Denitrifying bioreactor microbiome: Understanding pollution swapping and potential for improved performance.

Denitrifying woodchip bioreactors are a best management practice to reduce nitrate-nitrogen (NO3 -N) loading to surface waters from agricultural subsurface drainage. Their effectiveness has been proven in many studies, although variable results with respect to performance indicators have been observed. This paper serves the purpose of synthesizing the current state of the science in terms of the microbial community, its impact on the consistency of bioreactor performance, and its role in the production of potential harmful by-products including greenhouse gases, sulfate reduction, and methylmercury. Microbial processes other than denitrification have been observed in these bioreactor systems, including dissimilatory nitrate reduction to ammonia (DNRA) and anaerobic ammonium oxidation (anammox). Specific gene targets for denitrification, DNRA, anammox, and the production of harmful by-products are identified from bioreactor studies and other environmentally relevant systems for application in bioreactor studies. Lastly, cellulose depletion has been observed over time via increasing ligno-cellulose indices, therefore, the microbial metabolism of cellulose is an important function for bioreactor performance and management. Future work should draw from the knowledge of soil and wetland ecology to inform the study of bioreactor microbiomes.

Antimicrobial resistance in integrated agroecosystems: State of the science and future opportunities.

As the Journal of Environmental Quality (JEQ) celebrates 50 years of publication, the division of environmental microbiology is one of the newest additions to the journal. During this time, significant advances in understanding of the interconnected microbial community and impact of the microbiome on natural and designed environmental systems have occurred. In this review, we highlight the intractable challenge of antimicrobial resistance (AMR) on humans, animals, and the environment, with particular emphasis on the role of integrated agroecosystems and by highlighting contributions published in JEQ. From early studies of phenotypic resistance of indicator organisms in waters systems to current calls for integrating AMR assessment across "One Health," publications in JEQ have advanced our understanding of AMR. As we reflect on the state of the science, we emphasize future opportunities. First, integration of phenotypic and molecular tools for assessing environmental spread of AMR and human health risk continues to be an urgent research need for a one health approach to AMR. Second, monitoring AMR levels in manure is recommended to understand inputs and potential spread through agroecosystems. Third, baseline knowledge of AMR levels is important to realize the impact of manure inputs on water quality and public health risk; this can be achieved through background monitoring or identifying the source-related genes or organisms. And finally, conservation practices designed to meet nutrient reduction goals should be explored for AMR reduction potential.

Catchment-scale export of antibiotic resistance genes and bacteria from an agricultural watershed in central Iowa.

Antibiotics are administered to livestock in animal feeding operations (AFOs) for the control, prevention, and treatment of disease. Manure from antibiotic treated livestock contains unmetabolized antibiotics that provide selective pressure on bacteria, facilitating the expression of anti-microbial resistance (AMR). Manure application on row crops is an agronomic practice used by growers to meet crop nutrient needs; however, it can be a source of AMR to the soil and water environment. This study in central Iowa aims to directly compare AMR indicators in outlet runoff from two adjacent (221 to 229 ha) manured and non-manured catchments (manure comparison), and among three catchments (600 to 804 ha) with manure influence, no known manure application (control), and urban influences (mixed land use comparison). Monitored AMR indicators included antibiotic resistance genes (ARGs) ermB, ermF (macrolide), tetA, tetM, tetO, tetW (tetracycline), sul1, sul2 (sulfonamide), aadA2 (aminoglycoside), vgaA, and vgaB (pleuromutilin), and tylosin and tetracycline resistant enterococci bacteria. Results of the manure comparison showed significantly higher (p<0.05) tetracycline and tylosin resistant bacteria from the catchment with manure application in 2017, but no differences in 2018, possibly due to changes in antibiotic use resulting from the Veterinary Feed Directive. Moreover, the ARG analysis indicated a larger diversity of ARGs at the manure amended catchment. The mixed land use comparison showed the manure amended catchment had significantly higher (p<0.05) tetracycline resistant bacteria in 2017 and significantly higher tylosin resistant bacteria in 2017 and 2018 than the urban influenced catchment. The urban influenced catchment had significantly higher ermB concentrations in both sampling years, however the manure applied catchment runoff consisted of higher relative abundance of total ARGs. Additionally, both catchments showed higher AMR indicators compared to the control catchment. This study identifies four ARGs that might be specific to AMR as a result of agricultural sources (tetM, tetW, sul1, sul2) and optimal for use in watershed scale monitoring studies for tracking resistance in the environment.

Strategies for Building Computing Skills To Support Microbiome Analysis: a Five-Year Perspective from the EDAMAME Workshop.

Here, we report our educational approach and learner evaluations of the first 5 years of the Explorations in Data Analysis for Metagenomic Advances in Microbial Ecology (EDAMAME) workshop, held annually at Michigan State University's Kellogg Biological Station from 2014 to 2018. We hope this information will be useful for others who want to organize computing-intensive workshops and will encourage quantitative skill development among microbiologists.IMPORTANCE High-throughput sequencing and related statistical and bioinformatic analyses have become routine in microbiology in the past decade, but there are few formal training opportunities to develop these skills. A weeklong workshop can offer sufficient time for novices to become introduced to best computing practices and common workflows in sequence analysis. We report our experiences in executing such a workshop targeted to professional learners (graduate students, postdoctoral scientists, faculty, and research staff).

Temporal Dynamics of Bacterial Communities in Soil and Leachate Water After Swine Manure Application.

Application of swine manure to agricultural land allows recycling of plant nutrients, but excess nitrate, phosphorus and fecal bacteria impact surface and drainage water quality. While agronomic and water quality impacts are well studied, little is known about the impact of swine manure slurry on soil microbial communities. We applied swine manure to intact soil columns collected from plots maintained under chisel plow or no-till with corn and soybean rotation. Targeted 16S-rRNA gene sequencing was used to characterize and to identify shifts in bacterial communities in soil over 108 days after swine manure application. In addition, six simulated rainfalls were applied during this time. Drainage water from the columns and surface soil were sampled, and DNA was extracted and sequenced. Unique DNA sequences (OTU) associated with 12 orders of bacteria were responsible for the majority of OTUs stimulated by manure application. Proteobacteria were most prevalent, followed by Bacteroidetes, Firmicutes, Actinobacteria, and Spirochaetes. While the majority of the 12 orders decreased after day 59, relative abundances of genes associated with Rhizobiales and Actinomycetales in soil increased. Bacterial orders which were stimulated by manure application in soil had varied responses in drainage waters over the course of the experiment. We also identified a "manure-specific core" of five genera who comprised 13% of the manure community and were not significantly abundant in non-manured control soils. Of these five genera, Clostridium sensu stricto was the only genus which did not return to pre-manure relative abundance in soil by day 108. Our results show that enrichment responses after manure amendment could result from displacement of native soil bacteria by manure-borne bacteria during the application process or growth of native bacteria using manure-derived available nutrients.

Allelic Variation in Outer Membrane Protein A and Its Influence on Attachment of Escherichia coli to Corn Stover.

Understanding the genetic factors that govern microbe-sediment interactions in aquatic environments is important for water quality management and reduction of waterborne disease outbreaks. Although chemical properties of bacteria have been identified that contribute to initiation of attachment, the outer membrane proteins that contribute to these chemical properties still remain unclear. In this study we explored the attachment of 78 Escherichia coli environmental isolates to corn stover, a representative agricultural residue. Outer membrane proteome analysis led to the observation of amino acid variations, some of which had not been previously described, in outer membrane protein A (OmpA) at 10 distinct locations, including each of the four extracellular loops, three of the eight transmembrane segments, the proline-rich linker and the dimerization domain. Some of the polymorphisms within loops 1, 2, and 3 were found to significantly co-occur. Grouping of sequences according to the outer loop polymorphisms revealed five distinct patterns that each occur in at least 5% of our isolates. The two most common patterns, I and II, are encoded by 33.3 and 20.5% of these isolates and differ at each of the four loops. Statistically significant differences in attachment to corn stover were observed among isolates expressing different versions of OmpA and when different versions of OmpA were expressed in the same genetic background. Most notable was the increased corn stover attachment associated with a loop 3 sequence of SNFDGKN relative to the standard SNVYGKN sequence. These results provide further insight into the allelic variation of OmpA and implicate OmpA in contributing to attachment to corn stover.

The genome and developmental transcriptome of the strongylid nematode Haemonchus contortus.

BACKGROUND: The barber's pole worm, Haemonchus contortus, is one of the most economically important parasites of small ruminants worldwide. Although this parasite can be controlled using anthelmintic drugs, resistance against most drugs in common use has become a widespread problem. We provide a draft of the genome and the transcriptomes of all key developmental stages of H. contortus to support biological and biotechnological research areas of this and related parasites. RESULTS: The draft genome of H. contortus is 320 Mb in size and encodes 23,610 protein-coding genes. On a fundamental level, we elucidate transcriptional alterations taking place throughout the life cycle, characterize the parasite's gene silencing machinery, and explore molecules involved in development, reproduction, host-parasite interactions, immunity, and disease. The secretome of H. contortus is particularly rich in peptidases linked to blood-feeding activity and interactions with host tissues, and a diverse array of molecules is involved in complex immune responses. On an applied level, we predict drug targets and identify vaccine molecules. CONCLUSIONS: The draft genome and developmental transcriptome of H. contortus provide a major resource to the scientific community for a wide range of genomic, genetic, proteomic, metabolomic, evolutionary, biological, ecological, and epidemiological investigations, and a solid foundation for biotechnological outcomes, including new anthelmintics, vaccines and diagnostic tests. This first draft genome of any strongylid nematode paves the way for a rapid acceleration in our understanding of a wide range of socioeconomically important parasites of one of the largest nematode orders.