A study was undertaken to compare the rumen microbial community composition, methane yield, rumen fermentation, and CAZyme profiles between cattle and buffaloes. The primary aim of this study was to ascertain the impact of the host species on the above when diet and environmental factors are fixed. A total of 43 phyla, 200 orders, 458 families, and 1722 microbial genera were identified in the study. Bacteroidetes was the most prominent bacterial phylum and constituted >1/3rd of the ruminal microbiota; however, their abundances were comparable between cattle and buffaloes. Firmicutes were the second most abundant bacteria, found to be negatively correlated with the Bacteroidetes. The abundances of Firmicutes as well as the F/B ratio were not different between the two host species. In this study, archaea affiliated with the nine phyla were identified, with Euryarchaeota being the most prominent. Like bacterial phyla, the abundances of Euryarchaeota methanogens were also similar between the cattle and buffaloes. At the order level, Methanobacteriales dominated the archaea. Methanogens from the Methanosarcinales, Methanococcales, Methanomicrobiales, and Methanomassiliicoccales groups were also identified, but at a lower frequency. Methanobrevibacter was the most prevalent genus of methanogens, accounting for approximately three percent of the rumen metagenome. However, their distribution was not different between the two host species. CAZymes affiliated with five classes, namely CBM, CE, GH, GT, and PL, were identified in the metagenome, where the GH class was the most abundant and constituted ~70% of the total CAZymes. The protozoal numbers, including Entodiniomorphs and Holotrichs, were also comparable between the cattle and buffaloes. Results from the study did not reveal any significant difference in feed intake, nutrient digestibility, and rumen fermentation between cattle and buffaloes fed on the same diet. As methane yield due to the similar diet composition, feed ingredients, rumen fermentation, and microbiota composition did not vary, these results indicate that the microbiota community structure and methane emissions are under the direct influence of the diet and environment, and the host species may play only a minor role until the productivity does not vary. More studies are warranted to investigate the effect of different diets and environments on microbiota composition and methane yield. Further, the impact of variable productivity on both the cattle and buffaloes when the diet and environmental factors are fixed needs to be ascertained.

A series of in vitro studies were conducted to explore the anti-methanogenic potential of five seaweeds collected from the Indian sea and to optimize the level(s) of incorporation of the most promising seaweed(s) into a straw and concentrate diet to achieve a significant reduction in methane (CH₄) production without disturbing rumen fermentation characteristics. A chemical composition analysis revealed a notable ash content varying between 55 and 70% in seaweeds. The crude protein content was highly variable and ranged between 3.25 and 15.3% of dry matter. Seaweeds contained appreciable concentrations of tannins and saponins. Among the seaweeds, Spyridia filamentosa exhibited significantly higher CH₄ production, whereas the percentage of CH₄ in total gas was significantly lower in the cases of Kappaphycus alvarezii and Sargassum wightii. The ranking of seaweeds in terms of CH₄ production (mL/g OM) is as follows: Sargassum wightii < Kappaphycus alvarezii < Acanthophora specifera < Padina gymnospora < Spyridia filamentosa. A remarkable decrease of 31–42% in CH₄ production was recorded with the incremental inclusion of Kappaphycus alvarezii at levels of 3–5% of the dry matter in the diet. The addition of Sargassum wightii led to a significant decrease of 36–48% in CH₄ emissions when incorporated at levels of 4–5% of dry matter, respectively. The findings of this study revealed a significant decrease in the numbers of total protozoa and Entodinomorphs, coupled with increasing abundances of sulfate-reducing microbes and minor methanogens. Metagenome data revealed that irrespective of the seaweed and treatment, the predominant microbial phyla included Bacteroidota, Bacillota, Pseudomonadota, Actinomycetota, Fibrobacterota, and Euryarchaeota. The prevalence of Methanobrevibacter was similar across treatments, constituting the majority (~79%) of the archaeal community. The results also demonstrated that the supplementation of Kappaphycus alvarezii and Sargassum wightii did not alter the feed fermentation pattern, and therefore, the reduction in CH₄ production in the present study could not be attributed to it. Animal studies are warranted to validate the extent of reduction in CH₄ production and the key processes involved by supplementation with Kappaphycus alvarezii and Sargassum wightii at the recommended levels.

The transformation of oat brewery waste (OBW) into livestock feed could be a potential replacement for the expensive concentrate and one of the effective approaches for avoiding health hazards due to the accumulation of oat brewery waste in the environment. To explore the potential of OBW as a methane (CH₄) mitigating agent, an in vitro study was undertaken to investigate the effect of graded replacement of concentrate with OBW on CH₄ production, microbiota, feed fermentation, and CAZymes. A total of five treatments with variable proportions of OBW were formulated. The results indicated a linear decrease in the total gas production and a 38–52% decrease in CH₄ production with a 60 and 100% replacement of concentrate with OBW. The inclusion of OBW also affected the abundance of microbes such as Firmicutes, Euryarchaeota, Methanobrevibacter, and protozoa numbers. This study demonstrated that OBW can partially replace the concentrate and effectively mitigate CH₄ production; however, the concurrent decrease in fermentation cautioned for the partial replacement of concentrate with OBW at an appropriate level at which the fermentation remains unaffected while decreasing CH₄ production. Therefore, waste from oat breweries can contribute to curtailing the accumulation of greenhouse gases (GHGs) in the atmosphere.

A study was conducted on growing sheep to investigate the effect of two selected levels of biowaste of Padina gymnospora on feed intake, digestibility, daily enteric methane (CH₄) emission, growth performance, and rumen metagenome. We randomly divided the 18 growing male sheep into three groups of six animals each. The animals were fed on a basal diet comprising finger millet straw (Eleusine coracana) and a concentrate mixture in a 35:65 ratio. The sheep in the control group (C) were offered a concentrate mixture without waste, whereas the wheat bran in the concentrate mixture in test group I (A₂) and test group II (A₅) was replaced (w/w) with the biowaste of Padina gymnospora at a level of 3.07 and 7.69%, respectively. The biowaste of Padina gymnospora at the above levels in concentrate constituted 2 and 5% of the diet. A significant decrease of 28.4% in daily enteric CH₄ emission (g/d) was reported in the A₅ group, whereas the difference in daily enteric CH₄ emission between the C and A₂ & A₂ and A₅ groups did not prove significant. The inclusion of Padina gymnospora biowaste did not affect the nutrient intake and digestibility among the groups. The inclusion of Padina gymnospora biowaste in the A₅ group resulted in a significant reduction (p = 0.0012) in daily CH₄ emissions compared with group C; however, no significant differences were observed in daily CH₄ emissions between groups C–A₂ (p = 0.0793) and A₂–A₅ (p = 0.3269). Likewise, the adjustment of data to CH₄ emissions per 100 g of organic matter intake indicated a substantial decrease in the A₅ group relative to C. The energy loss in CH₄ as a percentage of GE relative to group C decreased significantly (−23.4%) in the A₅ group; however, this reduction was not associated with an increase in productivity, as almost similar average daily gain (p = 0.827) was observed in the groups. The replacement of wheat bran with the biowaste of Padina gymnospora significantly decreased the numbers of total protozoa and holotrichs in the A₅ group. Irrespective of the group, the Bacteroidota was the single largest phylum in the rumen metagenome, representing >60% of the microbiota. However, the abundance of Bacteroidota was similar among the groups. The methanogenic phyla Euryarchaeota was the 5th most abundant; however, it constituted only 3.14% of the metagenome. The abundance of Desulfovibrio was significantly higher in the A₅ group as compared with the control. In conclusion, the significant increase in the abundance of sulfate-reducing bacteria and reduction in protozoal numbers led to a significant reduction in CH₄ emissions with the incorporation of biowaste of Padina gymnospora at a 5% level of the diet.

In vitro studies were conducted in a series to investigate if the ruminal microbes are capable of degrading chlorpyrifos. This in vitro study presents the results from three experiments: Exp. I was conducted without feed, while Exp II and III were conducted with feed, either with or without methanol for dissolving chlorpyrifos, respectively. A basal diet comprising finger millet straw and concentrate was prepared. Incubation medium with feed but without chlorpyrifos served as the control. A total of six replicates each of control and chlorpyrifos spiked were used for the incubation. The pesticide concentration in the incubation medium before and after 24 h of incubation was analyzed using GC-MS/MS. The genomic DNA was isolated from the incubation fluid of the individual samples, and the shotgun metagenomic sequencing was performed. The clean reads were taxonomically classified using the Kraken2 database, and microbial classification at different taxonomic ranks was separated using Pavian v1.0. The microbial genes in the metagenome data were predicted and assigned functional roles using the MetaErg v1.2.3 pipeline. The assigned KEGG Orthology (KO), EC numbers (Enzyme Commission number), Gene Ontology (GO), and corresponding NCBI taxonomy information relevant to chlorpyrifos metabolism/degradation were retrieved. Results from the study revealed that the chlorpyrifos concentration was decreased from 5.78 to 1.64 ppm over 24 h of in vitro incubation with feed. Similar alpha and beta diversity indices between control and chlorpyrifos treatments revealed that the richness and the evenness of the microbial population were not affected by the presence of chlorpyrifos in the rumen fluid. There was no difference in the microbiota affiliated to the major phyla such as Bacteroidota, Fibrobacterota, Bacillota, and Pseudomonadota. The EC 3.1.8.1, EC 3.1.3.1, EC 1.14.13.-, and EC 1.1.1.- reported for chlorpyrifos degradation were not detected in the metagenome, and only EC 3.1.1.1 was identified, which demonstrated that degradation of chlorpyrifos was carried out by the affiliated enzyme carboxylesterase. The presence of GO:0004035, GO:0004364, GO:0019637, GO:0016791, and GO:0042178 in the metagenome strengthens that the chlorpyrifos degradation in the present study was primarily assigned to the rumen microbiota. This in vitro study provided insights into the rumen microbiota involved in the chlorpyrifos degradation and the initial clue that the rumen microbes are capable of degrading chlorpyrifos. Further, the animal studies in different species with the variable levels of chlorpyrifos are also warranted to confirm the efficacy of rumen microbes in mixed syntrophy and determine the threshold capabilities of the ruminal microbes.