New publication on rumen microbes which may have uses in biofuel production

A scientific article led by my colleague Dr. Alaa Rabee at the Desert Research Center in Egypt was just published online and is now available! Dr. Rabee and I have been collaborating remotely on projects related to the bacteria in the rumen of camels, sheep, and cows, as Dr. Rabee’s work focuses on the isolation of bacteria which can degrade plant materials efficiently and could be used to produce biofuels. He will be spending 6 months working in my lab as a visiting scholar, which was delayed until this year because of the pandemic.


Rabee, A.E., Sayed Alahl, A.A., Lamara, M., Ishaq, S.L. 2022. Fibrolytic rumen bacteria of camel and sheep and their applications in the bioconversion of barley straw to soluble sugars for biofuel production. PLoS ONE 17(1): e0262304. Article.

Abstract

Lignocellulosic biomass such as barley straw is a renewable and sustainable alternative to traditional feeds and could be used as bioenergy sources; however, low hydrolysis rate reduces the fermentation efficiency. Understanding the degradation and colonization of barley straw by rumen bacteria is the key step to improve the utilization of barley straw in animal feeding or biofuel production. This study evaluated the hydrolysis of barley straw as a result of the inoculation by rumen fluid of camel and sheep. Ground barley straw was incubated anaerobically with rumen inocula from three fistulated camels (FC) and three fistulated sheep (FR) for a period of 72 h. The source of rumen inoculum did not affect the disappearance of dry matter (DMD), neutral detergent fiber (NDFD). Group FR showed higher production of glucose, xylose, and gas; while higher ethanol production was associated with cellulosic hydrolysates obtained from FC group. The diversity and structure of bacterial communities attached to barley straw was investigated by Illumina Mi-Seq sequencing of V4-V5 region of 16S rRNA genes. The bacterial community was dominated by phylum Firmicutes and Bacteroidetes. The dominant genera were RC9_gut_group, RuminococcusSaccharofermentansButyrivibrioSucciniclasticumSelenomonas, and Streptococcus, indicating the important role of these genera in lignocellulose fermentation in the rumen. Group FR showed higher RC9_gut_group and group FC revealed higher RuminococcusSaccharofermentans, and Butyrivibrio. Higher enzymes activities (cellulase and xylanase) were associated with group FC. Thus, bacterial communities in camel and sheep have a great potential to improve the utilization lignocellulosic material in animal feeding and the production of biofuel and enzymes.

Paper published on animal feed and rumen bacteria

I’m delighted to announce that a paper was published on the effect of a dietary additive on the rumen and fecal bacterial communities in dairy cattle, in the journal Animal: The International Journal of Animal Biosciences!

A lot of factors can be manipulated to help get the most out of one’s diet, including the source and processing method of the ingredients – in most cases in livestock feed: plants. Growing plants for animal feed can be expensive, and often nutrients in plants become more available to the animal after the plant has been processed/broken down in some way. This sometimes allows for food byproducts to be reused for animal feed, and one common example is used brewers’ grains. Once the grains have been fermented to produce alcohol, the simple sugars have been used up but a lot of the complex sugar carbohydrates – in other words: fiber – are left over. Ruminants don’t need simple sugars, but they do need a lot of fiber, and brewers’ grains have been investigated for their usefulness for animal nutrition because they are a cheap, readily-available, and common source of fiber, as well as protein.

The original experiment for this work took place several years ago, and involved an animal feeding trial which added reduced-fat distillers’ grains with solubles into dairy cattle feed. The research team found no negatives effect on milk production or animal health, and that work was previously published. To add to that project, the original research team wanted to know if the diet would drastically change the bacterial community living in the rumen, which would have implications for feed digestion and animal health.

A collaborator of mine donated the cow microbial community DNA data to my AVS 590 special topics in DNA Sequencing Data Analysis course in spring 2020 (now formally registered as AVS 454/554). I worked with UMaine graduate students Adwoa Dankwa and Usha Humagain over the semester to train them in coding and develop the manuscript. The diet only had minimal effects on the bacterial community profiles, which in this case is a good finding – we want to be able to feed a cheap, nutritional source like distillers’ grains without harming the cow or its microbes.


Dankwa, A.S., U. Humagain, S.L. Ishaq, C.J. Yeoman, S. Clark , D.C. Beitz, and E. D. Testroet. 2021. Determination of the microbial community in the rumen and fecal matter of lactating dairy cows fed on reduced-fat dried distillers grains with solubles. Animal 15(7):100281.

Abstract

Reduced-fat dried distillers’ grains with solubles (RF-DDGS) is a co-product of ethanol production and contains less fat than traditional distillers’ grains. The fat in corn is ~ 91% unsaturated, and it is toxic to rumen microorganisms so it could influence the composition of the rumen microbiome. It has been demonstrated that RF-DDGS is a suitable ration ingredient to support the high-producing dairy cow, and this feedstuff is a promising alternative protein source for lactating dairy cows. The current study aims to better understand the effect of RF-DDGS on the rumen and fecal bacterial composition in lactating dairy cows. Thirty-six multiparous (2 or 3), mid-lactation Holstein cows (BW = 680 ± 11 kg; 106 ± 27 DIM) were randomly assigned to two groups which were fed a control diet made up of corn, corn silage, and alfalfa hay supplemented with expeller soybean meal or with added RF-DDGS (20% of the dry matter (DM)) containing approximately 6.0% fat. Whole rumen contents (rumen fluid and digesta; esophageal tubing method) and feces (free catch method) were collected on day 35 of the experimental period, after the 14-d acclimation period. Rumen contents and feces from each cow were used for DNA extraction. The bacterial community composition in rumen and fecal samples was assessed via the 16S rRNA gene by using the Illumina MiSeq sequencing platform. Bacteroidetes, Actinobacteria, and Firmicutes were the most abundant phyla in rumen contents. The fecal microbiota was dominated by the phyla Firmicutes and Bacteroidetes, as well as Actinobacteria and Chloroflexi. RF-DGGS increased bacterial richness, evenness, and Shannon diversity in both rumen and fecal samples and was associated with several taxa that had different abundance in treatment versus control comparisons.  The RF-DGGS, however, did not significantly alter the bacterial community in the rumen or feces. In general, these findings demonstrated that dietary inclusion of RF-DDGS did not impose any serious short-term (within 30 days) health or production consequences, as would be expected. With this study, we present further evidence that inclusion of 20% (DM basis) RF-DDGS in the diet of lactating dairy cows can be done without consequence on the microbiome of the rumen.

Implications

Reduced-fat dried distillers’ grains with solubles is a quality, economical, and readily available protein source demonstrated to support the protein needs of high-producing dairy cows. In this study, the rumen and fecal bacterial communities of lactating dairy cows were not significantly influenced by 20% (dry matter basis) reduced-fat dried distillers’ grains with solubles and did not impose serious short-term (within 30 days) health or production consequences. This diet could potentially be introduced into Total Mixed Ration feeding of dairy cattle given the fact that it is readily available and relatively economical.

Ciencia y aventura en mexico

For the past four days, I have been in Monterrey, Mexico, where I have been fostering international scientific relations in meetings and in the mountains.  It was my very first trip to Mexico, and it was an amazing experience.

After arriving in Monterrey, I was greeted by a research friend of mine, Dr. Jose Garcia-Mazcorro.  We met a few years ago when Jose emailed me to ask questions about a recently published paper on Saccharomyces probiotic treatment in cattle, and our conversations on the ecological theory behind probiotics led to a review paper on the subject, led by Jose and I. It wasn’t until last August when Jose and I actually met in person, at the ISME conference in Leipzig, Germany.

At dinner, I had the opportunity to meet Jose’s wife, Alecia, who is also a researcher, and their son, and discuss everything from aflatoxin to Stephen King’s “It” (we discovered that their son and I were both reading It when I discussed living in Maine, not far from Derry).

The next day, I woke up at 3:45 am to travel to the mountains for an incredible experience: a small-group tour in Matacanes canyon led by Daniel, a mountaineer with 20 years of experience and owner of Todo Avetura. He and Omar, another guide, led us for 12 hours and taught us about Matacanes canyon while we trekked 13 kilometers (8.7 miles) down waterfalls, through caves, over boulders, and over cliffs. Even the drive into and out of the canyon was an adventure; the steep road into the mountains fords rivers and winds along cliff faces.  In the canyon, I got to do many things for the first time, including rappel down the side of two waterfalls; 27 m (88.6 ft) and 15 m (49.2 ft) into a cave, swim through the absolute dark of a river cave system, and jump off of several cliffs into the water below, including a 9.5 meter (31 ft) jump!  It was supposed to be 10 meters, but that looked just a little too terrifying to try so I chose a spot that looked friendlier.  Turns out that jumping 9.5 meters is a lot like getting up to present in front of a large audience, you just have to get up there and do it before you have time to think about and psych yourself out.

The guides were really passionate about the mountains, and they were particular about safety and not rushing or pushing us to the point where we would get hurt.  If you have the chance, and the cohones, to go to Matacanes, I highly recommend Todo Aventura.

On Monday, sore but no worse for wear, I gingerly toured some of the facilities where Jose is currently working, MNA, an animal nutrition company.  I met with company president and nutritionist Dr. Jorge Kawas, and Jose, Jorge, and I discussed the role of microbes in animal nutrition and health. 

Dr. Jose Garcia-Mazcorro and I at MNA in Monterrey.

Monday night, I got to chat one-on-one with Professor T.J. Nagaraja, an author on the Saccharomyces review and a prominent researcher in rumen acidosis, cattle health, and infectious disease. 

The main reason for my trip to Monterrey was to attend and speak at the XXII UANL-Engorda de Bovinos en Corral Symposium.  I presented the opening seminar titled “Raising feedlot cattle with good microbes in mind” (“Cria y engorda de ganado con buenos microbios en mente”).  The video can be found here, and slides with presentation notes here:

Organized by MNA and UANL, the university in Monterrey, the symposium brings together researchers, producers, animal industry professionals, and students to discuss animal health in feedlot cattle.  I was honored to give the opening talk, which will be available online soon, and pleased to hear that the audience did in fact like microbes more than before my seminar!  Usually when I start talking about the gut microbiome people have the urge to run off and wash their hands…

Unfortunately, I had to jump back on a plane shortly after my talk, as I am heading to give a different presentation at the Wildlife Society meeting in Reno, Nevada tomorrow! But, I have plenty of memories and new project ideas to remember my trip by, and hopefully I will come back to Monterrey soon!

Accepted a tenure track faculty position at the University of Maine, Orono!

I’m thrilled to announce that this fall I’ll be starting as a tenure-track Assistant Professor in animal microbiome studies at the University of Maine, Orono, in the Department of Animal and Veterinary Sciences!!!

I’ll be returning to the field I started in; gut microbiology, but I’ll be integrating things I picked up along the way over the past few years, including ongoing collaborations in soil and built environment microbiology. I’ll be right at home in the UMaine School of Food and Agriculture, which brings together animal science, nutrition, and plant and soil science.

In addition to formally starting my own lab, I’m looking forward to snowy winters and summers on the rivers.

In fact, I’ll be right at home anywhere in Orono.

Cookies in the Featured Image are from Mug Buddy Cookies, made in Biddeford, Maine!!

A study on the effect of diet particle size got published!

I’m pleased to announce that the “particle size” project is officially published!  I inherited this dataset of bacterial 16S rRNA sequences in 2015, while working for the Yeoman Lab.  This collaborative project combined nutrition, animal production, and microbial ecology to look at the effect of diet particle size on lambs and their rumen bacteria. While small in size, the project was large in scope – despite everything we know about how different diet components encourage different microbial communities to survive in the digestive tract, we know practically nothing about how the size of the particles in that diet might contribute.

Ruminants, like sheep, goats, cows, deer, moose, etc.,  have a four-chambered stomach, the largest of which is called the rumen.  The rumen houses symbiotic microorganisms which break down plant fibers that the animal can’t digest on its own.  It’s estimated that up to 80% of a ruminant’s energy need is met from the volatile fatty acids (also called short-chain fatty acids) that bacteria produce from digesting fiber, and that up to 85% of a ruminant’s protein need is met from microbial proteins.

A lot of factors can be manipulated to help get the most out of one’s diet, including adjusting ingredients for water content, palatability, ease of chewing, and how easy the ingredients are to digest.  For example, highly fibrous foods with larger particles/pieces require more chewing, as well as a longer time spent in the rumen digesting so that microorganisms have plenty of time to break the chemical bonds of large molecules.  Smaller food particles can reduce the time and effort spent chewing, allow for more surface area on plant fibers for microorganisms to attach to and digest faster, and speed up the movement of food through the digestive tract.  On the other hand, moving food too quickly could reduce the amount of time microorganisms can spend digesting, or time the ruminant can absorb nutrients across their GI tract lumen, or cause slow-growing microbial species to wash out.


Pelleted-hay alfalfa feed increases sheep wether weight gain and rumen bacterial richness over loose-hay alfalfa feed.

Suzanne L. Ishaq1, Medora M. Lachman2, Benjamin A. Wenner3, Amy Baeza2, Molly Butler2, Emily Gates2, Sarah Olivo1, Julie Buono Geddes2, Patrick Hatfield2, Carl J. Yeoman2

  1. Biology and the Built Environment Center, University of Oregon, Eugene, Oregon, United States of America
  2. Department of Animal and Range Sciences, Montana State University, Bozeman, Montana, United States of America
  3. Department of Animal Sciences, The Ohio State University, Columbus, Ohio, United States of America

Abstract

Diet composed of smaller particles can improve feed intake, digestibility, and animal growth or health, but in ruminant species can reduce rumination and buffering – the loss of which may inhibit fermentation and digestibility.  However, the explicit effect of particle size on the rumen microbiota remains untested, despite their crucial role in digestion.  We evaluated the effects of reduced particle size on rumen microbiota by feeding long-stem (loose) alfalfa hay compared to a ground and pelleted version of the same alfalfa in yearling sheep wethers during a two-week experimental period.  In situ digestibility of the pelleted diet was greater at 48 h compared with loose hay; however, distribution of residual fecal particle sizes in sheep did not differ between the dietary treatments at any time point (day 7 or 14).  Both average daily gain and feed efficiency were greater for the wethers consuming the pelleted diet.  Observed bacterial richness was very low at the end of the adaptation period and increased over the course of the study, suggesting the rumen bacterial community was still in flux after two weeks of adaptation.  The pelleted-hay diet group had a greater increase in bacterial richness, including common fibrolytic rumen inhabitants. The pelleted diet was positively associated with several Succiniclasticum, a Prevotella, and uncultured taxa in the Ruminococcaceae and Rickenellaceae families and Bacteroidales order. Pelleting an alfalfa hay diet for sheep does shift the rumen microbiome, though the interplay of diet particle size, retention and gastrointestinal transit time, microbial fermentative and hydrolytic activity, and host growth or health is still largely unexplored.

A collaborative paper on zinc and rumen bacteria in sheep got published!

Zinc is an important mineral in your diet; it’s required by many of your enzymes and having too much or too little can cause health problems. We know quite a bit about how important zinc is to sheep, in particular for their growth, immune system, and fertility.  We also know that organically- versus inorganically-sourced zinc differs in its bio-availability, or how easy it is for cells to access and use it.  Surprisingly, we know nothing about how different zinc formulations might affect gut microbiota, despite the knowledge that microorganisms may also need zinc.

This collaborative study was led by Dr. Whit Stewart and his then-graduate student, Chad Page, while they were at Montana State University (they are now both at the University of Wyoming).   Chad’s work focused on how different sources of zinc affected sheep growth and performance (previously presented, publication forthcoming), and I put together this  companion paper examining the effects on rumen bacteria.

The pre-print is available now for Journal of Animal Science members, and the finished proof should be available soon. JAS is the main publication for the American Society of Animal Science, and one of the flagship journals in the field.


Zinc amino acid supplementation alters yearling ram rumen bacterial communities but zinc sulfate supplementation does not.

Ishaq, S.L., Page, C.M., Yeoman, C.J., Murphy, T.W., Van Emon, M.L., Stewart, W.C. 2018. Journal of Animal Science. Accepted. Article.

Abstract

Despite the body of research into Zn for human and animal health and productivity, very little work has been done to discern whether this benefit is exerted solely on the host organism, or whether there is some effect of dietary Zn upon the gastrointestinal microbiota, particularly in ruminants. We hypothesized that 1) supplementation with Zn would alter the rumen bacterial community in yearling rams, but that 2) supplementation with either inorganically-sourced ZnSO4, or a chelated Zn amino acid complex, which was more bioavailable, would affect the rumen bacterial community differently. Sixteen purebred Targhee yearling rams were utilized in an 84 d completely-randomized design, and allocated to one of three pelleted dietary treatments: control diet without fortified Zn (~1 x NRC), a diet fortified with a Zn amino acid complex (~2 x NRC), and a diet fortified with ZnSO4 (~2 x NRC). Rumen bacterial community was assessed using Illumina MiSeq of the V4-V6 region of the 16S rRNA gene. One hundred and eleven OTUs were found with > 1% abundance across all samples. The genera PrevotellaSolobacteriumRuminococcusButyrivibrioOlsenellaAtopobium, and the candidate genus Saccharimonas were abundant in all samples. Total rumen bacterial evenness and diversity in rams were reduced by supplementation with a Zn-amino-acid complex, but not in rams supplemented with an equal concentration of ZnSO4, likely due to differences in bioavailability between organic and inorganically-sourced supplement formulations. A number of bacterial genera were altered by Zn supplementation, but only the phylum Tenericutes was significantly reduced by ZnSO4 supplementation, suggesting that either Zn supplementation formulation could be utilized without causing a high-level shift in the rumen bacterial community which could have negative consequences for digestion and animal health.

Featured Image Source: Wikimedia Commons

The Hungate 1000 Project is now published!

The Hungate 1000 Project was a massive undertaking: namely, sequencing the genome of 1000 microorganisms cultured from ruminant animals all over the world, and was both  coordinated and led by the Rumen Microbial Genomics Network.  After years of hard work by some incredible researchers, the Hungate 1000 has just been published in the Nature Biotechnology Journal! The […]