Researching the Molecular Components of Flourish Using Mass Spectroscopy

We looked into the molecular composition of a liquid probiotic known as Flourish and and the 13 components that make it up using mass spectroscopy

Analysis of a Putative Promoter in Mycobacteriophage JacoRen57

JacoRen57 is a cluster AB mycobacteriophage that infects Mycobacterium smegmatis mc²155. We recently reported on the characterization of a putative promoter in JacoRen57 using an mCherry reporter construct. This promoter is present in a gap upstream of a gene that is present in all AB phages. In all cases, these are forward genes immediately following a long series of reverse genes. The genes are most frequently identified as a RecA-like DNA recombinases but also as RepA by bioinformatics. To further analyze this putative promoter and gene product, NWC Molecular Genetics students cloned the RecA-like DNA recombinase into an E. coli expression vector with a TVMV removable N-terminal His-tag. They expressed and we purified the tagged protein and are using it to immunize Balb/c mice. We plan to use the antiserum to confirm RecA-like DNA recombinase expression patterns when JacoRen57 infects M. smegmatis

Microplastic Concentrations in Orange City Wastewater

Microplastics are increasingly polluting both terrestrial and aquatic ecosystems. Researchers have found that wastewater treatment plants are an entry point for microplastics into surface waters, and we wondered how effective our local wastewater treatment plant is in removing microplastics from wastewater, given that it was not engineered for the removal of microplastics. We sampled effluent water from the new wastewater treatment plant (WWTP) in Orange City, Iowa, to determine the amount of microplastics released from the plant into Orange City Slough. We found concentrations of 498 pieces of microplastic/cubic meter (0.498/L) in the effluent water, which is similar to published values for secondary wastewater treatment plants. Given that microplastics have been found in human blood and have been shown to cause various health effects in humans and other animals, we propose that WWTPs be engineered to prevent the release of microplastics into surface waters

The Effect of Cold Stratification on the Germination of Grassland Seeds

One factor that must be considered when reconstructing a prairie is how the prairie seeds being planted need to be prepared for germination. The probability of successful germination of these seeds is dependent on many factors, including exposure to cold temperatures for a prolonged period of time. To explore this idea, we collected seeds from 13 species of forbs and grasses, both native and non-native, and stored them at various temperatures for several weeks. We predicted that the germination of the seeds of native grassland species would be enhanced by cold stratification, while non-natives (especially forbs common to flower gardens) could be negatively affected by cold stratification, especially if they are native to an area with less-extreme winters. We also predicted that seeds stored at -80°C (a temperature much lower than they would experience in nature) would respond negatively to the treatment and be unable to germinate. We found that four species (Sow Thistle, Penstemon, Queen Anne’s Lace, and Yellow Foxtail) were significantly affected by the seeds being chilled or frozen, with non-native sow thistle and yellow foxtail responding negatively to being frozen and native foxglove penstemon germinating best at -80°C

Invertebrate Pitfall Surveys at Glacial Hills Preserve and Buena Vista County Park

Invertebrates are integral members of the ecosystems they inhabit. However, they are often overlooked and understudied. We performed two pitfall surveys in Buena Vista County in September 2021 to study the diversity and abundance of invertebrate species in the presence and absence of two invasive species: Eastern Redcedar (Juniperus virginiana) and Garlic Mustard (Alliaria petiola). We collected pitfall samples and brought them back to the lab for identification. We found no significant differences in invertebrate abundance between cedar and non-cedar samples, nor between mustard and non-mustard samples. We propose further research focusing on seasonal changes, as these invasives may have more impacts at certain times of year than others

Characterization of Sparus aurata osteonectin cDNA and in silico analysis of protein conserved features: Evidence for more than one osteonectin in Salmonidae

Osteonectin is a matricellular protein involved in various cellular mechanisms but its exact function remains unclear despite numerous studies. We present here the cloning of Sparus aurata partial osteonectin cDNA and the reconstruction of 15 other sequences from both vertebrates and invertebrates, almost doubling the set of available sequences (a total of 35 sequences is now available). Taking advantage of the resulting large amount of data, we have created multiple sequence alignments and identified osteonectin putative conserved features (intra- and inter-disulfide bonds, collagen- and calcium-binding domains and phosphorylation sites) likely to be important for protein structure and function. This work also provides the first evidence for the presence of more than one osteonectin in some species. Finally, S. aurata osteonectin gene expression has been shown to initiate during larval development shortly after gastrulation, and to be high in bone-derived cell lines while down-regulated during extracellular matrix mineralization, further emphasizing the important role of osteonectin in skeletal development and bone formation

Specific MRI abnormalities reveal severe perrault syndrome due to CLPP defects

In establishing a genetic diagnosis in heterogeneous neurological disease, clinical characterization and whole exome sequencing (WES) go hand-in-hand. Clinical data are essential, not only to guide WES variant selection and define the clinical severity of a genetic defect but also to identify other patients with defects in the same gene. In an infant patient with sensorineural hearing loss, psychomotor retardation, and epilepsy, WES resulted in identification of a novel homozygous CLPP frameshift mutation (c.21delA). Based on the gene defect and clinical symptoms, the diagnosis Perrault syndrome type 3 (PRLTS3) was established. The patient's brain-MRI revealed specific abnormalities of the subcortical and deep cerebral white matter and the middle blade of the corpus callosum, which was used to identify similar patients in the Amsterdam brain-MRI database, containing over 3000 unclassified leukoencephalopathy cases. In three unrelated patients with similar MRI abnormalities the CLPP gene was sequenced, and in two of them novel missense mutations were identified together with a large deletion that covered part of the CLPP gene on the other allele. The severe neurological and MRI abnormalities in these young patients were due to the drastic impact of the CLPP mutations, correlating with the variation in clinical manifestations among previously reported patients. Our data show that similarity in brain-MRI patterns can be used to identify novel PRLTS3 patients, especially during early disease stages, when only part of the disease manifestations are present. This seems especially applicable to the severely affected cases in which CLPP function is drastically affected and MRI abnormalities are pronounced

Specific MRI Abnormalities Reveal Severe Perrault Syndrome due to CLPP Defects

In establishing a genetic diagnosis in heterogeneous neurological disease, clinical characterization and whole exome sequencing (WES) go hand-in-hand. Clinical data are essential, not only to guide WES variant selection and define the clinical severity of a genetic defect but also to identify other patients with defects in the same gene. In an infant patient with sensorineural hearing loss, psychomotor retardation, and epilepsy, WES resulted in identification of a novel homozygous CLPP frameshift mutation (c.21delA). Based on the gene defect and clinical symptoms, the diagnosis Perrault syndrome type 3 (PRLTS3) was established. The patient's brain-MRI revealed specific abnormalities of the subcortical and deep cerebral white matter and the middle blade of the corpus callosum, which was used to identify similar patients in the Amsterdam brain-MRI database, containing over 3000 unclassified leukoencephalopathy cases. In three unrelated patients with similar MRI abnormalities the CLPP gene was sequenced, and in two of them novel missense mutations were identified together with a large deletion that covered part of the CLPP gene on the other allele. The severe neurological and MRI abnormalities in these young patients were due to the drastic impact of the CLPP mutations, correlating with the variation in clinical manifestations among previously reported patients. Our data show that similarity in brain-MRI patterns can be used to identify novel PRLTS3 patients, especially during early disease stages, when only part of the disease manifestations are present. This seems especially applicable to the severely affected cases in which CLPP function is drastically affected and MRI abnormalities are pronounce

Trees increase their P:N ratio with size

Aim: Phosphorus (P) tends to become limiting in ageing terrestrial ecosystems, and its resorption efficiency is higher than for other elements such as nitrogen (N). We thus hypothesized that trees should store more P than those other elements such as N when tree size increases and that this process should be enhanced in slow-growing late-successional trees.- Location: Catalan forests. -Methods: We used data from the Catalan Forest Inventory that contains field data on the P and N contents of total aboveground, foliar and woody biomasses of the diverse mediterranean, temperate and alpine forests of Catalonia (1018 sites). We used correlation and general linear models to analyse the allometric relationships between the nutrient contents of different aboveground biomass fractions (foliar, branches and stems) and total aboveground biomass. - Results: Aboveground forest P content increases proportionally more than aboveground forest N content with increasing aboveground biomass. Two mechanisms underlie this. First, woody biomass increases proportionally more than foliar biomass, with woody biomass having a higher P:N ratio than foliar biomass. Second, the P:N ratio of wood increases with tree size. These results are consistent with the generally higher foliar resorption of P than of N. Slow-growing species accumulate more total P aboveground with size than fast-growing species, mainly as a result of their large capacity to store P in wood. - Main conclusions: Trees may have developed long-term adaptive mechanisms to store P in biomass, mainly in wood, thereby slowing the loss of P from ecosystems, reducing its availability for competitors and implying an increase in the P:N ratio in forest biomass with ageing. This trend to accumulate more P than N with size is more accentuated in slow-growing, large, long-lived species of late successional stages. In this way they partly counterbalance the gradual decrease of P in the soil

What's New Is Old: Resolving the Identity of Leptothrix ochracea Using Single Cell Genomics, Pyrosequencing and FISH

Leptothrix ochracea is a common inhabitant of freshwater iron seeps and iron-rich wetlands. Its defining characteristic is copious production of extracellular sheaths encrusted with iron oxyhydroxides. Surprisingly, over 90% of these sheaths are empty, hence, what appears to be an abundant population of iron-oxidizing bacteria, consists of relatively few cells. Because L. ochracea has proven difficult to cultivate, its identification is based solely on habitat preference and morphology. We utilized cultivation-independent techniques to resolve this long-standing enigma. By selecting the actively growing edge of a Leptothrix-containing iron mat, a conventional SSU rRNA gene clone library was obtained that had 29 clones (42% of the total library) related to the Leptothrix/Sphaerotilus group (≤96% identical to cultured representatives). A pyrotagged library of the V4 hypervariable region constructed from the bulk mat showed that 7.2% of the total sequences also belonged to the Leptothrix/Sphaerotilus group. Sorting of individual L. ochracea sheaths, followed by whole genome amplification (WGA) and PCR identified a SSU rRNA sequence that clustered closely with the putative Leptothrix clones and pyrotags. Using these data, a fluorescence in-situ hybridization (FISH) probe, Lepto175, was designed that bound to ensheathed cells. Quantitative use of this probe demonstrated that up to 35% of microbial cells in an actively accreting iron mat were L. ochracea. The SSU rRNA gene of L. ochracea shares 96% homology with its closet cultivated relative, L. cholodnii, This establishes that L. ochracea is indeed related to this group of morphologically similar, filamentous, sheathed microorganisms