The International Panel on Climate Change (IPCC) estimated that in 2010 enteric methane (CH₄) from ruminant livestock [1.6 to 2.7 Gt of carbon dioxide equivalent (CO₂e) per year] accounted for 3% to 5% of the total global anthropogenic greenhouse gas (GHG) emissions by all sectors (49 Gt total; Smith et al., 2014). Gerber et al. (2013) estimated a slightly greater contribution of enteric CH₄ at 6% of global GHG emissions, representing 40% of all livestock emissions. Given the ambitious GHG reduction commitments made by many countries, mitigation of enteric CH₄ has become a subject of intense investigation by the global research community. Numerous mitigation approaches have been proposed, as summarized in recent reviews (e.g., Beauchemin et al., 2020; Ku-Vera et al., 2020; Arndt et al., 2022; Beauchemin et al., 2022). Some strategies include improving animal efficiency, diet reformulation (e.g., decreasing fiber concentration), lipid supplementation of diets, forage management to improve digestibility, improved grazing management, tannin-containing forages, seaweed (macroalgae), CH₄ inhibitors (e.g., 3-nitrooxypropanol), nitrates, essential oils, other secondary plant compounds, genetic selection of low-emitting animals, vaccines, and early-life microbiome programming.

Among the mitigation approaches, the use of seaweeds has gained wide-scale interest because seaweeds are “naturally” sourced and widely abundant, livestock has a long history of grazing seaweed along shorelines in coastal areas, and some seaweeds are highly effective CH₄ inhibitors (Abbott et al., 2020). Specifically, the red seaweed Asparagopsis spp. (e.g., A. taxiformis and A. armata) has been shown to consistently decrease CH₄ emissions from dairy and beef cattle production (Lean et al., 2021), with reductions of up to 98% reported in beef cattle fed a high-grain diet supplemented with A. taxiformis (Kinley et al., 2020). Asparagopsis spp. accumulates halogenated compounds (mainly bromoform and dibromochloromethane) that reduce the efficiency of methyltransferase during methanogenesis (Machado et al., 2016). A recent study examining the impact of A. taxiformis on the rumen microbiome found that it rapidly reduced the abundance of rumen methanogens and led to a total collapse of the methanogen community (O’Hara et al., 2022). However, bromoform-containing seaweeds are not yet widely available to farmers because of regulatory issues, high production costs, and uncertainty as to whether or not bromoform and iodine accumulate in milk and meat. There is also a need for more efficient production practices to lower the CO₂ emissions from growing, harvesting, processing, extracting, and transporting seaweeds on a large scale.

Given the current limitations to the wide-scale use of Asparagopsis spp., other red, brown, and green seaweeds have garnered interest for CH₄ mitigation because they contain specific carbohydrates, lipids, peptides, saponins, and phlorotannins that have the potential to inhibit methanogens, albeit to a lesser extent than bromoform-containing seaweeds (Vijn et al., 2020; Abbott et al., 2020). To date, most of the research on these alternative seaweeds for ruminants has been conducted in vitro. As summarized by Abbott et al. (2020), decreases in in vitro CH₄ production have been reported for Cladophora patentiramea (green seaweed), Cystoseira trinodis (brown seaweed), Dictyota bartayresii (brown seaweed), Gigartina spp. (red seaweed), Padina australis (brown seaweed), and Ulva spp. (green seaweed), but efficacy has not been confirmed in vivo. In fact, few in vivo studies have evaluated non-bromoform-containing seaweeds for their ability to inhibit CH₄ production. Lean et al. (2021) searched the literature prior to 2021 to conduct a meta-analysis and identified only one study (Antaya et al., 2019) that evaluated a non-bromoform-containing seaweed and CH₄ production in ruminants. The study by Antaya et al. (2019), which provided grazing dairy cows with Ascophyllum nodosum (kelp meal) at approximately 0.7% of dry matter (DM) intake (DMI), reported a 10.5% decrease in CH₄ production (g/day) in the first month of feeding, with no effects thereafter. However, CH₄ yield (g/kg DMI) was not affected during that study. A more recent meta-analysis by Sofyan et al. (2022) included an additional publication (Katwal et al., 2021) in which the feeding of Sargassum johnstonii, at 8% of dietary DM, had no effect on CH₄ yields of lactating cows (Katwal et al., 2021). Thus, there is a paucity of support for the use of non-bromoform-containing seaweeds for CH₄ mitigation, and further in vivo research is required.

The red seaweed Mazzaella japonica is native to Republic of Korea, Japan, and Russia, and it is a non-indigenous species in other areas, including the coastal waters of western Canada where it is licensed for beach harvest (Holden et al., 2018). Once harvested, the material is sun-dried and marketed primarily as a source of carrageenan, a thickening agent used in processed foods, cosmetics, and pharmaceuticals. It is also used by local farmers as a roughage source for beef cattle, which has prompted interest in its potential for CH₄ mitigation. The objective of this study was to evaluate the effects of including M. japonica in the diet of beef cattle on DMI, rumen fermentation, digestibility, nitrogen (N) utilization, and enteric CH₄ production. The hypothesis was that incorporating M. japonica into the diet of beef cattle would decrease CH₄ production without incurring any negative effects on DMI and digestibility.

There is tremendous global interest in the use of seaweed (macroalgae) as a means of mitigating enteric CH₄ emissions from ruminant livestock (Beauchemin et al., 2022). Most animal feeding studies have evaluated the red seaweed Asparagopsis spp. because it contains halogenated compounds that are highly effective at disrupting methanogenesis when fed in small quantities (≤ 5% of DM) to ruminant livestock (as reviewed by Lean et al., 2021). However, very few in vivo studies have examined the mitigation potential of other seaweed species, despite a number of in vitro studies showing reductions in CH₄ production for a range of red, brown, and green seaweeds (Abbott et al., 2020). The CH₄ mitigation effect of non-bromoform-containing seaweeds is thought to be related to the numerous secondary plant compounds they contain (Vijn et al., 2020; Abbott et al., 2020). The present study evaluated M. japonica because it is available in western Canada, and some farmers claim that feeding this seaweed to their beef cattle improves weight gain and decreases CH₄ emissions. These claims are based on anecdotal evidence because controlled studies are not available to confirm or refute these assertions. M. japonica was inadvertently introduced to the waters of coastal British Columbia in the early 1900s, and in the past decade, the government of British Columbia has implemented a licensed harvest with no apparent negative environmental impacts (British Columbia Government News, 2015).

Seaweeds have been used as animal feeds for decades, especially in countries in close proximity to the sea. However, when fed at high levels, many seaweeds are known to reduce animal growth due to a lack of palatability, low digestibility of complex carbohydrates, and high inorganic matter proportion (Abbott et al., 2020). Rather than using seaweed as a major source of energy in the diet, low inclusion rates (1% and 2%) of M. japonica were used in the current study to evaluate the effects on CH₄ production. These low inclusion levels had no adverse effects on the palatability or DMI of the heifers. In fact, the cattle receiving M. japonica had a slightly greater intake than that of the control cattle, which was not due to enhanced palatability given that the rates of TMR intake in the early hours post feeding were not different across the treatments. Rather, the heifers appeared to compensate for the slightly lower DM digestibility of the TMR supplemented with M. japonica by increasing DMI in the later hours post-feeding, such that intake of digestible DM was similar for all treatments (data not shown). The decreased digestibility of diets containing M. japonica was due to a less digestible inorganic fraction, as there were no treatment effects on the digestibility of the organic fraction.

As is the case with most seaweeds, M. japonica contains a relatively high inorganic fraction (ash, 28.7% DM), thus adding 1% or 2% to the TMR decreased the OM concentration of the TMR by about 1 percentage unit. M. japonica is relatively high in macrominerals: 0.68% calcium, 1.71% potassium, 0.92% magnesium, 4.09% sodium, 0.16% phosphorus, and 6.46% sulfur (Terry et al., 2022). Most ruminants require 0.18% to 0.24% DM of dietary sulfur (NASEM, 2016); thus, the high sulfur concentration of M. japonica could be a concern with inclusion rates of > 2%, especially in cases where sulfur concentration in water is high or when diets include other feed ingredients that contain high sulfur concentration (e.g., distillers’ grains). The iodine concentration in M. japonica is low, unlike that of many bromoform-containing seaweeds. Terry et al. (2022) reported that the iodine concentration of M. japonica (15.7 mg/kg DM) was substantially less than that of A. taxiformis (2,580 mg/kg DM). Elevated iodine concentration in bromoform-containing seaweeds can exceed the daily iodine intake recommendations for cattle (Roque et al., 2021) – a major limitation to the inclusion of A. taxiformis in ruminant diets.

The finding that supplementing a forage-based TMR with 1% or 2% M. japonica had no mitigating effect on methanogenesis, when CH₄ was expressed relative to DMI (i.e., CH₄ yield), is consistent with the lack of effects on propionate proportion, rumen pH, and protozoa numbers. It is well established that inhibiting methanogenesis shifts rumen fermentation from acetate to propionate (Ungerfeld et al., 2022), and a decrease in rumen pH lowers CH₄ production (Luna et al., 1998). Ciliate protozoa produce H₂, the main substrate for methanogenesis in the rumen, and, therefore, treatments that decrease protozoa numbers result in less CH₄ produced (Tapio et al., 2017). The observed 9.2% decrease in daily CH₄ production (g/day) observed for the 2% M. japonica treatment was attributed to the slightly lower DMI of these animals during the CH₄ measurements. This finding is important because numerous in vitro studies have shown a decrease in CH₄ production with various non- and low-bromoform-containing seaweeds (Abbott et al., 2020; Sofyan et al., 2022). However, in many cases, the decrease in in vitro CH₄ production can be explained by lower substrate fermentability and lower gas production. Our results emphasize the need to express CH₄ production on the basis of OM fermented when conducting in vitro studies, or as CH₄ yield for in vivo studies, rather than as the total amount of CH₄ produced. Similar to our finding, Antaya et al. (2019) reported a 10.5% decrease in CH₄ production (g/day) in grazing dairy cows receiving A. nodosum, with no differences when CH₄ was expressed as g/kg DMI. The lack of effect of M. japonica on CH₄ yield in the current study confirms a recent in vitro rumen simulation study by Terry et al. (2022) that reported that adding 2% M. japonica to a forage-based diet had no effect on CH₄, expressed on the basis of mL/day, mg/g DM disappeared, or mg/g DM incubated. Similarly, supplementing M. japonica at 2% in an in vitro RUSITEC system had no significant effects on the composition of bacterial and archaeal communities (O’Hara et al., 2022).

Compared with barley silage and straw, M. japonica is relatively high in CP concentration (20.7% DM). Owing to the relatively low CP concentration (8.3% ± 0.26) of the basal diet, adding 1% and 2% M. japonica increased the N intake of the cattle. However, fecal N excretion was also increased for heifers consuming M. japonica because the CP digestibility did not differ among the diets. A portion of dietary CP is degraded in the rumen by ruminal microorganisms to NH₃-N, amino acids, and peptides, with NH₃-N being the primary N source for ruminal microbial protein synthesis. The increase in NH₃-N concentration in the rumen fluid with increasing supplementation of M. japonica indicates that the CP in this seaweed was rumen degradable. However, the lack of treatment effects on microbial N flow indicates that the degradable N supply was already met by the basal diet (8.48 mmol NH₃-N/L). Schwab et al. (2005) suggested that bacteria require 5–11 mmol/L of ammonia-N for optimal microbial growth depending on fermentation conditions. Ammonia that is not incorporated into bacterial protein is absorbed, enters the portal vein, is transported to the liver, and is converted to urea for excretion in urine. The linear increase in urinary N excretion with increasing M. japonica is consistent with the increase in NH₃-N concentration in the rumen fluid. Therefore, despite the high CP concentration of M. japonica and the relatively low CP concentration of the basal diet, supplementing with M. japonica did not improve the N utilization of the cattle, probably because the N requirement of the animals was sufficiently met by the basal diet. The cattle used in the study were at physiological BW (mean BW 840 kg) and CP requirements were low (8% CP), and, therefore, the mean N retention (mean, 6%) was low compared with that expected for younger cattle (26%, Angelidis et al., 2019; 30%, Schuba et al., 2017). It should be emphasized that N retention, calculated by subtracting N losses in refusals, feces, and urine from N intake, is considered an overestimate as other potential losses are not accounted for (e.g., epithelial cells, hair, nails, and N gas from the rumen; NASEM, 2016).

Supplementing a forage-based diet with the red seaweed M. japonica at 1% or 2% of DM failed to lower the enteric CH₄ yield (g/kg DMI) of mature beef heifers. A 9.2% decrease in daily CH₄ production (g/day) of cattle fed 2% M. japonica was observed, but this decrease was attributed to the slightly lower intake of these animals during the CH₄ measurements. Diets containing M. japonica were slightly less digestible than the control diet because the seaweed replaced forage when incorporated into the basal diet and the M. japonica contained a relatively large inorganic fraction (28.7% of DM). The sun-dried M. japonica used in the study was highly palatable, and thus the slightly lower digestibility of diets containing seaweed was offset by the increased DMI of the cattle, such that intake of digestible DM was not different among treatments. The high CP concentration (20.7% of DM) of M. japonica increased the N intake of cattle. The CP was rumen degradable, resulting in increased NH₃-N concentrations in the rumen fluid with increasing supplementation of M. japonica. Thus, we conclude that M. japonica can be used in diets to help meet the CP requirements of cattle. However, its high inorganic fraction, particularly its high sulfur concentration, may limit inclusion rates. When comprising up to 2% of the diet, M. japonica cannot be recommended as a CH₄ inhibitor for beef cattle fed high-forage diets.

The raw data supporting the conclusions of this article will be made available by the authors, without undue reservation.

The experiment was conducted at the Beef Cattle Metabolism Facility of Agriculture and Agri-Food Canada, Lethbridge Research and Development Centre (Lethbridge, AB) in accordance with guidelines of the Canadian Council on Animal Care (2009) and was pre-approved (protocol #ACC1935) by the Institutional Animal Care and Use Committee.

ST conducted the statistical analyses, oversaw the study, and contributed to manuscript writing. TC conducted the respiration chamber component and reviewed the manuscript. RG contributed to the study design and reviewed the manuscript. DA contributed to the study design and reviewed the manuscript. KB procured the funding, conceptualized the study, prepared the manuscript draft, and reviewed the manuscript. All authors contributed to the article and approved the submitted version.