The Effects of Incremental Doses of Aflatoxin B1 on In Vitro Ruminal Nutrient Digestibility and Fermentation Profile of a Lactating Dairy Cow Diet in a Dual-Flow Continuous Culture System.

Aflatoxin B1 (AFB1) is a mycotoxin known to impair human and animal health. It is also believed to have a deleterious effect on ruminal nutrient digestibility under in vitro batch culture systems. The objective of this study was to evaluate the effects of increasing the dose of AFB1 on ruminal dry matter and nutrient digestibility, fermentation profile, and N flows using a dual-flow continuous culture system fed a diet formulated for lactating dairy cows. Eight fermenter vessels were used in a replicated 4 × 4 Latin square design with 10 d periods (7 d adaptation and 3 d sample collection). Treatments were randomly applied to fermenters on diet DM basis: (1) 0 µg of AFB1/kg of DM (Control); (2) 50 µg of AFB1/kg of DM (AF50); (3) 100 µg of AFB1/kg of DM (AF100); and (4) 150 µg of AFB1/kg of DM (AF150). Treatments did not affect nutrient digestibility, fermentation, and N flows. Aflatoxin B1 concentration in ruminal fluid increased with dose but decreased to undetectable levels after 4 h post-dosing. In conclusion, adding incremental doses of AFB1 did not affect ruminal fermentation, digestibility of nutrients, and N flows in a dual-flow continuous culture system fed diets formulated for lactating dairy cows.

Burden of anemia and intestinal parasites in farmers and family members and owned livestock in two geographic locations in Senegal before and during the rainy seasons.

Different environmental conditions can impact the burden of anemia and intestinal parasite infections in human and livestock populations. The objective of this study was to compare the prevalence of anemia and intestinal parasite infections in farmers, family members, and owned sheep in two geographic locations along the Senegal River in June (end of the dry season) and September (rainy season). In Diawara, the prevalence of anemia in humans was high in June (74%) and remained high in September (75%) (p = 0.91). The prevalence of intestinal parasite infections increased from 7% in June to 54% in September (p < 0.05). Anemia was associated with age (children) and sex (women) (p < 0.05); but not with a positive diagnosis of intestinal parasite infection (p = 0.73). In sheep, the prevalence of anemia increased from 43% in June to 73% in September (p < 0.05), and the prevalence of intestinal parasite infections increased from 75% in June to 100% in September (p < 0.05). A positive diagnosis of Haemonchus contortus was associated with anemia (p = 0.05) and loss of body weight (2.4 kg) (p = 0.08). In Mpal, similar anemia and parasite infection trends were observed in children and sheep. The persistent high prevalence of anemia, and the impact of the rainy season on the burden of intestinal parasite infections in farmers, family members, and owned sheep can justify a One Health approach, where Senegal's ministries of health and of agriculture share resources for implementation and evaluation of government program efforts to reduce anemia in children and women, as well as morbidity and mortality in owned livestock; particularly in remote areas where public health services and veterinary services are very limited.

Assessment of peri-urban livestock producers' willingness to pay for improved forages as cash crops.

Availability of quality feed is a major constraint for livestock production in Burkina Faso. Despite previous efforts to test improved forages at research stations to overcome the dry-season feed gap, little has been done to promote them as cash crops that can contribute to meeting the growing feed demand in the country. This study was undertaken to evaluate the willingness to pay (WTP) for improved forage by livestock producers in the peri-urban livestock production systems of Burkina Faso. A total of 202 livestock producers were interviewed using semi-structured questionnaires. The contingent valuation method and Tobit econometric model were used to analyze the survey data. Exactly 79% of the interviewed livestock producers were willing to pay for improved forages for their livestock. Key factors that significantly affect this decision were the price of cottonseed cakes used as supplemental feed (P = .001), farmers' knowledge about improved forage crops (P = .001), farmers' ethnicity (P = .05), and farmers' practice of daily grazing and transhumance (P = .01). The estimated WTP for improved forage as a cash crop was US$0.32 kg-1 for all livestock producers and $0.58 kg-1 for those who only expressed a positive WTP. The positive WTP for improved forages and factors affecting that decision suggest that producing improved forages is a viable alternative to expensive cottonseed cakes and the practice of transhumance to overcome the dry-season feed gap. Therefore, dissemination of improved forages is recommended to market-oriented crop farmers to meet the growing feed demand in Burkina Faso.

Aflatoxin M1 in milk does not contribute substantially to global liver cancer incidence.

BACKGROUND: For 60 y, it has been known that aflatoxin B1 (AFB1), a mycotoxin produced by Aspergillus fungi in certain food and feed crops, causes hepatocellular carcinoma (liver cancer; HCC) in humans. The annual global burden of AFB1-related HCC has been estimated. However, much less is known about the potential carcinogenic impact of a metabolite of AFB1 called aflatoxin M1 (AFM1), which is secreted in milk when dairy animals consume AFB1-contaminated feed. The cancer risk of AFM1 to humans from milk consumption has not yet been evaluated. OBJECTIVES: We sought to estimate the global risk of AFM1-related liver cancer through liquid milk consumption, accounting for possible synergies between AFM1 and chronic infection with hepatitis B virus (HBV) in increasing cancer risk. METHODS: We conducted a quantitative cancer risk assessment by analyzing extensive datasets of national population sizes, dairy consumption patterns, AFM1 concentrations in milk in 40 nations, and chronic HBV prevalence. Two separate cancer risk assessments were conducted: assuming a possible synergy between AFM1 and HBV in increasing cancer risk in a manner similar to that of AFB1 and HBV, and assuming no such synergy. RESULTS: If there is no synergy between AFM1 and HBV, AFM1 may contribute ∼0.001% of total annual HCC cases globally. If there is synergy between AFM1 and HBV infection, AFM1 may contribute ∼0.003% of all HCC cases worldwide. In each case, the total expected AFM1-attributable cancer cases are ∼13-32 worldwide. CONCLUSION: AFM1 exposure through liquid milk consumption does not substantially increase liver cancer risk in humans. Policymakers should consider this low risk against the nutritional benefits of milk consumption, particularly to children, in a current global situation of milk being discarded because of AFM1 concentrations exceeding regulatory standards.

Anemia and intestinal parasites in farmers and family members and sheep in two agro-ecological zones in Senegal.

The burden of anemia in Senegal is high, particularly in children and women in rural households. The main objectives of the study reported here were (i) to measure and compare the prevalence of anemia and intestinal parasitic infections in farmers and family members and sheep in two agro-ecological zones in Senegal and (ii) to examine the association between anemia and age or sex in farmers and family members. The study was conducted in Mpal (250 km from Dakar, the capital city) and Diawara (700 km from Dakar, a remote location near the Malian border). In humans, the prevalence of anemia was higher in Diawara (64/86 = 74%), compared to Mpal (13/29 = 45%) (p < 0.01). Using logistic regression, the odds of anemia were 20.3, 5.7, and 3.2 times higher in children 1-4 years old, children 5-12 years-old, and teenagers 13-19 years old, respectively, compared to adults 20-60 years old, after controlling for study site and sex (p < 0.05). In Diawara, the odds of anemia were 2.9 times higher in women, compared to men, after controlling for age (p = 0.06). The prevalence of intestinal parasites (Giardia sp.) was the same (7%) at both locations. In sheep, the prevalence of low packed cell volume (PCV) and low body condition was higher in Diawara (48/60 = 60% and 11/60 = 18%, respectively), compared to Mpal (23/46 = 50% and 0/46 = 0%, respectively) (p < 0.05). Clinical anemia was associated (p < 0.01) with low PCV and a positive diagnosis of H. contortus. Overall, the prevalence of anemia was higher in farmers and family members and owned sheep in Diawara. In addition, anemia was more common in children and women, an indication that intra-household food allocation may be regulated in favor of men and older age groups. The consequences of livestock affected with anemia and undernutrition can be significant. High morbidity and mortality in livestock can lead to low household income, inadequate household access to and individual consumption of animal source foods, and subsequent risk of anemia in children and women in rural households in Senegal.

Effect of Lactobacillus hilgardii, Lactobacillus buchneri, or their combination on the fermentation and nutritive value of sorghum silage and corn silage.

The objective of this study was to determine the effect of inoculation with Lactobacillus hilgardii with or without Lactobacillus buchneri on the fermentation, chemical composition, and aerobic stability of sorghum and corn silage after 2 ensiling durations. Sorghum forage was harvested at 27% dry matter (DM; experiment 1), and different corn hybrids were harvested at late (43.8% DM; experiment 2) or normal maturity (34% DM; experiment 3). All harvested forages were chopped and ensiled in quadruplicate in vacuum-sealed nylon-polyethylene bags (40 × 61 cm) for 30 and 90 d after treatment with (1) deionized water (uninoculated) or (2) L. buchneri (1.5 × 105 cfu/g of fresh weight; LB); (3) L. hilgardii (1.5 × 105 cfu/g of fresh weight; LH); or (4) L. buchneri and L. hilgardii (1.5 × 105 cfu/g of fresh weight of each inoculant). Data for each experiment were analyzed separately accounting for the 2 × 2 × 2 factorial treatment arrangement. Inoculating sorghum forage with LB or LH separately increased acetate and 1,2 propanediol concentration, tended to increase DM loss, reduced lactate concentration and the lactate-to-acetate ratio, and increased aerobic stability after 90 but not after 30 d of ensiling. Inoculating late-harvested corn silage with LB or LH separately increased and decreased DM loss, respectively, increased 1,2 propanediol concentration, reduced lactate-to-acetate ratio and yeast counts but did not affect aerobic stability. Inoculating normal-harvested corn silage with LH reduced DM loss and increased 1,2 propanediol concentration and yeast counts; LB reduced lactate concentration, lactate-to-acetate ratio, and total acids. Either inoculant alone increased aerobic stability after 30 or 90 d. The main benefit of combining LB with LH was prevention of increases in DM losses by LH or LB separately. No improvement in aerobic stability resulted from applying LH instead of LB separately or from combining them. Application of LB or LH separately improved aerobic stability of sorghum silage after 90 d and normal-harvested corn silage after 30 or 90 d but did not affect that of late-harvested corn silage.

Effects of exogenous α-amylases, glucoamylases, and proteases on ruminal in vitro dry matter and starch digestibility, gas production, and volatile fatty acids of mature dent corn grain.

Two separate experiments were carried out to evaluate the effects of incremental doses of 10 exogenous endo-acting α-amylase and exo-acting glucoamylase; 1LAT (bacterial α-amylase), 2AK, 3AC, 4Cs4, 5Trga, 6Afuga, 7Fvga, and 10Tg (fungal α-amylases, glucoamylases, and α-glucosidase), 8Star and 9Syn (fungal amylase-mixtures; experiment 1) and three exogenous proteases; 11P14L, 12P7L, and 13P30L (bacterial proteases; experiment 2) on in vitro dry matter digestibility (IVDMD) and in vitro starch digestibility (IVSD) of mature dent corn grain using a batch culture system. Incremental doses of the exogenous enzymes (0, 0.25, 0.50, 0.75, and 1.00 mg/g of dried substrate) were applied directly to the substrate (0.5 g of ground corn, 4 mm) in sextuplicate (experiment 1) or quadruplicate (experiment 2) within F57 filter bags, which were incubated at 39 °C in buffered rumen fluid for 7 h. Rumen fluid was collected 2-3 h after the morning feeding from three lactating dairy cows and pooled. Cows were consuming a midlactation total mixed ration (TMR; 1.60 Mcal/kg DM and 15.4%; net energy of lactation and crude protein, respectively). Three independent runs were carried out for each experiment. Data were analyzed as a randomized complete block design using run as the blocking factor. Dose was used as a fixed factor while run was considered a random factor. Linear, quadratic, and cubic orthogonal contrasts were also tested. In experiment 1, enzymes 2AK, 3AC, and 10Tg did not increase (P > 0.10) IVDMD and IVSD, whereas 0.25 mg of enzymes 1LAT, 5Trga, and 8Star increased (P < 0.01) IVDMD by 23%, 47%, and 62% and IVSD by 35%, 41%, and 58%, respectively, compared with the control. Enzymes 4Cs4, 6Afuga, 7Fvga, and 9Syn linearly increased IVDMD and IVSD (P < 0.01). Greatest increases in IVDMD (82.9%) and IVSD (85.9%) resulted with 1 mg of 6Afuga compared to control. In experiment 2, the lowest dose of exogenous proteases 11P14L and 12P7L increased (P < 0.01) IVDMD by 98% and 87% and IVSD by 57% and 64%, respectively, whereas the highest dose of 13P30L increased (P = 0.02) IVDMD by 44.8% and IVSD by 30%, relative to the control. In conclusion, IVSD and IVDMD were increased by one α-amylase, certain glucoamylases, and all proteases tested, with the glucoamylase 6Afuga in experiment 1 and the neutral protease 12P7L in experiment 2, increasing IVDMD and IVSD to the greater extents. Future in vivo studies are required to validate these findings before these enzyme additives can be recommended for improving the digestibility of mature dent corn grain.

Meta-analysis of effects of inoculation with Lactobacillus buchneri, with or without other bacteria, on silage fermentation, aerobic stability, and performance of dairy cows.

A meta-analysis of 158 peer-reviewed articles was conducted to examine effects of inoculation with Lactobacillus buchneri (LB)-based inoculants (LBB) that did or did not include homolactic or obligate heterolactic bacteria on silage fermentation and aerobic stability. A complementary meta-analysis of 12 articles examined LBB inoculation effects on dairy cow performance. Raw mean differences between inoculant and control treatment means weighted by inverse variance were compared with a hierarchical effects model that included robust variance estimation. Meta-regression and subgrouping analysis were used to identify effects of covariates including forage type, application rate (≤104, 105, 106, or ≥ 107 cfu/g as fed), bacteria type (LB vs. LB plus other bacteria), enzyme inclusion, ensiling duration, and silo type (laboratory or farm scale). Inoculation with LBB increased acetate (62%), 1, 2 propanediol (364%) and propionate (30%) concentration and aerobic stability (73.8%) and reduced lactate concentration (7.2%), yeast counts (7-fold) and mold counts (3-fold). Feeding inoculated silage did not affect milk yield, dry matter intake, and feed efficiency in lactating dairy cows. However, forage type, inoculant composition, and dose effects on silage quality measures were evident. Inoculation with LBB increased aerobic stability of all silages except tropical grasses. Adding obligate homolactic or facultative heterolactic bacteria to LB prevented the small increase in DM losses caused by LB alone. The 105 and 106 cfu/g rates were most effective at minimizing DM losses while aerobic stability was only increased with 105, 106, and ≥ 107 cfu/g rates. Inoculation with LBB increased acetate concentration, reduced yeast counts and improved aerobic stability but did not improve dairy cow performance.

Effects of feeding a live yeast on rumen fermentation and fiber degradability of tropical and subtropical forages.

BACKGROUND: The effect of live yeast Saccharomyces cerevisiae strain CNCM I-1077 (SC) on the ruminal degradability of different forages commonly found in dairy diets in South America was evaluated. We also assessed if SC supplementation interacts with forage group to affect ruminal fiber degradability. Four non-lactating rumen-cannulated Holstein cows were randomly assigned to two treatment sequences: Control-SC-Control or SC-Control-SC, in a switchback design, with three 30-day periods. Cows in the SC treatment were supplied with 1 × 1010 colony-forming units of yeast daily via rumen cannula. In situ degradability of dry matter (DM) and neutral detergent fiber (aNDF) was measured in 15 forages collected in South America. Forages were assigned to one of three groups: corn silages; tropical grasses (sugarcane silages and tropical grass silages); and temperate grasses and alfalfa (oat silages, ryegrass silages, alfalfa silage, and alfalfa hay). RESULTS: Cows supplemented with SC had higher (P = 0.05) counts of yeasts and lower (P = 0.03) concentration of lactate in rumen fluid. There was no interaction between forage group and yeast supplementation (P > 0.10) on in situ degradability. The SC increased DM (by 4.6%) and aNDF degradation (by 10.3%) at 24 h of incubation (P < 0.05). Metabolomics revealed that a chemical entity (C17 H29 N6 O3 , m/z 365.2284 [M + H]+ ) from the family of lipids and related molecules was suppressed in the rumen fluid of cows supplemented with SC. CONCLUSION: The SC supplementation improved DM and aNDF degradability regardless of the forage group. © 2021 Society of Chemical Industry.

Aflatoxin in Dairy Cows: Toxicity, Occurrence in Feedstuffs and Milk and Dietary Mitigation Strategies.

Aflatoxins are poisonous carcinogens produced by fungi, mainly Aspergillus flavus and Aspergillus parasiticus. Aflatoxins can contaminate a variety of livestock feeds and cause enormous economic losses, estimated at between US$52.1 and US$1.68 billion annually for the U.S. corn industry alone. In addition, aflatoxin can be transferred from the diet to the milk of cows as aflatoxin M1 (AFM1), posing a significant human health hazard. In dairy cows, sheep and goats, chronic exposure to dietary aflatoxin can reduce milk production, impair reproduction and liver function, compromise immune function, and increase susceptibility to diseases; hence, strategies to lower aflatoxin contamination of feeds and to prevent or reduce the transfer of the toxin to milk are required for safeguarding animal and human health and improving the safety of dairy products and profitability of the dairy industry. This article provides an overview of the toxicity of aflatoxin to ruminant livestock, its occurrence in livestock feeds, and the effectiveness of different strategies for preventing and mitigating aflatoxin contamination of feeds.

Improving adoption of technologies and interventions for increasing supply of quality livestock feed in low- and middle-income countries.

The global increase in the demand for and production of animal-source foods (four-to five-fold increase between 1960 and 2015), which has been mostly concentrated in low- and middle-income countries (LMIC), provides smallholder livestock producers with an opportunity for improving their livelihoods and food and nutrition security. However, across livestock production systems in many LMIC, limited supplies and high cost of good quality feed severely constrains exploitation of this opportunity. In many of such countries, feeds and feeding-related issues are often ranked as the primary constraint to livestock production and increased consumption of animal-source foods. Here we review the complex biophysical, socio-economic and technological challenges related to improving quality feed supply and the reasons for generally low adoption of apparently proven feed enhancement technologies. We describe also successful interventions and conclude by recommending strategies for improving quality feed supply in LMIC that account for and overcome the prevailing challenges.

MILK Symposium Introduction: Dairy production in developing countries.

In low- and middle-income countries (LMICs), dairy production is highly valued, and demand for milk is projected to continue to increase markedly over the next few decades. This presents a tremendous opportunity to improve the nutrition, health, incomes, and livelihoods of millions of people with the high-quality protein and bioavailable micronutrients in dairy products. However, low dairy consumption levels, due to low affordability, accessibility, and availability, still typify several LMICs. This is caused by inadequate feeding, management, and genetics; poor transport, cooling, and processing infrastructure; unconducive policy environments; and sociocultural and demographic factors. Strategies to address some of these factors were presented at the MILK Symposium hosted by the Feed the Future Innovation Lab for Livestock Systems during the 2019 American Dairy Science Association Annual Meeting. The papers presented are full manuscripts in this Special Issue of the Journal of Dairy Science. They address the importance of dairy products for human health, strategies to address feed, management, health, and food safety challenges in dairy production systems, and sustainability of dairy production in LMICs. They collectively show how strategic interventions can lead to marked improvements in dairy production in developing countries. These will ultimately contribute to meeting the growing global demand for milk and to achievement of the United Nations Sustainable Development Goals.

MILK Symposium review: Improving control of mastitis in dairy animals in Nepal.

Dairy animals are an important source of income, food, and nutritional security, and improvements in the productivity of dairy animals substantially improve the wellbeing of smallholder dairy farmers. As in other developing countries, dairy animals are key for rural livelihoods in Nepal but often suffer from mastitis-a production disease causing economic losses to farmers, challenges to the dairy processing industry, and possible health hazards to consumers. Studies show that the prevalence of subclinical mastitis in Africa and Asia typically exceeds 50%, threatening animal wellbeing, farmers, dairy processors, and consumers. We conducted a study in Nepal to develop a technology training package to control mastitis in dairy animals. Following identification of knowledge gaps, a technology package consisting of (1) developing good husbandry practices, implementing mastitis detection and control technologies; and (2) training technicians and farmers was implemented. A strategy was subsequently established to provide feedback to farmers in dairy cooperatives on the subclinical mastitis status of their cows. The package was applied in the mid-western region of Nepal. Six months after implementation, we observed a reduction in subclinical mastitis prevalence: from 55% (baseline) to 28% (endline; n = 432) in dairy cows and from 78% to 18% (n = 216) in buffalo. These positive study outcomes strongly suggest that the mastitis technology training package should be scaled across smallholder farmers within and beyond Nepal to control mastitis in dairy animals.

Effects of Wormwood (Artemisia montana) Essential Oils on Digestibility, Fermentation Indices, and Microbial Diversity in the Rumen.

This study investigated the effects of essential oil (EO) from three Korean wormwood (Artemisia Montana) plants on in vitro ruminal digestibility, fermentation, and microbial diversity. Dried (0.5 g) soybean meal (SBM) or bermudagrass hay (BGH) were incubated in buffered rumen fluid (40 mL) for 72 h with or without EO (5 mg/kg) from Ganghwa (GA), Injin (IN), or San (SA) wormwood (Experiment 1). Both SA and IN improved (p < 0.05) dry matter digestibility (DMD) of BGH, while GA reduced (p < 0.05) total short-chain fatty acid of BGH and SBM. Besides, SA increased (p < 0.05) numbers of Ruminococcus albus and Streptococcus bovis in SBM. Experiment 2 examined different doses (0, 0.1, 1, and 10 mg/kg) of SA, the most promising EO from Experiment 1. Applying SA at 10 mg/kg gave the highest DMD (L; p < 0.01) and neutral detergent fiber (Q; p < 0.05) digestibility for BGH. Applying SA at 1 mg/kg gave the highest R. albus population (Q; p < 0.05) in SBM. Therefore, SA was better than GA and IN at improving rumen fermentation, and the 0.1 to 1 and 10 mg/kg doses improved ruminal fermentation and in vitro digestibility of SBM and BGH, respectively.

Effects of Class IIa Bacteriocin-Producing Lactobacillus Species on Fermentation Quality and Aerobic Stability of Alfalfa Silage.

The effects of two strains of class IIa bacteriocin-producing lactic acid bacteria, Lactobacillus delbrueckii F17 and Lactobacillus plantarum (BNCC 336943), or a non-bacteriocin Lactobacillus plantarum MTD/1 (NCIMB 40027), on fermentation quality, microbial counts, and aerobic stability of alfalfa silage were investigated. Alfalfa was harvested at the initial flowering stage, wilted to a dry matter concentration of approximately 32%, and chopped to 1 to 2 cm length. Chopped samples were treated with nothing (control, CON), Lactobacillus delbrueckii F17 (F17), Lactobacillus plantarum (BNCC 336943) (LPB), or Lactobacillus plantarum MTD/1 (NCIMB 40027) (LPN), each at an application rate of 1 × 106 colony-forming units/g of fresh weight. Each treatment was ensiled in quadruplicate in vacuum-sealed polyethylene bags packed with 500 g of fresh alfalfa per bag and ensiled at ambient temperature (25 ± 2 °C) for 3, 7, 14, 30, and 60 days. The samples were then subjected to an aerobic stability test after 60 days of ensiling. Compared with the CON silage, the inoculants reduced the pH after 14 days of ensiling. After 60 days, pH was lowest in the LPB-treated silage, followed by the F17 and LPN-treated silages. Inoculation of F17 increased concentrations of lactic acid in silages fermented for 7, 14, 30, and 60 days relative to other treatments, except for the LPN-treated silages ensiled for 30 and 60 days, in which the lactic acid concentrations were similar to that of F17 silage. Application of F17 and LPB decreased the number of yeast and mold relative to CON and LPN-treated silages. Compared with the CON silage, inoculant-treated silages had greater aerobic stability, water-soluble carbohydrate, and crude protein concentrations, and lower neutral detergent fiber, amino acid nitrogen, and ammonia nitrogen concentrations. The LPB-treated silage had the greatest aerobic stability followed by the F17-treated silage. Both class IIa bacteriocin producing inoculants improved alfalfa silage fermentation quality, reduced the growth of yeasts and molds, and improved the aerobic stability of the ensiled forage to a greater extent than the proven LPN inoculant. However, higher crude protein concentration and lower ammonia nitrogen concentration were observed in LPN-treated silage relative to other treatments.

An expansin-like protein expands forage cell walls and synergistically increases hydrolysis, digestibility and fermentation of livestock feeds by fibrolytic enzymes.

Bacterial expansin-like proteins have synergistically increased cellulose hydrolysis by cellulolytic enzymes during the initial stages of biofuel production, but they have not been tested on livestock feeds. The objectives of this study were to: isolate and express an expansin-like protein (BsEXLX1), to verify its disruptive activity (expansion) on cotton fibers by immunodetection (Experiment 1), and to determine the effect of dose, pH and temperature for BsEXLX1 and cellulase to synergistically hydrolyze filter paper (FP) and carboxymethyl cellulose (CMC) under laboratory (Experiment 2) and simulated ruminal (Experiment 3) conditions. In addition, we determined the ability of BsEXLX1 to synergistically increase hydrolysis of corn and bermudagrass silages by an exogenous fibrolytic enzyme (EFE) (Experiment 4) and how different doses of BsEXLX1 and EFE affect the gas production (GP), in vitro digestibility and fermentation of a diet for dairy cows (Experiment 5). In Experiment 1, immunofluorescence-based examination of cotton microfiber treated without or with recombinant expansin-like protein expressed from Bacillus subtilis (BsEXLX1) increased the surface area by > 100% compared to the untreated control. In Experiment 2, adding BsEXLX1 (100 µg/g FP) to cellulase (0.0148 FPU) increased release of reducing sugars compared to cellulase alone by more than 40% (P < 0.01) at optimal pH (4.0) and temperature (50°C) after 24 h. In Experiment 3 and 4, adding BsEXLX1 to cellulase or EFE, synergistically increased release of reducing sugars from FP, corn and bermudagrass silages under simulated ruminal conditions (pH 6.0, 39°C). In Experiment 5, increasing the concentration of BsEXLX1 linearly increased (P < 0.01) GP from fermentation of a diet for dairy cows by up to 17.8%. Synergistic effects between BsEXLX1 and EFE increased in vitro NDF digestibility of the diet by 23.3% compared to the control. In vitro digestibility of hemicellulose and butyrate concentration were linearly increased by BsEXLX1 compared to the control. This study demonstrated that BsEXLX1 can improve the efficacy of cellulase and EFE at hydrolyzing pure substrates and dairy cow feeds, respectively.