The effect of fungal probiotics added to a high-grain diet on the gastrointestinal tract of sheep

Author contributions

Conceptualization: R. Fabino Neto, F. O. A. Pessoa, M. M. Godoy; E.S. Miyagi; Formal analysis: F. O. A. Pessoa, R. Fabino Neto; Funding acquisition: F. O. A. Pessoa; M.M. Godoy; Investigation: F.O.A. Pessoa; R. Fabino Neto, T. D. Silva, V. V. Santana Neto, D. K. S. Lima, R. J. M. Silva; Project administration: R. Fabino Neto, F. O. A. Pessoa, M. M. Goday, E. S. Miyagi; Resources: R. Fabino Neto, F. O. A. Pessoa; Supervision: F. O. A. Pessoa, R. Fabino Neto, E. S. Miyagi; Validation: F. O. A. Pessoa, E. S. Miyagi; Visualization: F.O.A. Pessoa, M.M.A. Brainer, R. Fabino Neto; Writing (original draft): F. O. A. Pessoa, M. M. A. Brainer, R. Fabino Neto; Writing (review and editing): F. O. A. Pessoa, M. M. A. Brainer, R. Fabino Neto.

Treatments	Grain corn	Ground corn	Base mix	Probiotics
Treatments	(%)			spore/animal/day^a a Concentration of 9.33 x 1011 spore per animal per day. WGC: whole grain corn; GGC: ground grain corn; TE: control; RZ: Rhizomucor spp.; AT: Aspergillus terreus; MX: mixture of the two fungi;
WGC-TE	85	-	15	No
GGC-TE	-	85	15	No
WGC-RZ	85	-	15	Rhizomucor spp.
GGC-RZ	-	85	15	Rhizomucor spp.
WGC-AT	85	-	15	Aspergillus terreus
GGC-AT	-	85	15	Aspergillus terreus
WGC-MX	85	-	15	Rhizomucor spp. + Aspergillus terreus
GGC-MX	-	85	15	Rhizomucor spp. + Aspergillus terreus

Components (g/kg)	Corn	Whole grain corn diet	Ground grain corn diet
Dry matter	925.98	890.95	889.20
Mineral matter	10.88	36.50	39.45
Organic matter	989.12	963.50	960.55
Neutral detergent fiber	323.18	250.24	183.63
Acid detergent fiber	242.73	31.57	38.10
Ether extract	11.00	27.00	23.50
Non-fibrous carbohydrate⁴4 Paulino PVRP, Oliveira TST, Gionbeli MPM, Gallo SBS. Dietas Sem Forragem para Terminação de Animais Ruminantes. Rev Científica Produção Anim [Internet]. 2013 Dec 31;15(2):161-72. Available from: http://www.bibliotekevirtual.org/index.php/2013-02-07-03-02-35/2013-02-07-03-03-11/352-rcpa/v15n02/2877-v15n02a06.html http://www.bibliotekevirtual.org/index.p...	572.59	562.94	620.23
Crude protein	82.35	123.33	124.09

Physicochemical factor	Whole					Ground					TOTAL		Total
Physicochemical factor	RZ	AT	TE	MX	-	RZ	AT	TE	MX	-	WGC	GGC	Total
Coloration
Blackish-brown - BB	2	0	1	2		1	1	4	3		5	9	14
Brown - BN	1	2	1	2		1	2	0	2		6	5	11
Greenish brown - GB	0	1	0	0		1	1	1	0		1	3	4
Grayish milky - GM	0	2	3	1		0	0	0	0		6	0	6
Milky brown - MB	2	0	0	0		2	1	0	0		2	3	5
Odor
Ammoniacal - AI	1	1	0	0		0	0	0	0		2	0	2
Acid - AC	0	0	0	0		2	0	0	0		0	2	2
Aromatic - AR	3	3	2	3		1	2	3	4		11	10	21
Penetrating acid - PA	0	1	0	0		0	0	0	0		1	0	1
Insipid to acid - IA	0	0	1	0		1	0	0	0		1	1	2
Odorless - OL	1	0	2	2		1	3	2	1		5	7	12
Viscosity
Aqueous - AQ	1	2	3	3		2	2	1	2		9	7	16
Aqueous to foamy - AF	2	1	0	1		1	0	1	0		4	2	6
Thick - TC	2	2	2	1		0	2	2	2		7	6	13
Thick to aqueous - AT	0	0	0	0		2	1	1	1		0	5	5
MBRP (min)	< 3	< 3	< 3	< 3		< 3	< 3	< 3	< 3		< 3	< 3	< 3

Variables	Processing		Probiotics				P-value			CV
Variables	Whole	Ground	RZ	AT	MX	TE	Proc	Prob	Proc:Prob	CV
N-NH₃, mg/dℓ	8.99	9.34	9.66	9.30	9.54	8.1473	0.786	0.808	0.554	33.32
pH	5.51^b	6.15^a	5.77	5.73	5.99	5.838	0.001	0.579	0.748	7.60

Variables	Whole				Ground
Variables	RZ	TA	MX	TE	RZ	TA	MX	TE
Coccus	+++	+++	+++	+++	+++	+++	+++	++
Diplococcus	+++	+++	+++	+++	+++	+++	+++	+++
Streptococcus	+	++	+	++	+	++	++	+
Staphylococci	-	+	+	+	+	+	+	+
Spirillum	++	++	++	++	++	+	++	++
Vibrion	+	++	++	+	++	+	+	+
Bacilli	+++	+++	+++	+++	+++	++	+++	+++
Yeast	++	+	+	++	++	++	++	+
Gram positive, %	82.8^a	69.7^a	72.5^a	80.1^a	86.4^a	70.1^a	80.2^a	76.5^a
Gram-negative, %	17.2^b	33.5^b	30.7^b	26.5^b	25.1^b	25.0^b	26.3^b	25.1^b
Total	100	100	100	100	100	100	100	100

Variable	Whole grain corn CFU/mL^-1				Mean
Variable	RZ	AT	MX	TE	Mean
Lac+ bacteria	2.6 x 10⁴	2.6 x 10⁴	6.3 x 10⁴	7.4 x 10⁴	3.8 x 10⁴
Lac- bacteria	1.9 x 10⁴	1.8 x 10⁴	6.9 x 10³	1.5 x 10⁴	1.4 x 10⁴
Filamentous fungi	1.4 x 10³	9.6 x 10²	1.3 x 10³	8.6 x 10²	1.1 x 10³
Yeasts	2.2 x 10⁴	7.8 x 10³	5.0 x 10³	4.8 x 10³	4.9 x 10³
Small protozoa	2.0 x 10⁷	0	9.0 x 10⁵	1.8 x 10⁵	5.27 x 10⁶
Medium protozoa	5.8 x 10⁵	0	2.7 x 10⁴	2.3 x 10³	1.45 x 10⁵
Large protozoa	0	0	0	2.6 x 10³	6.50 x 10²
	Ground grain corn CFU/mL^-1
Lac+ bacteria	7.6 x 10⁴	2.8 x 10⁴	6.1 x 10⁴	1.1 x 10⁵	6.87 x 10⁴
Lac- bacteria	1.8 x 10⁴	1.1 x 10⁴	1.7 x 10⁴	2.3 x 10⁴	1.72 x 10⁴
Filamentous fungi	2.8 x 10³	1.2 x 10³	1.5 x 10³	1.9 x 10³	1.85 x 10³
Yeasts	8.3 x 10³	2.8 x 10³	6.7 x 10³	1.7 x 10³	4.87 x 10³
Small protozoa	2.6 x 10⁴	0	8.7 x 10⁴	2.0 x 10⁵	7.82 x 10⁴
Medium protozoa	60	0	1.0 x 10⁴	2.2 x 10²	2.57 x 10³
Large protozoa	0	0	1.7 x 10⁴	0	4.25 x 10³

Treatment	Quantification CFU/mL	Positive cultures (%)
Treatment	Quantification CFU/mL	Total	Lac+	Lac-
Whole grain corn
Rhizomucor spp.	2.27 x 10⁵	17.59	57.92ª	42.07^b
Aspergillus terreus	2.22 x 10⁵	17.23	58.36ª	41.64^b
Mixture of fungi	3.52 x 10⁵	27.31	90.12ª	9.87^b
Control	4.89 x 10⁵	37.86	76.29ª	23.70^b
Total WGC	1.29 x 10 ⁶A	100	-	-
Ground grain corn
Rhizomucor spp.	4.72 x 10⁵	26.95	80.75ª	19.25^b
Aspergillus terreus	2.38 x 10⁵	13.61	58.26ª	41.74^b
Mix of fungi	3.91 x 10⁵	22.31	78.07ª	21.93^b
Control	6.50 x 10⁵	37.13	88.27ª	11.73^b
Total GGC	1.75 x 10 ⁶B	100	-	-

Bacteria	Total	Whole								Ground
	Total	RZ		AT		MX		TE		RZ		AT		MX		TE
	N^* * Refers to the sum of observations between the bacterial genera.	n	%	N	%	n	%	N	%	n	%	N	%	N	%	n	%
Alcaligenes	37	4	10.8	8	21.6	4	10.8	1	2.7	9	24.3	2	5.4	6	16.2	3	8.1
Escherichia coli	22	4	18.2	2	9.1	3	13.6	3	13.6	-	-	6	27.3	1	4.5	3	13.6
Klebsiella	11	1	9.1	2	18.2	1	9.1	-	-	2	18.2	2	18.2	2	18.2	1	9.1
Shigella	10	1	10.0	-	-	1	10.0	3	30.0	1	10.0	-	-	1	10.0	3	30.0
Enterobacter	6	1	16.7	1	16.7	1	16.7	-	-	-	-	-	-	-	-	3	50.0
Edwardsiella	2	-	-	-	-	-	-	-	-	2	100	-	-	-	-	-	-
Proteus	1	-	-	1	100	-	-	-	-	-	-	-	-	-	-	-	-
Salmonella	1	-	-	-	-	-	-	-	-	1	100	-	-	-	-	-	-
Total	91	11	12.1	14	15.4	10	11.0	7	7.7	15	16.5	10	11.0	10	11.0	13	14.3

	Treatments - CFU/mL-1										P-value
Protozoan	Processing		Whole grain corn				Ground grain corn				P-value
Protozoan	Whole	Ground	RZ	AT	MX	TE	RZ	AT	MX	TE	Proc	Prob
Entodinium	51.65	23.61	110.6	-	16	80	3	-	2.5	80	0.80	0.63
Charonina	8	-	24	-	2	6	-	-	-	-	0.74	0.84

Variables	Processing (µm)		Probiotic (µm)				P-value			CV
Variables	Whole	Ground	RZ	AT	MX	TE	Proc	Prob	Proc:Prob	CV
LPS	2093	2267	1920	2224	22880	2288	0.297	0.346	0.128	34.84
WPB	771^b	9774^a	1080^a	752^b	711^b	949^a	0.001^* * Variable followed by different lowercase letters in the row shows significant difference at a 5% probability by the Tukey test. LPS: height of the lamina propria-submucosa of the ruminal epithelium; WPB: width of the base of the rumen papillae; TMT: thickness of the muscular tunica of the ruminal epithelium; SVH: small intestine villus height; SCD: small intestine crypt depth; and LCD: large intestine crypt depth. Proc: processing; Prob: probiotic; Proc:Prob: processing:probiotic.	0.001^* * Variable followed by different lowercase letters in the row shows significant difference at a 5% probability by the Tukey test. LPS: height of the lamina propria-submucosa of the ruminal epithelium; WPB: width of the base of the rumen papillae; TMT: thickness of the muscular tunica of the ruminal epithelium; SVH: small intestine villus height; SCD: small intestine crypt depth; and LCD: large intestine crypt depth. Proc: processing; Prob: probiotic; Proc:Prob: processing:probiotic.	0.001	27.67
TMT	1530ª	1380^b	1596^a	1250^c	1403^b	1572^a	0.001^* * Variable followed by different lowercase letters in the row shows significant difference at a 5% probability by the Tukey test. LPS: height of the lamina propria-submucosa of the ruminal epithelium; WPB: width of the base of the rumen papillae; TMT: thickness of the muscular tunica of the ruminal epithelium; SVH: small intestine villus height; SCD: small intestine crypt depth; and LCD: large intestine crypt depth. Proc: processing; Prob: probiotic; Proc:Prob: processing:probiotic.	0.001^* * Variable followed by different lowercase letters in the row shows significant difference at a 5% probability by the Tukey test. LPS: height of the lamina propria-submucosa of the ruminal epithelium; WPB: width of the base of the rumen papillae; TMT: thickness of the muscular tunica of the ruminal epithelium; SVH: small intestine villus height; SCD: small intestine crypt depth; and LCD: large intestine crypt depth. Proc: processing; Prob: probiotic; Proc:Prob: processing:probiotic.	0.001	14.78
SVH	507ª	385^b	352^b	500^a	511^a	421^b	0.001^* * Variable followed by different lowercase letters in the row shows significant difference at a 5% probability by the Tukey test. LPS: height of the lamina propria-submucosa of the ruminal epithelium; WPB: width of the base of the rumen papillae; TMT: thickness of the muscular tunica of the ruminal epithelium; SVH: small intestine villus height; SCD: small intestine crypt depth; and LCD: large intestine crypt depth. Proc: processing; Prob: probiotic; Proc:Prob: processing:probiotic.	0.001^* * Variable followed by different lowercase letters in the row shows significant difference at a 5% probability by the Tukey test. LPS: height of the lamina propria-submucosa of the ruminal epithelium; WPB: width of the base of the rumen papillae; TMT: thickness of the muscular tunica of the ruminal epithelium; SVH: small intestine villus height; SCD: small intestine crypt depth; and LCD: large intestine crypt depth. Proc: processing; Prob: probiotic; Proc:Prob: processing:probiotic.	0.241	27.34
PCD	451ª	408^a	574^a	387^b	353^b	403^b	0.096	0.001^* * Variable followed by different lowercase letters in the row shows significant difference at a 5% probability by the Tukey test. LPS: height of the lamina propria-submucosa of the ruminal epithelium; WPB: width of the base of the rumen papillae; TMT: thickness of the muscular tunica of the ruminal epithelium; SVH: small intestine villus height; SCD: small intestine crypt depth; and LCD: large intestine crypt depth. Proc: processing; Prob: probiotic; Proc:Prob: processing:probiotic.	0.463	28.90
SCD	564^b	753^a	520^b	652^a	755^a	706^a	0.001^* * Variable followed by different lowercase letters in the row shows significant difference at a 5% probability by the Tukey test. LPS: height of the lamina propria-submucosa of the ruminal epithelium; WPB: width of the base of the rumen papillae; TMT: thickness of the muscular tunica of the ruminal epithelium; SVH: small intestine villus height; SCD: small intestine crypt depth; and LCD: large intestine crypt depth. Proc: processing; Prob: probiotic; Proc:Prob: processing:probiotic.	0.001^* * Variable followed by different lowercase letters in the row shows significant difference at a 5% probability by the Tukey test. LPS: height of the lamina propria-submucosa of the ruminal epithelium; WPB: width of the base of the rumen papillae; TMT: thickness of the muscular tunica of the ruminal epithelium; SVH: small intestine villus height; SCD: small intestine crypt depth; and LCD: large intestine crypt depth. Proc: processing; Prob: probiotic; Proc:Prob: processing:probiotic.	0.015	27.62

[1] *correspondent: ronaildofabino@discente.ufg.br

Brasil

Brasil

The effect of fungal probiotics added to a high-grain diet on the gastrointestinal tract of sheep

Abstract