A decade of seasonal dynamics and co-occurrences within freshwater bacterioplankton communities from eutrophic Lake Mendota,WI, USA

With an unprecedented decade-long time series from a temperate eutrophic lake, we analyzed bacterial and environmental co-occurrence networks to gain insight into seasonal dynamics at the community level. We found that (1) bacterial co-occurrence networks were non-random, (2) season explained the network complexity and (3) co-occurrence network complexity was negatively correlated with the underlying community diversity across different seasons. Network complexity was not related to the variance of associated environmental factors. Temperature and productivity may drive changes in diversity across seasons in temperate aquatic systems, much as they control diversity across latitude. While the implications of bacterioplankton network structure on ecosystem function are still largely unknown, network analysis, in conjunction with traditional multivariate techniques, continues to increase our understanding of bacterioplankton temporal dynamics.     

Reduction of product-related species during the fermentation and purification of a recombinant IL-1 receptor antagonist at the laboratory and pilot scale

Through a parallel approach of tracking product quality through fermentation and purification development, a robust process was designed to reduce the levels of product-related species. Three biochemically similar product-related species were identified as byproducts of host-cell enzymatic activity. To modulate intracellular proteolytic activity, key fermentation parameters (temperature, pH, trace metals, EDTA levels, and carbon source) were evaluated through bioreactor optimization, while balancing negative effects on growth, productivity, and oxygen demand. The purification process was based on three non-affinity steps and resolved product-related species by exploiting small charge differences. Using statistical design of experiments for elution conditions, a high-resolution cation exchange capture column was optimized for resolution and recovery. Further reduction of product-related species was achieved by evaluating a matrix of conditions for a ceramic hydroxyapatite column. The optimized fermentation process was transferred from the 2-L laboratory scale to the 100-L pilot scale and the purification process was scaled accordingly to process the fermentation harvest. The laboratory- and pilot-scale processes resulted in similar process recoveries of 60 and 65%, respectively, and in a product that was of equal quality and purity to that of small-scale development preparations. The parallel approach for up- and downstream development was paramount in achieving a robust and scalable clinical process.     

The Genome Sequences of Cellulomonas fimi and “Cellvibrio gilvus” Reveal the Cellulolytic Strategies of Two Facultative Anaerobes,Transfer of “Cellvibrio gilvus” to the Genus Cellulomonas,and Proposal of Cellulomonas gilvus sp. nov

Actinobacteria in the genus Cellulomonas are the only known and reported cellulolytic facultative anaerobes. To better understand the cellulolytic strategy employed by these bacteria, we sequenced the genome of the Cellulomonas fimi ATCC 484^T. For comparative purposes, we also sequenced the genome of the aerobic cellulolytic “Cellvibrio gilvus” ATCC 13127^T. An initial analysis of these genomes using phylogenetic and whole-genome comparison revealed that “Cellvibrio gilvus” belongs to the genus Cellulomonas. We thus propose to assign “Cellvibrio gilvus” to the genus Cellulomonas. A comparative genomics analysis between these two Cellulomonas genome sequences and the recently completed genome for Cellulomonas flavigena ATCC 482^T showed that these cellulomonads do not encode cellulosomes but appear to degrade cellulose by secreting multi-domain glycoside hydrolases. Despite the minimal number of carbohydrate-active enzymes encoded by these genomes, as compared to other known cellulolytic organisms, these bacteria were found to be proficient at degrading and utilizing a diverse set of carbohydrates, including crystalline cellulose. Moreover, they also encode for proteins required for the fermentation of hexose and xylose sugars into products such as ethanol. Finally, we found relatively few significant differences between the predicted carbohydrate-active enzymes encoded by these Cellulomonas genomes, in contrast to previous studies reporting differences in physiological approaches for carbohydrate degradation. Our sequencing and analysis of these genomes sheds light onto the mechanism through which these facultative anaerobes degrade cellulose, suggesting that the sequenced cellulomonads use secreted, multidomain enzymes to degrade cellulose in a way that is distinct from known anaerobic cellulolytic strategies.     

Taxonomic Classification of Bacterial 16S rRNA Genes Using Short Sequencing Reads: Evaluation of Effective Study Designs

Massively parallel high throughput sequencing technologies allow us to interrogate the microbial composition of biological samples at unprecedented resolution. The typical approach is to perform high-throughout sequencing of 16S rRNA genes, which are then taxonomically classified based on similarity to known sequences in existing databases. Current technologies cause a predicament though, because although they enable deep coverage of samples, they are limited in the length of sequence they can produce. As a result, high-throughout studies of microbial communities often do not sequence the entire 16S rRNA gene. The challenge is to obtain reliable representation of bacterial communities through taxonomic classification of short 16S rRNA gene sequences. In this study we explored properties of different study designs and developed specific recommendations for effective use of short-read sequencing technologies for the purpose of interrogating bacterial communities, with a focus on classification using naïve Bayesian classifiers. To assess precision and coverage of each design, we used a collection of ∼8,500 manually curated 16S rRNA gene sequences from cultured bacteria and a set of over one million bacterial 16S rRNA gene sequences retrieved from environmental samples, respectively. We also tested different configurations of taxonomic classification approaches using short read sequencing data, and provide recommendations for optimal choice of the relevant parameters. We conclude that with a judicious selection of the sequenced region and the corresponding choice of a suitable training set for taxonomic classification, it is possible to explore bacterial communities at great depth using current technologies, with only a minimal loss of taxonomic resolution.     

Intestinal Methanobrevibacter smithii but not total bacteria is related to diet‐induced weight gain in rats

It is increasingly understood that gastrointestinal (GI) methanogens, including Methanobrevibacter smithii, influence host metabolism.

Objective:

Therefore, we compared M. smithii colonization and weight gain in a rat model under different dietary conditions.

Design and Methods:

Sprague‐Dawley rats were inoculated with M. smithii or vehicle (N = 10/group), fed normal chow until day 112 postinoculation, high‐fat chow until day 182, then normal chow until day 253. Thereafter, five rats from each group were fed high‐fat and normal chow until euthanasia.

Results:

Both groups exhibited M. smithii colonization, which increased following inoculation only for the first 9 days. Change to high‐fat chow correlated with significant increases in weight (P < 0.00001) and stool M. smithii (P < 0.01) in all rats, with stool M. smithi decreasing on return to normal chow. Rats switched back to high‐fat on day 253 further increased weight (P < 0.001) and stool M. smithii (P = 0.039). Euthanasia revealed all animals had higher M. smithii, but not total bacteria, in the small intestine than in the colon. Rats switched back to high‐fat chow had higher M. smithii levels in the duodenum, ileum, and cecum than those fed normal chow; total bacteria did not differ in any bowel segment. Rats which gained more weight had more bowel segments colonized, and the lowest weight recorded was in a rat on high‐fat chow which had minimal M. smithii colonization.

Conclusions:

We conclude that M. smithii colonization occurs in the small bowel as well as in the colon, and that the level and extent of M. smithii colonization is predictive of degree of weight gain in this animal model.     

Modulation of Force below 1 Hz: Age-Associated Differences and the Effect of Magnified Visual Feedback

Oscillations in force output change in specific frequency bins and have important implications for understanding aging and pathological motor control. Although previous studies have demonstrated that oscillations from 0–1 Hz can be influenced by aging and visuomotor processing, these studies have averaged power within this bandwidth and not examined power in specific frequencies below 1 Hz. The purpose was to determine whether a differential modulation of force below 1 Hz contributes to changes in force control related to manipulation of visual feedback and aging. Ten young adults (25±4 yrs, 5 men) and ten older adults (71±5 yrs, 4 men) were instructed to accurately match a target force at 2% of their maximal isometric force for 35 s with abduction of the index finger. Visual feedback was manipulated by changing the visual angle (0.05°, 0.5°, 1.5°) or removing it after 15 s. Modulation of force below 1 Hz was quantified by examining the absolute and normalized power in seven frequency bins. Removal of visual feedback increased normalized power from 0–0.33 Hz and decreased normalized power from 0.66–1.0 Hz. In contrast, magnification of visual feedback (visual angles of 0.5° and 1.5°) decreased normalized power from 0–0.16 Hz and increased normalized power from 0.66–1.0 Hz. Older adults demonstrated a greater increase in the variability of force with magnification of visual feedback compared with young adults (P = 0.05). Furthermore, older adults exhibited differential force modulation of frequencies below 1 Hz compared with young adults (P<0.05). Specifically, older adults exhibited greater normalized power from 0–0.16 Hz and lesser normalized power from 0.66–0.83 Hz. The changes in force modulation predicted the changes in the variability of force with magnification of visual feedback (R² = 0.80). Our findings indicate that force oscillations below 1 Hz are associated with force control and are modified by aging and visual feedback.     

Evaluating environmental DNA‐based quantification of ranavirus infection in wood frog populations

A variety of challenges arise when monitoring wildlife populations for disease. Sampling tissues can be invasive to hosts, and obtaining sufficient sample sizes can be expensive and time‐consuming, particularly for rare species and when pathogen prevalence is low. Environmental DNA (eDNA)‐based detection of pathogens is an alternative approach to surveillance for aquatic communities that circumvents many of these issues. Ranaviruses are emerging pathogens of ectothermic vertebrates linked to die‐offs of amphibian populations. Detecting ranavirus infections is critical, but nonlethal methods have the above issues and are prone to false negatives. We report on the feasibility and effectiveness of eDNA‐based ranavirus detection in the field. We compared ranavirus titres in eDNA samples collected from pond water to titres in wood frog (Lithobates sylvaticus; n = 5) tadpoles in sites dominated by this one species (n = 20 pond visits). We examined whether ranavirus DNA can be detected in eDNA from pond water when infections are present in the pond and if viral titres detected in eDNA samples correlate with the prevalence or intensity of ranavirus infections in tadpoles. With three 250 mL water samples, we were able to detect the virus in all visits with infected larvae (0.92 diagnostic sensitivity). Also, we found a strong relationship between the viral eDNA titres and titres in larval tissues. eDNA titres increased prior to observed die‐offs and declined afterwards, and were two orders of magnitude higher in ponds with a die‐off. Our results suggest that eDNA is useful for detecting ranavirus infections in wildlife and aquaculture.     

Interspecific differences in milkweeds alter predator density and the strength of trophic cascades

Trophic cascades occur when predators benefit plants by consuming herbivores, but the overall strength of a trophic cascade depends upon the way species interactions propagate through a system. For example, plant resistance to, or tolerance of, herbivores reduces the potential magnitude of a trophic cascade. At the same time, plants can also affect predator foraging or consumption in ways that either increase or decrease the strength of trophic cascades. In this study, we investigated the effects of plant variation on cascade strength by manipulating predator access to aphid populations on two species of milkweed: the slower-growing, putatively more-defended Asclepias syriaca and the faster-growing, putatively less-defended Asclepias incarnata. Predatory insects increased plant growth and survival for both species, but the strength of these trophic cascades was greater on A. incarnata, which supported more aphid growth early in the season than did A. syriaca. More predators were observed per aphid on A. incarnata, and cage treatments generated significant patterns consistent with predator aggregation on A. incarnata, but not A. syriaca. Although predators strongly affected aphids, this effect did not differ consistently between milkweed species. Plant tolerance to herbivory may therefore be the primary driver of the difference in trophic cascade strength observed. Importantly, we observed that the timing of predator exclusion affected plant growth and survival differently, indicating that measures of “cascade strength” may change with phenology and plant physiological responses. Together, our results suggest a mechanism by which differences in resource allocation patterns could explain differences in growth, phenology, and cascade strength between species.