Effect of Moniliformis moniliformis density on distribution within the definitive host population (Rattus norvegicus)

The population dynamics of Moniliformis moniliformis was studied in ‘free-ranging’ laboratory rats, Rattus norvegicus, presented with different relative density levels of M. moniliformis in cockroaches, Periplaneta americana. Changes in selected population parameters of the negative binomial distribution were evaluated as indicators of changes in aggregation. A significant increase in the degree of aggregation of parasites occurred as a result of the increase in relative density of infective stages available to the rats. This increase in aggregation was due to the increase in over-dispersion that occurred in female rats only. The degree of aggregation in females was found to be significantly higher than that in males at both treatment levels. The best indicators of the degree of aggregation were found to be the ratio of the variance to the relative density and the ratio of the log-variance to log-relative density. Changes in k were not correlated with changes in over-dispersion or the relative density.     

The controlling sequence for site-specific chromosome breakage in Tetrahymena

Site-specific chromosome breakage occurs in many ciliated protozoa during nuclear differentiation. We have determined the cis-acting sequence that controls this process in Tetrahymena thermophila. The Tetrahymena ribosomal RNA gene is bounded by two breakage sites. Injection of this gene into developing macronuclei leads to breakage at these sites. Deletion analysis has localized the sequences essential for breakage to a 28 bp region that includes a 15 bp sequence (Cbs) known to be present in other breakage sites. Insertions of Cbs allow breakage to occur at new sites, which is accompanied by elimination of surrounding DNAs and formation of telomeric sequences, as it is at natural sites. Thus, Cbs is the necessary and sufficient sequence signal for chromosome breakage in Tetrahymena.     

Characterization of soybean choline kinase cDNAs and their expression in yeast and Escherichia coli.

An expressed sequence tag from Arabidopsis that displayed sequence homology to mammalian and yeast choline kinases was used to isolate choline kinase-like cDNAs from soybean (Glycine max L.). Two distinct cDNAs, designated GmCK1 and GmCK2, were recovered that possessed full-length reading frames, each sharing approximately 32% identity at the predicted amino acid level with the rat choline kinase. A third unique choline kinase-like cDNA, GmCK3, was also identified but was not full length. Heterologous expression of GmCK1 in yeast (Saccharomyces cerevisiae) and GmCK2 in both yeast and Escherichia coli demonstrated that each encodes choline kinase activity. In addition to choline, other potential substrates for the choline kinase enzyme include ethanolamine, monomethylethanolamine (MME), and dimethylethanolamine (DME). Both soybean choline kinase isoforms demonstrated negligible ethanolamine kinase activity. Competitive inhibition assays, however, revealed very distinct differences in their responses to DME and MME. DME effectively inhibited only the GmCK2-encoded choline kinase activity. Although MME failed to effectively inhibit either reaction, an unexpected enhancement of choline kinase activity was observed specifically with the GmCK1-encoded enzyme. These results show that choline kinase is encoded by a small, multigene family in soybean comprising two or more distinct isoforms that exhibit both similarities and differences with regard to substrate specificity.     

Use of 16S rRNA Gene for Identification of a Broad Range of Clinically Relevant Bacterial Pathogens

According to World Health Organization statistics of 2011, infectious diseases remain in the top five causes of mortality worldwide. However, despite sophisticated research tools for microbial detection, rapid and accurate molecular diagnostics for identification of infection in humans have not been extensively adopted. Time-consuming culture-based methods remain to the forefront of clinical microbial detection. The 16S rRNA gene, a molecular marker for identification of bacterial species, is ubiquitous to members of this domain and, thanks to ever-expanding databases of sequence information, a useful tool for bacterial identification. In this study, we assembled an extensive repository of clinical isolates (n = 617), representing 30 medically important pathogenic species and originally identified using traditional culture-based or non-16S molecular methods. This strain repository was used to systematically evaluate the ability of 16S rRNA for species level identification. To enable the most accurate species level classification based on the paucity of sequence data accumulated in public databases, we built a Naïve Bayes classifier representing a diverse set of high-quality sequences from medically important bacterial organisms. We show that for species identification, a model-based approach is superior to an alignment based method. Overall, between 16S gene based and clinical identities, our study shows a genus-level concordance rate of 96% and a species-level concordance rate of 87.5%. We point to multiple cases of probable clinical misidentification with traditional culture based identification across a wide range of gram-negative rods and gram-positive cocci as well as common gram-negative cocci.     

Impacts of soil temperature,phenology and plant community composition on invertebrate herbivory in a natural warming experiment

Species and community-level responses to warming are well documented, with plants and invertebrates known to alter their range, phenology or composition as temperature increases. The effects of warming on biotic interactions are less clearly understood, but can have consequences that cascade through ecological networks. Here, we used a natural soil temperature gradient of 5–35°C in the Hengill geothermal valley, Iceland, to investigate the effects of temperature on plant community composition and plant–invertebrate interactions. We quantified the level of invertebrate herbivory on the plant community across the temperature gradient and the interactive effects of temperature, plant phenology (i.e. development stage) and vegetation community composition on the probability of herbivory for three ubiquitous plant species, Cardamine pratensis, Cerastium fontanum and Viola palustris. We found that the percentage cover of graminoids and forbs increased, while the amount of litter decreased, with increasing soil temperature. Invertebrate herbivory also increased with soil temperature at the plant community level, but this was underpinned by different effects of temperature on herbivory for individual plant species, mediated by the seasonal development of plants and the composition of the surrounding vegetation. This illustrates the importance of considering the development stage of organisms in climate change research given the variable effects of temperature on susceptibility to herbivory at different ontogenetic stages.     

Diagnostics method for the rapid quantitative detection and identification of low-level contamination of high-purity water with pathogenic bacteria

High-purity water (HPW) can be contaminated with pathogenic microorganisms, which may result in human infection. Current culture-based techniques for the detection of microorganisms from HPW can be slow and laborious. The aim of this study was to develop a rapid method for the quantitative detection and identification of pathogenic bacteria causing low-level contamination of HPW. A novel internally controlled multiplex real-time PCR diagnostics assay was designed and optimized to specifically detect and identify Pseudomonas aeruginosa and the Burkholderia genus. Sterile HPW, spiked with a bacterial load ranging from 10 to 10³ cfu/100 ml, was filtered and the bacterial cells were removed from the filters by sonication. Total genomic DNA was then purified from these bacteria and subjected to testing with the developed novel multiplex real-time PCR diagnostics assay. The specific P. aeruginosa and Burkholderia genus assays have an analytical sensitivity of 3.5 genome equivalents (GE) and 3.7 GE, respectively. This analysis demonstrated that it was possible to detect a spiked bacterial load of 1.06 × 10² cfu/100 ml for P. aeruginosa and 2.66 × 10² cfu/100 ml for B. cepacia from a 200-ml filtered HPW sample. The rapid diagnostics method described can reliably detect, identify, and quantify low-level contamination of HPW with P. aeruginosa and the Burkholderia genus in <4 h. We propose that this rapid diagnostics method could be applied to the pharmaceutical and clinical sectors to assure the safety and quality of HPW, medical devices, and patient-care equipment.     

Survey of Innate Immune Responses to Burkholderia pseudomallei in Human Blood Identifies a Central Role for Lipopolysaccharide

B. pseudomallei is a gram-negative bacterium that causes the tropical infection melioidosis. In northeast Thailand, mortality from melioidosis approaches 40%. As exemplified by the lipopolysaccharide-Toll-like receptor 4 interaction, innate immune responses to invading bacteria are precipitated by activation of host pathogen recognition receptors by pathogen associated molecular patterns. Human melioidosis is characterized by up-regulation of pathogen recognition receptors and pro-inflammatory cytokine release. In contrast to many gram-negative pathogens, however, the lipopolysaccharide of B. pseudomallei is considered only weakly inflammatory. We conducted a study in 300 healthy Thai subjects to investigate the ex vivo human blood response to various bacterial pathogen associated molecular patterns, including lipopolysaccharide from several bacteria, and to two heat-killed B. pseudomallei isolates. We measured cytokine levels after stimulation of fresh whole blood with a panel of stimuli. We found that age, sex, and white blood cell count modulate the innate immune response to B. pseudomallei. We further observed that, in comparison to other stimuli, the innate immune response to B. pseudomallei is most highly correlated with the response to lipopolysaccharide. The magnitude of cytokine responses induced by B. pseudomallei lipopolysaccharide was significantly greater than those induced by lipopolysaccharide from Escherichia coli and comparable to many responses induced by lipopolysaccharide from Salmonella minnesota despite lower amounts of lipid A in the B. pseudomallei lipopolysaccharide preparation. In human monocytes stimulated with B. pseudomallei, addition of polymyxin B or a TLR4/MD-2 neutralizing antibody inhibited the majority of TNF-α production. Challenging existing views, our data indicate that the innate immune response to B. pseudomallei in human blood is largely driven by lipopolysaccharide, and that the response to B. pseudomallei lipopolysaccharide in blood is greater than the response to other lipopolysaccharide expressing isolates. Our findings suggest that B. pseudomallei lipopolysaccharide may play a central role in stimulating the host response in melioidosis.     

Androgen receptor expression in satellite cells of the neonatal levator ani of the rat

Androgens are thought to mediate sexual differentiation of spinal nucleus of the bulbocavernosus (SNB) motoneurons via actions on androgen receptors (ARs) within their target muscles bulbocavernosus and levator ani (LA). However, the cells within these muscles which mediate masculinization of the SNB remain undefined. Until recently, myocytes were thought to be the most likely candidate cell type. However, genetic tests of AR function in myocytes have failed to support a sufficient role for these cells in producing masculine SNB morphology, suggesting the involvement of other cell types. To identify other candidate cell types in the LA, we evaluated whether satellite cells or fibroblasts express AR. Fluorescent immunohistochemistry and confocal microscopy were used to evaluate whether satellite cells and fibroblasts express AR in neonatal male and female rats in the LA and an adjacent sexually monomorphic control muscle (CM). We found that a small proportion of satellite cells in the LA express AR and that this proportion is significantly greater in the LA compared to the CM. No sex differences were found between the proportions of satellite cells expressing AR in either muscle. Less colocalization of satellite cells and AR was seen in postnatal day 3 muscle than in postnatal day 1 muscle. In contrast, only negligible amounts of fibroblasts labeled with S100A4 express AR in either the LA or the CM. Together, findings support satellite cells, but not fibroblasts, as a candidate cell type involved in the sexual differentiation of the SNB neuromuscular system. © 2012 Wiley Periodicals, Inc. Develop Neurobiol 73: 448–454, 2013.     

Wetland Waterbird Food Resources Increased by Harvesting Invasive Cattails

The conservation of many freshwater marsh waterbirds (i.e., waterfowl, shorebirds, wading birds, and secretive marshbirds) in the Laurentian Great Lakes requires managing invasive emergent macrophytes, which degrade waterbird habitat by creating dense, litter-clogged stands, and excluding plants that produce nutritionally balanced and high-energy food (seeds, tubers, and submerged aquatic vegetation). The most commonly used management approach in the United States Great Lakes region involves the application of herbicides, which can stimulate waterbird forage plants but does not address the accumulation of plant litter, the underlying cause of plant community diversity loss and habitat degradation. We experimentally evaluated the effects of an alternative approach, harvesting invasive plants and their litter followed by flooding, on plant communities, focusing on the effects of these treatments to increase the abundance of high-energy wetland plants. At the Shiawassee National Wildlife Refuge in Michigan, USA, we experimentally treated an invasive cattail (Typha × glauca)-dominated wetland in August and September of 2016, 2017, and 2018, using a randomized block design with 4 blocks and 3 treatments (sediment surface harvest, above ground harvest, and control). We monitored the effects of these treatments on the abundance and dominance of waterbird forage-producing plants, plant diversity, and plant communities prior to (Jul 2016) and during the summer following each treatment (late Jul or early Aug 2017, 2018, and 2019). Additionally, we used pre- and post-treatment waterbird use-day data collected at the unit scale and compared values with satellite imagery-derived land cover changes. Compared to control plots, 3 years of harvesting and flooding significantly increased plant species diversity, increased the abundance of waterbird seed- and tuber-producing plant species by 5 times, and increased annual plant dominance by more than 10 times, while substantially reducing all measures of cattail and its litter. Use-days increased for total waterbirds, including waterfowl and dabbling ducks, following treatment. Cattail cover decreased and open water and non-cattail emergent vegetation cover increased. Harvesting invasive plant biomass coupled with flooding promoted a plant community composition and structure beneficial to waterbirds. © 2020 The Wildlife Society.