A Feeding Trial to Investigate Strategies to Mitigate the Impacts of Pimelea Poisoning in Australian Cattle.

Pimelea poisoning of cattle causes distinct symptoms and frequently death, attributable to the toxin simplexin. Pimelea poisoning was induced via addition of ground Pimelea trichostachya plant to the daily feed in a three-month trial with Droughtmaster steers. The trial tested four potential mitigation treatments, namely, biochar, activated biochar, bentonite, and a bacterial inoculum, and incorporated negative and positive control groups. All treatments tested were unable to prevent the development of simplexin poisoning effects. However, steers consuming a bentonite adsorbent together with Pimelea showed lesser rates-of-decline for body weight (P < 0.05) and four hematological parameters (P < 0.02), compared to the positive control group fed Pimelea only. Microbiome analysis revealed that despite displaying poisoning symptoms, the rumen microbial populations of animals receiving Pimelea were very resilient, with dominant bacterial populations maintained over time. Unexpectedly, clinical edema developed in some animals up to 2 weeks after Pimelea dosing was ceased.

Binding of the plant-derived toxin simplexin to bovine protein kinase C: insights from molecular dynamics.

Pimelea poisoning of cattle is toxicologically linked to the activation of bovine protein kinase C (PKC) by the plant-derived toxin simplexin. To understand the affinity of PKC for simplexin, we performed molecular dynamics (MD) studies of simplexin, simplexin analogues, and several other activators of PKC. Binding enthalpy calculations indicated that simplexin had the strongest affinity for PKCα-C1B among the activators studied. Key to simplexin's affinity is its ability to form more hydrogen bonds to PKC, compared to the other activators. The C-3 carbonyl group and C-20 hydroxyl group of simplexin were identified as especially important for stabilizing the PKC binding interaction. The hydrophobic alkyl chain of simplexin induces deep membrane embedding of the PKC-simplexin complex, enhancing the protein-ligand hydrogen bonding. Our findings align with previous experiments on structure-activity relationships (SAR) for simplexin analogues, and provide insights that may guide the development of interventions or treatments for Pimelea poisoning.

Degradation of the Indospicine Toxin from Indigofera spicata by a Mixed Population of Rumen Bacteria.

The leguminous plant species, Indigofera linnaei and Indigofera spicata are distributed throughout the rangeland regions of Australia and the compound indospicine (L-2-amino-6-amidinohexanoic acid) found in these palatable forage plants acts as a hepatotoxin and can accumulate in the meat of ruminant livestock and wild camels. In this study, bovine rumen fluid was cultivated in an in vitro fermentation system provided with Indigofera spicata plant material and the ability of the resulting mixed microbial populations to degrade indospicine was determined using UPLC-MS/MS over a 14 day time period. The microbial populations of the fermentation system were determined using 16S rRNA gene amplicon sequencing and showed distinct, time-related changes occurring as the rumen-derived microbes adapted to the fermentation conditions and the nutritional substrates provided by the Indigofera plant material. Within eight days of commencement, indospicine was completely degraded by the microbes cultivated within the fermenter, forming the degradation products 2-aminopimelamic acid and 2-aminopimelic acid within a 24 h time period. The in vitro fermentation approach enabled the development of a specifically adapted, mixed microbial population which has the potential to be used as a rumen drench for reducing the toxic side-effects and toxin accumulation associated with ingestion of Indigofera plant material by grazing ruminant livestock.

Rumen Virus Populations: Technological Advances Enhancing Current Understanding.

The rumen contains a multi-kingdom, commensal microbiome, including protozoa, bacteria, archaea, fungi and viruses, which enables ruminant herbivores to ferment and utilize plant feedstuffs that would be otherwise indigestible. Within the rumen, virus populations are diverse and highly abundant, often out-numbering the microbial populations that they both predate on and co-exist with. To date the research effort devoted to understanding rumen-associated viral populations has been considerably less than that given to the other microbial populations, yet their contribution to maintaining microbial population balance, intra-ruminal microbial lysis, fiber breakdown, nutrient cycling and genetic transfer may be highly significant. This review follows the technological advances which have contributed to our current understanding of rumen viruses and drawing on knowledge from other environmental and animal-associated microbiomes, describes the known and potential roles and impacts viruses have on rumen function and speculates on the future directions of rumen viral research.

Metatranscriptomics reveals mycoviral populations in the ovine rumen.

The rumen is known to contain DNA-based viruses, although it is not known whether RNA-based viruses that infect fungi (mycoviruses) are also present. Analysis of publicly available rumen metatranscriptome sequence data from sheep rumen samples (n = 20) was used to assess whether RNA-based viruses exist within the ovine rumen. A total of 2466 unique RNA viral contigs were identified that had homology to nine viral families. The Partitiviridae was the most consistently observed mycoviral family. High variation in the abundance of each detected mycovirus suggests that rumen mycoviral populations vary greatly between individual sheep. Functional analysis of the genes within the assembled mycoviral contigs suggests that the mycoviruses detected had simple genomes, often only carrying the machinery required for replication. The fungal population of the ovine rumen was also assessed using metagenomics data from the same samples, and was consistently dominated by the phyla Ascomycota and Basidomycota. The strictly anaerobic phyla Neocallimastigomycota were also present in all samples but at a low abundance. This preliminary investigation has provided clear evidence that mycoviruses with RNA genomes exist in the rumen, with further in-depth studies now required to characterise this mycoviral community and determine its role in the rumen.

Toward Understanding Phage:Host Interactions in the Rumen; Complete Genome Sequences of Lytic Phages Infecting Rumen Bacteria.

The rumen is known to harbor dense populations of bacteriophages (phages) predicted to be capable of infecting a diverse range of rumen bacteria. While bacterial genome sequencing projects are revealing the presence of phages which can integrate their DNA into the genome of their host to form stable, lysogenic associations, little is known of the genetics of phages which utilize lytic replication. These phages infect and replicate within the host, culminating in host lysis, and the release of progeny phage particles. While lytic phages for rumen bacteria have been previously isolated, their genomes have remained largely uncharacterized. Here we report the first complete genome sequences of lytic phage isolates specifically infecting three genera of rumen bacteria: Bacteroides, Ruminococcus, and Streptococcus. All phages were classified within the viral order Caudovirales and include two phage morphotypes, representative of the Siphoviridae and Podoviridae families. The phage genomes displayed modular organization and conserved viral genes were identified which enabled further classification and determination of closest phage relatives. Co-examination of bacterial host genomes led to the identification of several genes responsible for modulating phage:host interactions, including CRISPR/Cas elements and restriction-modification phage defense systems. These findings provide new genetic information and insights into how lytic phages may interact with bacteria of the rumen microbiome.

In vitro detection and primary cultivation of bacteria producing materials inhibitory to ruminal methanogens.

A novel method for screening bacterial isolates for their potential to inhibit the growth of ruminal methanogenic Archaea was developed using a modification of the soft agar overlay technique, formally used for the isolation of lytic bacteriophages. This method may be used in the specific, hydrogen-rich conditions required for the growth of ruminal methanogenic Archaea.