| White blood cells, x 109/L | 9.25 | 13.12 | 1.34 | 0.160 | 11-22 x 109/L |
| Red blood cells, x 1012/L | 6.00 | 6.06 | 0.08 | 0.745 | 5.0-9.5 x 1012/L |
| Hemoglobin, g/L | 110.00 | 109.30 | 1.39 | 0.813 | 99-165 g/L |
| Hematocrit, % | 38.43 | 39.25 | 0.53 | 0.474 | 32-50 % |
The hematological parameters are currently used as indicators of human and animal health (Iser et al. 2016). In general, it is necessary to determine if a new product, such as EM, can cause changes in these blood parameters. It has been reported that some beneficial microorganisms orally used can induce changes in leukocytes, mainly by activating the immune system to eliminate exogenous material and/or possible toxic and allergenic compounds (Betancur et al. 2021). The obtained results show that the use of EM in suckling pigs did not modify these parameters and they are consistent with previous studies (Tufarelli et al. 2017 and Rehan and Hotha 2019). However, Vera-Mejía et al. (2018), when using a probiotic strain in young pigs, found variable changes in eosinophils, lymphocytes and monocytes, justified by a better specific immune response, although, without notable changes for the red series and the values were within normal parameters.
Despite that in the control treatment there was a higher incidence of diarrhea, compared to the treatment that included the microbial additive (EM), this did not cause a significant value (P = 0.474) for the hematocrit (table 3). This hematological indicator (Ht) has been related to diarrheal syndrome because the loss of electrolytes and water, and subsequent dehydration causes a hemoconcentration of the blood and, in turn, an increase in hematocrit. Apparently, the diarrheal syndrome did not cause affectations that produced changes in this indicator and in the morbidity and mortality of pigs (unpublished data).
